Transcriptional enhancers and their communication with gene promoters

In the paper

“The mechanisms of pioneer factor action are still being established [37], but at least some of them (such as FOXA, OCT4 and SOX2/SOX11) have been shown to initiate chromatin opening by direct displacement of histones [38–41], whilst many others recruit chromatin remodelers [42–50].”

What the sources say

• Supported

  The Results section (“FoxA binding displaces linker histone H1 from nucleosomes”; page unavailable) reports that FoxA binding displaces linker histone H1, making underlying nucleosomes more accessible. This supports the FOXA example for which this source is cited; the paper does not establish the corresponding mechanism for OCT4 or SOX2/SOX11. (LLM, conf=0.90)

  cites [38] Iwafuchi-Doi M, Donahue G, Kakumanu A et al (2016) The pioneer transcription factor FoxA maintains an accessible nucleosome configuration at enhancers for tissue-specific gene activation. Mol Cell 62:79–91 source
• Disputed

  The Introduction says OCT4 may help convert closed chromatin to open chromatin, but it does not attribute that opening to direct histone displacement. The Results section “Linker histone H1 competes with OCT4 for nucleosome binding” reports the opposite of the claimed mechanism for OCT4: adding OCT4 did not displace H1; adding H1 removed OCT4 from the nucleosome. The Discussion describes SOX2/SOX11 peeling DNA from histones, not displacing the histones. (LLM, conf=0.88)

  Checked by a reasoning model — may be wrong.

  cites [39] Echigoya K, Koyama M, Negishi L et al (2020) Nucleosome binding by the pioneer transcription factor OCT4. Sci Rep 10:11832 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [40] Dodonova SO, Zhu F, Dienemann C et al (2020) Nucleosome-bound SOX2 and SOX11 structures elucidate pioneer factor function. Nature 580:669–672 source
• Partly supported

  The introductory passage (labelled Abstract in our copy; page unavailable) describes pioneer factors as transcription factors that can displace nucleosomes and names FOXA1 and SOX2 as examples. The Results section, “Functional Grainyhead motifs are embedded in a specific DNA context” (page unavailable), also associates pioneer-factor binding with nucleosome displacement. But the paper does not establish direct histone displacement for each named factor: it mentions OCT4 as a possible additional factor needed for SOX2-dependent enhancer activation and does not mention SOX11. (LLM, conf=0.84)

  cites [41] Jacobs J, Atkins M, Davie K et al (2018) The transcription factor Grainy head primes epithelial enhancers for spatiotemporal activation by displacing nucleosomes. Nat Genet 50:1011–1020 sourceExperimental — accuracy not yet measured

In the paper

“The majority of active enhancers are themselves transcribed, producing enhancer RNAs (eRNAs) in a bi-directional manner [73–77].”

What the sources say

• Partly supported

  The Abstract says enhancer transcription is widespread and that eRNAs are produced bidirectionally. The Discussion also reports bidirectional initiation at thousands of enhancers. Neither passage establishes that a majority of active enhancers are transcribed; the nearly 80% figure in the Abstract concerns promoters, not enhancers. (LLM, conf=0.85)

  cites [73] Core LJ, Martins AL, Danko CG et al (2014) Analysis of nascent RNA identifies a unified architecture of initiation regions at mammalian promoters and enhancers. Nat Genet 46:1311–1320 sourceExperimental — accuracy not yet measured
• Supported

  The Results section ('Bidirectional pairs of capped RNAs identify active enhancers'; page not provided) describes similar levels of capped RNA in both directions at enhancers and calls bidirectional transcription a signature feature of active enhancers. The Conclusions add that few untranscribed candidate enhancers showed activity, supporting the claim's broad characterization of active enhancers. (LLM, conf=0.87)

  cites [74] Andersson R, Gebhard C, Miguel-Escalada I et al (2014) An atlas of active enhancers across human cell types and tissues. Nature 507:455–461 source
• Partly supported

  The passage reporting GRO-Seq results (section labeled Abstract; page unavailable) found eRNA initiation at 76% of Rev-Erb binding sites in enhancer-like regions and bidirectional transcription at 56% of those sites. This supports the pattern in the studied macrophage-associated sites, but not the broader claim that a majority of active enhancers generally are transcribed bidirectionally. (LLM, conf=0.86)

  cites [75] Lam MTY, Cho H, Lesch HP et al (2013) Rev-Erbs repress macrophage gene expression by inhibiting enhancer-directed transcription. Nature 498:511–515 sourceExperimental — accuracy not yet measured
• Disputed

  The Discussion says that non-transcribed enhancers greatly outnumber transcribed ones, contrary to the claim that a majority are transcribed. The Introduction also gives an example of unidirectional enhancer-region transcription; this paper does not establish bidirectional production as the general pattern. No page numbers are available in our copy. (LLM, conf=0.90)

  Checked by a reasoning model — may be wrong.

  cites [76] De Santa F, Barozzi I, Mietton F et al (2010) A large fraction of extragenic RNA pol II transcription sites overlap enhancers. PLoS Biol 8:e1000384 sourceExperimental — accuracy not yet measured
• Partly supported

  The Transcription at enhancers section reports that eRNAs are produced bi-directionally at neuronal activity-regulated enhancers. It does not support the claim that a majority of active enhancers are transcribed: the Methods Summary reports detectable eRNAs at 44% of extragenic enhancers, and the findings are limited to neuronal enhancers. Page numbers are not provided in our copy. (LLM, conf=0.89)

  cites [77] Kim T-K, Hemberg M, Gray JM et al (2010) Widespread transcription at neuronal activity-regulated enhancers. Nature 465:182–187 sourceExperimental — accuracy not yet measured

In the paper

“Nonetheless, a significant minority of enhancers in human ESCs also show pre-existing “permissive” promoter contacts prior to activation [141, 394].”

What the sources say

• Supported

  The Discussion (Reconfiguration of cis-regulatory units upon lineage commitment; page unavailable) defines permissive interactions as contacts present before gene activation and says the study found such preformed interactions during ESC differentiation. The Results (Rewiring and recolouring of promoter-interacting regions; page unavailable) report that 25% of promoter-interacting regions retained across ESCs and NECs changed chromatin state, supporting the claim’s qualitative description of a substantial minority. (LLM, conf=0.86)

  cites [141] Freire-Pritchett P, Schoenfelder S, Várnai C et al (2017) Global reorganisation of regulatory units upon lineage commitment of human embryonic stem cells. Elife 6:e21926 source
• Disputed

  The Abstract says the pre-established enhancer–promoter contacts studied were present in epidermal progenitor cells but not in embryonic stem cells. Its analysis of hESC promoter-contact data likewise reports reduced contact signal relative to keratinocytes. The paper places these pre-established contacts after pluripotency, not in hESCs. (LLM, conf=0.88)

  Checked by a reasoning model — may be wrong.

  cites [394] Rubin AJ, Barajas BC, Furlan-Magaril M et al (2017) Lineage-specific dynamic and pre-established enhancer–promoter contacts cooperate in terminal differentiation. Nat Genet 49:1522–1528 sourceExperimental — accuracy not yet measured

In the paper

“In the classic scenario, this boundary is presented by CTCF binding to its sequence motifs located in a divergent orientation at either end of the loop [277, 438–440]”

What the sources say

• Disputed

  The Results section reports that 92% of CTCF motif pairs at the examined loop loci are convergent, not divergent, and that convergent pairs are overwhelmingly more frequent than divergent pairs. The Discussion likewise says that motifs at loop anchors are convergent in more than 90% of cases. Page numbers were not supplied in our copy. (LLM, conf=0.95)

  Checked by a reasoning model — may be wrong.

  cites [277] Rao SSP, Huntley MH, Durand NC et al (2014) A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping. Cell 159:1665–1680 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [438] Guo Y, Monahan K, Wu H et al (2012) CTCF/cohesin-mediated DNA looping is required for protocadherin α promoter choice. Proc Natl Acad Sci USA 109:21081–21086 source
• Partly supported

  In Results, under “Directional CTCF Binding in Genome-wide DNA Looping Specificity” (page unavailable), the paper reports reverse-forward CTCF-binding-site pairs at boundaries between neighboring domains, consistent with divergent sites at a boundary. But it reports that most CTCF-mediated interactions within loops involve forward-reverse, convergent site pairs. It therefore does not support describing the sites at either end of a loop as divergent. (LLM, conf=0.87)

  cites [439] Guo Y, Xu Q, Canzio D et al (2015) CRISPR inversion of CTCF sites alters genome topology and enhancer/promoter function. Cell 162:900–910 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results section reports that divergent CTCF-site patterns are enriched at TAD boundaries and suggests they confer boundary function (page not provided). But the paper distinguishes boundaries from loop ends: its Results section says loop bases involve pairs of convergent CTCF sites. It therefore does not support placing divergent motifs at either end of a loop. (LLM, conf=0.87)

  cites [440] Nanni L, Ceri S, Logie C (2020) Spatial patterns of CTCF sites define the anatomy of TADs and their boundaries. Genome Biol 21:197 sourceExperimental — accuracy not yet measured

In the paper

“In Drosophila, the Zinc Finger Early Drosophila Activator (Zelda) is critical for lowering the nucleosome barrier at enhancers for genes driving zygotic gene activation [33–35].”

What the sources say

• Partly supported

  The paper’s Abstract reports that Zelda binds regulatory TAGteam sites and plays a key role in activating the early zygotic genome. It does not say that Zelda lowers a nucleosome barrier at enhancers; the passages retrieved for nucleosome occupancy and chromatin accessibility do not establish that mechanism. No page numbers are available in our copy. (LLM, conf=0.88)

  cites [33] Liang H-L, Nien C-Y, Liu H-Y et al (2008) The zinc-finger protein Zelda is a key activator of the early zygotic genome in Drosophila. Nature 456:400–403 sourceExperimental — accuracy not yet measured
• Partly supported

  The Discussion (page unavailable) says Zelda is important at promoters and enhancers and proposes that it promotes zygotic genome activation by controlling chromatin accessibility. But it does not establish that Zelda lowers a nucleosome barrier; it explicitly allows that Zelda may maintain chromatin that is already accessible rather than actively open it. (LLM, conf=0.88)

  cites [34] Harrison MM, Li X-Y, Kaplan T et al (2011) Zelda binding in the early Drosophila melanogaster embryo marks regions subsequently activated at the maternal-to-zygotic transition. PLoS Genet 7:e1002266 sourceExperimental — accuracy not yet measured
• Supported

  The Discussion (page unavailable), including Figure 7’s caption, says Zelda determines chromatin accessibility at early embryonic enhancers, allowing other transcription factors to bind. It also proposes that Zelda is essential for creating open chromatin that drives expression of the earliest zygotic genes. The paper limits this role to a subset of regions, rather than all Zelda-bound enhancers. (LLM, conf=0.87)

  cites [35] Schulz KN, Bondra ER, Moshe A et al (2015) Zelda is differentially required for chromatin accessibility, transcription factor binding, and gene expression in the early Drosophila embryo. Genome Res 25:1715–1726 source

In the paper

“Mechanistically, a key function of sequence-specific TFs is the recruitment of “workhorse” cofactors that facilitate enzymatic chromatin remodelling, histone modification and act as scaffolds for recruitment of additional factors.”

What the sources say

• Partly supported

  The section on differential activity of trans factors (page unavailable) says sequence-specific TFs recruit cofactors, including the SWI/SNF complex and p300, and identifies Mof as an acetyltransferase. The section on activating micro-environments describes cofactor activity leading to recruitment of BRD4, but does not establish the claimed scaffold role for recruiting additional factors. (LLM, conf=0.83)

  cites [56] Zabidi MA, Stark A (2016) Regulatory enhancer-core-promoter communication via transcription factors and cofactors. Trends Genet 32:801–814 sourceExperimental — accuracy not yet measured
• Partly supported

  The introduction and “FACT Function and Architecture” section describe FACT as a histone chaperone that reorganizes nucleosomes during transcription. The “Modifying” FACT Function section says histone modifications can recruit chromatin-altering machinery. The copy does not establish the claim’s central mechanism—that sequence-specific transcription factors recruit cofactors—or describe those cofactors as scaffolds for additional factors. (LLM, conf=0.78)

  cites [57] Winkler DD, Luger K (2011) The histone chaperone FACT: structural insights and mechanisms for nucleosome reorganization. J Biol Chem 286:18369–18374 sourceExperimental — accuracy not yet measured

In the paper

“P300 and CBP also serve as scaffolds that connect RNA polymerase II (RNAP) and RNAP-associated “general transcription factors” to the chromatin [64, 65].”

What the sources say

• ?Not checked — no full text of the sourcecites [64] Chan HM, La Thangue NB (2001) p300/CBP proteins: HATs for transcriptional bridges and scaffolds. J Cell Sci 114:2363–2373 source
• Partly supported

  The Results and Discussion (page unavailable) show that interactions between CBP and the IFN-β enhanceosome help recruit an RNA polymerase II holoenzyme containing general transcription factors to a promoter. The paper does not establish the same role for p300 or show that either protein acts as a scaffold connecting those factors to chromatin; its experiments used an immobilized DNA template in vitro. (LLM, conf=0.85)

  cites [65] Kim TK, Kim TH, Maniatis T (1998) Efficient recruitment of TFIIB and CBP-RNA polymerase II holoenzyme by an interferon-beta enhanceosome in vitro. Proc Natl Acad Sci USA 95:12191–12196 sourceExperimental — accuracy not yet measured

In the paper

“Recent experiments degrading Mediator and BRD4 have shown that these proteins are critical for gene expression [69].”

What the sources say

• Partly supported

  The Discussion (page unavailable) says BRD4 and Mediator play a key role in the transcription of many genes. The Introduction describes this paper’s use of a BRD4-degrading compound, but its experiments reduced Mediator binding rather than degrading Mediator; it mentions MED14 degradation only in a separate study of enhancer–promoter contact. Thus the gene-expression conclusion is supported, but the description of the experiments is not fully supported. (LLM, conf=0.86)

  cites [69] Crump NT, Ballabio E, Godfrey L et al (2021) BET inhibition disrupts transcription but retains enhancer–promoter contact. Nat Commun 12:223 sourceExperimental — accuracy not yet measured

In the paper

“It soon became clear that enhancers also played vital roles in humans, with non-coding enhancer deletions starting to be linked to severe disease phenotypes such as beta thalassaemia and polydactyly [25–28].”

What the sources say

• ?Not checked — no full text of the sourcecites [25] Kioussis D, Vanin E, deLange T et al (1983) Beta-globin gene inactivation by DNA translocation in gamma beta-thalassaemia. Nature 306:662–666 source
• Supported

  The Discussion reports a patient with γδβ-thalassemia whose de novo deletion removed three upstream regulatory hypersensitive sites while leaving the structural globin genes intact. This supports the beta-thalassemia example attributed to this paper; the sentence’s other citations may address polydactyly. (LLM, conf=0.92)

  cites [26] Driscoll MC, Dobkin CS, Alter BP (1989) Gamma delta beta-thalassemia due to a de novo mutation deleting the 5’ beta-globin gene activation-region hypersensitive sites. Proc Natl Acad Sci USA 86:7470–7474 source
• Partly supported

  The “ZRS and polydactyly” section links mutations in a non-coding regulatory element to preaxial polydactyly. It specifies single-base-pair mutations, not enhancer deletions, and does not address beta thalassaemia. Page numbers are unavailable in our copy. (LLM, conf=0.88)

  cites [27] Lettice LA, Heaney SJH, Purdie LA et al (2003) A long-range Shh enhancer regulates expression in the developing limb and fin and is associated with preaxial polydactyly. Hum Mol Genet 12:1725–1735 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [28] Sharpe J, Lettice L, Hecksher-Sørensen J et al (1999) Identification of Sonic hedgehog as a candidate gene responsible for the polydactylous mouse mutant Sasquatch. Curr Biol 9:97-S1 source

In the paper

“Often, pioneer factors must recruit additional non-pioneer sequence-specific TFs for efficient enhancer activation [30, 31].”

What the sources say

• ?Not checked — no full text of the sourcecites [30] Spitz F, Furlong EEM (2012) Transcription factors: from enhancer binding to developmental control. Nat Rev Genet 13:613–626 source
• Supported

  In “Active and passive means by which pioneer factors are useful in development,” the paper says that the vast majority of promoters and enhancers require a combination of transcription factors for activity. It explains that pioneer factors can bind first, allowing other transcription factors to bind later and contribute to activation. This supports the claim’s substance, although “recruit” is a loose paraphrase of that process. (LLM, conf=0.88)

  cites [31] Zaret KS, Carroll JS (2011) Pioneer transcription factors: establishing competence for gene expression. Genes Dev 25:2227–2241 source

In the paper

“At the DNA level, enhancers represent clusters of binding sites for sequence-specific transcription factors (TFs) (Fig. 1A) (reviewed in [29, 30]).”

What the sources say

• Supported

  The Introduction section “Enhancers as clusters of TF binding sites” states that functional enhancers are composed of concentrated clusters of transcription-factor recognition motifs. This directly supports the citing sentence’s description of enhancers at the DNA level. (LLM, conf=0.95)

  cites [29] Long HK, Prescott SL, Wysocka J (2016) Ever-changing landscapes: transcriptional enhancers in development and evolution. Cell 167:1170–1187 source
• ?Not checked — no full text of the sourcecites [30] Spitz F, Furlong EEM (2012) Transcription factors: from enhancer binding to developmental control. Nat Rev Genet 13:613–626 source

In the paper

“For instance, Zelda promotes the formation of local TF hubs involving important transcription factors required for developmental patterning, such as Bicoid and Dorsal [51–54].”

What the sources say

• Supported

  The Results section, “Formation of BCD hubs in the posterior embryo is dependent on ZLD” (page unavailable), reports that Bicoid hubs were abolished in Zelda-null embryos. This supports the Bicoid example for which this paper is offered among the grouped citations. (LLM, conf=0.89)

  cites [51] Mir M, Reimer A, Haines JE et al (2017) Dense Bicoid hubs accentuate binding along the morphogen gradient. Genes Dev 31:1784–1794 source
• Partly supported

  The Results section ('Bicoid binding events are enriched in Zelda hubs'; page unavailable) reports Bicoid binding enriched within Zelda hubs, supporting that example. The cited paper does not establish Dorsal’s involvement in those hubs, so the Dorsal example is unsupported by this source. (LLM, conf=0.88)

  cites [52] Mir M, Stadler MR, Ortiz SA et al (2018) Dynamic multifactor hubs interact transiently with sites of active transcription in embryos. Elife 7:e40497 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results and Discussion section (‘Zelda accumulates into subnuclear hubs’; page unavailable) reports local, dynamic Zelda hubs and suggests they could promote cooperation with Bicoid. The Introduction and ‘Zelda fosters temporal transcriptional coordination’ section say Zelda potentiates Bicoid and Dorsal activity, but the paper does not establish that either factor participates in the observed Zelda hubs or that Zelda promotes formation of hubs containing them. (LLM, conf=0.87)

  cites [53] Dufourt J, Trullo A, Hunter J et al (2018) Temporal control of gene expression by the pioneer factor Zelda through transient interactions in hubs. Nat Commun 9:5194 sourceExperimental — accuracy not yet measured
• Supported

  The Abstract reports increased Dorsal accumulation at the transcription site and describes transcription-factor hubs induced by Zelda. The Discussion reports Bicoid enrichment in Zelda hubs and suggests that Zelda raises transcription factors’ local concentration. Page numbers are not provided in our copy. (LLM, conf=0.90)

  cites [54] Yamada S, Whitney PH, Huang S-K et al (2019) The drosophila pioneer factor zelda modulates the nuclear microenvironment of a dorsal target enhancer to potentiate transcriptional output. Curr Biol 29:1387-1393.e5 source

In the paper

“Subsequent landmark experiments by the groups of Walter Schaffner [4] and Pierre Chambon [5] confirmed that a 72-bp repeat sequence element in SV40 [8, 9] was an “enhancing” sequence, capable of vastly upregulating gene expression from a plasmid upon transfection into mammalian cells.”

What the sources say

• ?Not checked — no full text of the sourcecites [4] Banerji J, Rusconi S, Schaffner W (1981) Expression of a beta-globin gene is enhanced by remote SV40 DNA sequences. Cell 27:299–308 source
• ?Not checked — no full text of the sourcecites [5] Moreau P, Hen R, Wasylyk B et al (1981) The SV40 72 base repair repeat has a striking effect on gene expression both in SV40 and other chimeric recombinants. Nucleic Acids Res 9:6047–6068. https://doi.org/10.1093/nar/9.22.6047 source
• ?Not checked — no full text of the sourcecites [8] Benoist C, Chambon P (1981) In vivo sequence requirements of the SV40 early promoter region. Nature 290:304–310 source
• Supported

  The citation to this paper is attached to the identification of the 72-bp repeat element in SV40; the sentence attributes the plasmid-enhancement experiments to other citations. The paper’s abstract identifies the 72-base-pair tandem repeats and reports that they form an essential element of the early viral transcriptional promoter. (LLM, conf=0.91)

  cites [9] Gruss P, Dhar R, Khoury G (1981) Simian virus 40 tandem repeated sequences as an element of the early promoter. Proc Natl Acad Sci USA 78:943–947 source

In the paper

“A subset of TFs known as pioneer factors are capable of overcoming this constraint through chromatin remodelling (reviewed in [31, 32]).”

What the sources say

• Supported

  The section “Active and passive means by which pioneer factors are useful in development” says pioneer factors can bind highly compacted chromatin and open or organize it locally, allowing other transcription factors to bind. The summary clarifies that this access applies to certain forms of compacted chromatin, not all forms. (LLM, conf=0.92)

  cites [31] Zaret KS, Carroll JS (2011) Pioneer transcription factors: establishing competence for gene expression. Genes Dev 25:2227–2241 source
• Supported

  The Abstract describes pioneer factors as a specialized class of transcription factors that bind histone-wrapped DNA and establish accessible chromatin domains. It also lists mechanisms by which they increase accessibility, including histone eviction and recruitment of chromatin-modifying enzymes. The Conclusions likewise describe interactions with chromatin-remodeling enzymes. Page numbers are not provided in our copy. (LLM, conf=0.93)

  cites [32] Larson ED, Marsh AJ, Harrison MM (2021) Pioneering the developmental frontier. Mol Cell 81:1640–1650 source

In the paper

“At active enhancers, the BRD4 protein co-localises with the Mediator complex, which in turn assembles the pre-initiation complex (PIC) and RNAP to initiate transcription [66–68].”

What the sources say

• Partly supported

  The “Mediator roles in pol II transcription” and “PIC assembly” sections say that Mediator is generally required for PIC assembly and enables RNA polymerase II recruitment. The “DNA looping” section links Mediator to enhancer activity, but the passages retrieved for BRD4 do not establish that BRD4 co-localises with Mediator at active enhancers. Page numbers are not available in our copy. (LLM, conf=0.88)

  cites [66] Allen BL, Taatjes DJ (2015) The mediator complex: a central integrator of transcription. Nat Rev Mol Cell Biol 16:155–166 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [67] Flanagan PM, Kelleher RJ, Sayre MH et al (1991) A mediator required for activation of RNA polymerase II transcription in vitro. Nature 350:436–438 source
• ?Not checked — no full text of the sourcecites [68] Kelleher RJ, Flanagan PM, Kornberg RD (1990) A novel mediator between activator proteins and the RNA polymerase II transcription apparatus. Cell 61:1209–1215 source

In the paper

“SWI/SNF recruits the chromatin activators P300 and CBP, which mediate the acetylation of histone H3 lysine 27 through their intrinsic histone acetyltransferase (HAT) activity [59–62].”

What the sources say

• ?Not checked — no full text of the sourcecites [59] Chan HM, Shikama N, La Thangue NB (2001) Control of gene expression and the cell cycle. Essays Biochem 37:87–96 source
• Partly supported

  The Introduction identifies CBP and p300 as histone acetyltransferase co-activators, and “Substrate and site specificities of CBP/p300” reports that they are required for over 90% of H3K27 acetylation in mouse embryonic fibroblasts. The cited paper does not establish the sentence’s additional claim that SWI/SNF recruits CBP and p300. (LLM, conf=0.88)

  cites [60] Jin Q, Yu L-R, Wang L et al (2011) Distinct roles of GCN5/PCAF-mediated H3K9ac and CBP/p300-mediated H3K18/27ac in nuclear receptor transactivation. EMBO J 30:249–262 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [61] Raisner R, Kharbanda S, Jin L et al (2018) Enhancer activity requires CBP/P300 bromodomain-dependent histone H3K27 acetylation. Cell Rep 24:1722–1729 source
• Partly supported

  The Results section ('SWI/SNF is required for p300 activity at enhancers'; page not provided) says SWI/SNF recruits p300 to enhancer loci and regulates H3K27 acetylation through interaction with this histone acetyltransferase. It does not establish that SWI/SNF also recruits CBP, so the claim's inclusion of CBP is unsupported by the cited Results passages. (LLM, conf=0.85)

  cites [62] Alver BH, Kim KH, Lu P et al (2017) The SWI/SNF chromatin remodelling complex is required for maintenance of lineage specific enhancers. Nat Commun 8:14648 sourceExperimental — accuracy not yet measured

In the paper

“Synthetic enhancers recruiting multiple heterotypic TFs showed stronger transcriptional effects compared with those recruiting high amounts of a single TF, providing a functional rationale for combinatorial TF recruitment to natural enhancers [55].”

What the sources say

• Supported

  The Results section, “Increased binding site complexity leads to stronger expression,” reports that heterotypic synthetic regulatory elements generally drove stronger expression than homotypic elements, even when the number of binding sites was controlled. The paper tested binding-site arrangements rather than directly measuring TF recruitment, but the claim accurately paraphrases the reported comparison. (LLM, conf=0.93)

  cites [55] Smith RP, Taher L, Patwardhan RP et al (2013) Massively parallel decoding of mammalian regulatory sequences supports a flexible organizational model. Nat Genet 45:1021–1028 source

In the paper

“The mechanisms of pioneer factor action are still being established [37], but at least some of them (such as FOXA, OCT4 and SOX2/SOX11) have been shown to initiate chromatin opening by direct displacement of histones [38–41], whilst many others recruit chromatin remodelers [42–50].”

What the sources say

• Partly supported

  The Results and Discussion section (page not available) reports that Oct4 physically interacts with components of several chromatin-remodeling and other epigenetic complexes. It does not establish that many pioneer factors recruit remodelers, or that Oct4’s interactions constitute recruitment to open chromatin. (LLM, conf=0.84)

  cites [42] Ding J, Xu H, Faiola F et al (2012) Oct4 links multiple epigenetic pathways to the pluripotency network. Cell Res 22:155–167 sourceExperimental — accuracy not yet measured
• Partly supported

  The Discussion (page unavailable) reports that Oct4 copurifies with subunits of the SWI/SNF nucleosome-remodeling complex, supporting an Oct4–remodeler association. It does not show that Oct4 recruits the complex, or establish that many pioneer factors recruit chromatin remodelers. (LLM, conf=0.84)

  cites [43] Pardo M, Lang B, Yu L et al (2010) An expanded Oct4 interaction network: implications for stem cell biology, development, and disease. Cell Stem Cell 6:382–395 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results (An Oct4-Centered Interaction Network; page not provided) report that several embryonic-stem-cell transcription factors associate with chromatin-modifying complexes, including SWI/SNF. The Discussion (Multiple Network Connections with Chromatin- and Protein-Modifying Factors; page not provided) proposes that recruitment may occur through initial local targeting, but does not establish that many pioneer factors recruit remodelers. The claim presents that proposed mechanism as a demonstrated, broader finding. (LLM, conf=0.86)

  cites [44] van den Berg DLC, Snoek T, Mullin NP et al (2010) An Oct4-centered protein interaction network in embryonic stem cells. Cell Stem Cell 6:369–381 sourceExperimental — accuracy not yet measured
• Partly supported

  The Discussion (page unavailable) reports that OCT4 recruits BRG1 to its binding sites and suggests that many pioneer factors may rely on chromatin remodelling. It does not establish that many other pioneer factors recruit remodelers; it presents that broader mechanism as a possibility requiring further study. (LLM, conf=0.83)

  cites [45] King HW, Klose RJ (2017) The pioneer factor OCT4 requires the chromatin remodeller BRG1 to support gene regulatory element function in mouse embryonic stem cells. Elife 6:e22631 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results section (“BAF and p63 binding reinforce open chromatin”; page not provided) reports that p63 enhances the chromatin targeting of the BAF remodeling complex. This supports an example of a transcription factor recruiting a remodeler, but the paper does not establish that many pioneer factors do so. (LLM, conf=0.82)

  cites [46] Bao X, Rubin AJ, Qu K et al (2015) A novel ATAC-seq approach reveals lineage-specific reinforcement of the open chromatin landscape via cooperation between BAF and p63. Genome Biol 16:284 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [47] Engelen E, Akinci U, Bryne JC et al (2011) Sox2 cooperates with Chd7 to regulate genes that are mutated in human syndromes. Nat Genet 43:607–611 source
• Partly supported

  In “BRG1 becomes associated with the GATA1 enhancers during differentiation of HSCs” (page unavailable), the authors say their results are consistent with GATA1 helping recruit BRG1, a chromatin remodeler. The paper does not establish that many pioneer factors recruit remodelers; its recruitment finding concerns GATA1 and is framed as an inference. (LLM, conf=0.86)

  cites [48] Hu G, Schones DE, Cui K et al (2011) Regulation of nucleosome landscape and transcription factor targeting at tissue-specific enhancers by BRG1. Genome Res 21:1650–1658 sourceExperimental — accuracy not yet measured
• Supported

  The Discussion states that GATA3’s pioneering activity correlates with recruitment of the SWI/SNF chromatin-remodeling complex. This paper supports the cited sentence as an example of a pioneer factor that recruits a remodeler; the sentence’s broader claim draws on multiple citations. (LLM, conf=0.90)

  cites [49] Takaku M, Grimm SA, Shimbo T et al (2016) GATA3-dependent cellular reprogramming requires activation-domain dependent recruitment of a chromatin remodeler. Genome Biol 17:36 source
• Partly supported

  The Results section ('AP-1 TFs recruit the BAF complex to enhancers'; page unavailable) supports recruitment of a chromatin remodeler by AP-1. The Discussion describes AP-1 nucleosome binding as a possible pioneer-factor property, but the paper does not establish that many other pioneer factors recruit remodelers. (LLM, conf=0.85)

  cites [50] Vierbuchen T, Ling E, Cowley CJ et al (2017) AP-1 transcription factors and the BAF complex mediate signal-dependent enhancer selection. Mol Cell 68:1067-1082.e12 sourceExperimental — accuracy not yet measured

In the paper

“In contrast, later studies of Drosophila developmental enhancers found evidence for much less rigid motif organisation [90, 91].”

What the sources say

• ?Not checked — no full text of the sourcecites [90] Kulkarni MM, Arnosti DN (2003) Information display by transcriptional enhancers. Development 130:6569–6575 source
• Supported

  The Results report no consistent spacing or orientation of transcription-factor motifs in the Drosophila cardiac enhancers studied, despite their shared function. The Introduction likewise describes their regulation as occurring without consistent motif grammar. (LLM, conf=0.94)

  cites [91] Junion G, Spivakov M, Girardot C et al (2012) A transcription factor collective defines cardiac cell fate and reflects lineage history. Cell 148:473–486 source

In the paper

“Consistent with this, homologous enhancers in a pair of distantly related species can retain their function in the non-host organism despite differences in DNA sequence [101, 110, 111].”

What the sources say

• ?Not checked — no full text of the sourcecites [101] Wong ES, Zheng D, Tan SZ et al (2020) Deep conservation of the enhancer regulatory code in animals. Science 370:eaax8137 source
• Supported

  The Results (Experimental validation of computationally predicted enhancers; page unavailable) report that mouse and zebrafish enhancer sequences were tested in zebrafish embryos, where some drove consistent expression patterns. The paper identifies these as functionally homologous enhancers with divergent sequences, supporting the claim that enhancer function can persist in a non-host organism despite sequence differences. (LLM, conf=0.92)

  cites [110] Yang S, Oksenberg N, Takayama S et al (2015) Functionally conserved enhancers with divergent sequences in distant vertebrates. BMC Genomics 16:882 source
• Supported

  In Results, under “Evolutionary conservation of CE64 sequence versus functional organisation” (page unavailable), the spotted gar CE64 sequence drove expression in transgenic mouse embryos despite lacking a region present in mouse CE64. The activity did not fully reproduce that of the mouse enhancer, but it supports the claim that homologous enhancers can retain function across species despite sequence differences. (LLM, conf=0.90)

  cites [111] Hörnblad A, Bastide S, Langenfeld K et al (2021) Dissection of the Fgf8 regulatory landscape by in vivo CRISPR-editing reveals extensive intra- and inter-enhancer redundancy. Nat Commun 12:439 source

In the paper

“Interestingly, at non-dimeric sites, physical interactions between co-bound TFs were not necessary [119], consistent with evidence from Drosophila where co-occupied DNA-bound TFs were often spaced quite widely apart (> 50 bp) [118].”

What the sources say

• Supported

  The Discussion reports high co-occupancy among TFs unlikely to physically interact and explicitly says TF–TF interactions are not mandatory for it. It contrasts this with dimeric motifs, where such interactions stabilize binding. (LLM, conf=0.93)

  cites [119] Sönmezer C, Kleinendorst R, Imanci D et al (2021) Molecular co-occupancy identifies transcription factor binding cooperativity in vivo. Mol Cell 81:255-267.e6 source

In the paper

“SMF has also recently been adapted to mammalian cells, detecting widespread co-occupancy of cooperative TFs at enhancers [119].”

What the sources say

• Partly supported

  The Introduction says the authors adapted SMF for mammalian genomes and found DNA co-occupancy widespread at sites of TF cooperativity. The Results describe measurements at mouse cis-regulatory elements, but the passages do not establish that the widespread finding applies specifically to enhancers rather than cis-regulatory elements more broadly. (LLM, conf=0.82)

  cites [119] Sönmezer C, Kleinendorst R, Imanci D et al (2021) Molecular co-occupancy identifies transcription factor binding cooperativity in vivo. Mol Cell 81:255-267.e6 sourceExperimental — accuracy not yet measured

In the paper

“The prevalence of flexible enhancer organisation is also supported by rapid evolutionary turnover of enhancers, which exceeds that of proximal promoter regions [29, 99, 100].”

What the sources say

• Partly supported

  Section I, “General principles of enhancer function” (page unavailable), describes high rates of enhancer turnover and links compensatory motif turnover to flexible enhancer organisation. The passages do not compare enhancer turnover with turnover in proximal promoter regions, so the claimed difference in rates is not supported by this source. (LLM, conf=0.84)

  cites [29] Long HK, Prescott SL, Wysocka J (2016) Ever-changing landscapes: transcriptional enhancers in development and evolution. Cell 167:1170–1187 sourceExperimental — accuracy not yet measured
• Supported

  The Results section, “Enhancer Evolution Is Appreciably More Rapid Than Proximal Promoter Evolution” (page not provided), reports rapid, widespread enhancer evolution and comparatively stable promoter activity in mammalian liver. The Discussion states that active enhancers have a mean lifetime three times shorter than active promoters. The link to flexible enhancer organisation is the citing authors’ interpretation, not a separate finding attributed to this source. (LLM, conf=0.93)

  cites [99] Villar D, Berthelot C, Aldridge S et al (2015) Enhancer evolution across 20 mammalian species. Cell 160:554–566 source
• Supported

  The Results section, “Enhancers are the main source of regulatory repurposing” (page not provided), reports that ancestral liver enhancers were lost at about 1.92 times the rate of ancestral promoters. This supports the attributed comparison in evolutionary turnover; the statement about flexible enhancer organisation is the citing authors’ framing. (LLM, conf=0.93)

  cites [100] Carelli FN, Liechti A, Halbert J et al (2018) Repurposing of promoters and enhancers during mammalian evolution. Nat Commun 9:4066 source

In the paper

“First developed in Drosophila, SMF can detect the binding of multiple TFs on a single DNA molecule [116].”

What the sources say

• Partly supported

  In Results, “Simultaneous Quantification of Multiple Transcription Initiation Intermediates” (page unavailable), the paper reports measuring multiple promoter-occupancy events on single molecules. It identifies footprints associated with the pre-initiation complex and RNA polymerase II, but does not show detection of multiple individual transcription factors on one molecule. The “Principles of Dual-Enzyme Single-Molecule Footprinting” section says the method was applied in Drosophila, not that SMF was first developed there. (LLM, conf=0.85)

  cites [116] Krebs AR, Imanci D, Hoerner L et al (2017) Genome-wide single-molecule footprinting reveals high RNA polymerase II turnover at paused promoters. Mol Cell 67:411–422.e4 sourceExperimental — accuracy not yet measured

In the paper

“Instead, enhancer conservation is typically “modular”, commonly preserving the sets of binding sites for the required TFs, but often not their exact sequence and orientation [101–109].”

What the sources say

• ?Not checked — no full text of the sourcecites [101] Wong ES, Zheng D, Tan SZ et al (2020) Deep conservation of the enhancer regulatory code in animals. Science 370:eaax8137 source
• Partly supported

  The Results sections “Identification of Sepsid Enhancers” and “Binding Site Composition and Organization in Sepsid Enhancers” describe similar collections of predicted binding sites for relevant transcription factors, while finding that few individual sites remain in the same location across families. The Discussion (“Binding Site Turnover and the Regulatory Machinery”) likewise reports extensive rearrangement with conserved enhancer function. These findings support the claim for the eve enhancers studied, but do not establish that this pattern is typical of enhancers generally; the passages also do not specifically assess binding-site orientation. Page numbers are unavailable in our copy. (LLM, conf=0.78)

  cites [102] Hare EE, Peterson BK, Iyer VN et al (2008) Sepsid even-skipped enhancers are functionally conserved in Drosophila despite lack of sequence conservation. PLoS Genet 4:e1000106 sourceExperimental — accuracy not yet measured
• Partly supported

  The paper’s discussion (page not provided in our copy) says that tissue-specific regulation can persist through regulatory architectures independent of sequence constraint, and describes nearby compensatory TF-binding events. This supports flexibility in binding-site sequence and location, but the study examines two liver TFs and does not establish that enhancers typically preserve the full set of required TF sites or that site orientation often changes. (LLM, conf=0.85)

  cites [103] Schmidt D, Wilson MD, Ballester B et al (2010) Five-vertebrate ChIP-seq reveals the evolutionary dynamics of transcription factor binding. Science 328:1036–1040 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [104] Dowell RD (2010) Transcription factor binding variation in the evolution of gene regulation. Trends Genet 26:468–475 source
• ?Could not verify

  The Results section on functional conservation of regulatory DNA motifs describes conserved motif–function associations, while the Discussion describes turnover of target genes within functional modules. Neither passage addresses whether individual enhancers preserve sets of transcription-factor binding sites while their exact sequences and orientations change; searches for enhancer conservation, binding-site sequence, and orientation did not establish that claim. (LLM, conf=0.73)

  cites [105] Habib N, Wapinski I, Margalit H et al (2012) A functional selection model explains evolutionary robustness despite plasticity in regulatory networks. Mol Syst Biol 8:619 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results sections “Specific selection of transcription factor sequence motifs” and “Compensatory changes stabilize regulatory output” link conserved enhancer activity to transcription-factor motifs rather than overall sequence identity, and show that lost motifs can be replaced at different positions. The paper does not establish that binding-site orientation is often unconserved, and its findings concern the assayed Drosophila enhancers rather than enhancers generally. Page numbers are unavailable in our copy. (LLM, conf=0.85)

  cites [106] Arnold CD, Gerlach D, Spies D et al (2014) Quantitative genome-wide enhancer activity maps for five Drosophila species show functional enhancer conservation and turnover during cis-regulatory evolution. Nat Genet 46:685–692 sourceExperimental — accuracy not yet measured
• Partly supported

  In ‘Dose of activation’ as an evolutionary trait (page unavailable), the paper says regulatory regions can retain function despite low base-by-base conservation and turnover of transcription-factor binding sites. In ‘Do “doses” add up at multi-TF enhancers?’ (page unavailable), it describes flexible enhancer organisation, but it does not establish that conserved enhancers commonly preserve the same sets of required transcription factors or address binding-site orientation. (LLM, conf=0.84)

  cites [107] Spivakov M (2014) Spurious transcription factor binding: non-functional or genetically redundant? BioEssays 36:798–806 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [108] Borok MJ, Tran DA, Ho MCW, Drewell RA (2010) Dissecting the regulatory switches of development: lessons from enhancer evolution in Drosophila. Development 137:5–13 source
• Partly supported

  The Results section, “Conserved enhancer occupancy is associated with enhancer function” (page unavailable), says evolutionary constraints likely select for overall enhancer composition rather than the positions of individual binding sites. The Results section, “TF binding site divergence is tolerated in the context of collective TF binding” (page unavailable), also reports conserved Tin occupancy despite motif divergence in enhancers bound by many factors. These findings support flexibility in some enhancers, but do not establish that enhancer conservation is typically modular or that exact binding-site orientation is often not preserved. (LLM, conf=0.83)

  cites [109] Khoueiry P, Girardot C, Ciglar L et al (2017) Uncoupling evolutionary changes in DNA sequence, transcription factor occupancy and enhancer activity. Elife 6:e28440 sourceExperimental — accuracy not yet measured

In the paper

“Flexibility of enhancer organisation with respect to binding site order and orientation was also observed in a massively parallel synthetic enhancer experiment [55].”

What the sources say

• Partly supported

  The Results and Discussion (page numbers unavailable) report that multiple binding-site orders drove strong expression in the massively parallel synthetic regulatory-element experiment, supporting flexibility with respect to order. The passages do not report a test or finding about binding-site orientation, so that part of the claim is unsupported. (LLM, conf=0.86)

  cites [55] Smith RP, Taher L, Patwardhan RP et al (2013) Massively parallel decoding of mammalian regulatory sequences supports a flexible organizational model. Nat Genet 45:1021–1028 sourceExperimental — accuracy not yet measured

In the paper

“While it is clear that enhancers typically favour the more flexible models of sequence organisation, the degree of necessary cooperation between TFs is less obvious, particularly since TF binding to DNA appears to be very transient [113–117].”

What the sources say

• Partly supported

  The Results and discussion report that each of four transcription factors occupies about 50% of its targeted regulatory modules transiently during Drosophila development. This supports changing DNA occupancy over developmental stages, but not the broader implication that transcription-factor binding to DNA is generally short-lived at the molecular level. (LLM, conf=0.86)

  cites [113] Wilczyński B, Furlong EEM (2010) Dynamic CRM occupancy reflects a temporal map of developmental progression. Mol Syst Biol 6:383 sourceExperimental — accuracy not yet measured
• Supported

  The ‘In vivo dynamics of transcription factors’ section says residence times on genomic elements can be short and that rapid exchange is a property of many transcription factors. It notes an exception for some site-specific DNA-binding proteins with longer residence times, which is consistent with the citing sentence’s qualified wording, ‘appears to be.’ (LLM, conf=0.92)

  cites [114] Voss TC, Hager GL (2014) Dynamic regulation of transcriptional states by chromatin and transcription factors. Nat Rev Genet 15:69–81 source
• ?Not checked — no full text of the sourcecites [115] Chen J, Zhang Z, Li L et al (2014) Single-molecule dynamics of enhanceosome assembly in embryonic stem cells. Cell 156:1274–1285 source
• Partly supported

  In Results, ‘The TATA Box Stabilizes Binding of the PIC’ (page unavailable), the paper says PIC formation at TATA-less promoters may be transient. It does not establish that transcription-factor binding to DNA is generally very transient; it instead reports more frequent PIC binding at TATA-containing promoters and notes that some general transcription factors may form a stable scaffold. (LLM, conf=0.87)

  cites [116] Krebs AR, Imanci D, Hoerner L et al (2017) Genome-wide single-molecule footprinting reveals high RNA polymerase II turnover at paused promoters. Mol Cell 67:411–422.e4 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [117] Stergachis AB, Debo BM, Haugen E et al (2020) Single-molecule regulatory architectures captured by chromatin fiber sequencing. Science 368:1449–1454 source

In the paper

“Additionally, a deep learning algorithm applied to binding of pluripotency TFs at base resolution in mouse embryonic stem cells (ESCs) suggested a “soft motif syntax” whereby TFs help each other to bind directly to the enhancer with a lenient distance-dependency between binding sites [98].”

What the sources say

• Supported

  Split into 2 atoms — 2 confirmed, 0 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [98] Avsec Ž, Weilert M, Shrikumar A et al (2021) Base-resolution models of transcription-factor binding reveal soft motif syntax. Nat Genet 53:354–366 source

In the paper

“Similarly, TFs governing serotonergic neuron differentiation showed synergy despite flexible sequence organisation of their cognate enhancers [95].”

What the sources say

• Supported

  The Results section, “HSN TF collective shows enhancer-context dependent synergistic relationships” (page not specified), reports synergy among factors regulating serotonergic HSN neuron genes. It also links reporter-specific synergy to the different arrangements of functional binding sites in each enhancer. The paper notes that the interactions depend on the enhancer and TF pair, rather than occurring universally. (LLM, conf=0.91)

  cites [95] Lloret-Fernández C, Maicas M, Mora-Martínez C et al (2018) A transcription factor collective defines the HSN serotonergic neuron regulatory landscape. Elife 7:e32785 source

In the paper

“In addition to the ‘primed’ and active states, enhancers can also reside in a ‘poised’ state exhibiting both H3K4me1 and H3 lysine 27 trimethylation (H3K27me3) that is associated with Polycomb repressive complexes [131, 135, 136, 138].”

What the sources say

• Partly supported

  The paper’s Extended Data Fig. 3 caption describes a cCRE that may be a poised enhancer in embryonic stem cells, iPSCs and neural progenitor cells. It associates that example with high DNase and H3K4me3 signals, not the claimed combination of H3K4me1 and H3K27me3; the passages found also do not link poised enhancers to Polycomb repressive complexes. (LLM, conf=0.86)

  cites [131] ENCODE Project Consortium, Moore JE, Purcaro MJ et al (2020) Expanded encyclopaedias of DNA elements in the human and mouse genomes. Nature 583:699–710 sourceExperimental — accuracy not yet measured
• Supported

  The paper describes poised enhancers in human embryonic stem cells as marked by H3K4me1 and enriched for H3K27me3 (Abstract; passage 8). It also reports simultaneous presence of the two marks and enrichment of SUZ12, a PRC2 component, at those elements (passage 2). (LLM, conf=0.94)

  cites [135] Rada-Iglesias A, Bajpai R, Swigut T et al (2011) A unique chromatin signature uncovers early developmental enhancers in humans. Nature 470:279–283 source
• Supported

  In “Identification of three major enhancer classes in mESCs” (page not provided), the paper classifies H3K4me1+, H3K27me3+ enhancers as poised. The same section reports substantial binding of Polycomb group proteins at poised enhancers. The paper also identifies poised enhancers without H3K27me3, but that does not conflict with the claim that enhancers can occupy the described state. (LLM, conf=0.93)

  cites [136] Zentner GE, Tesar PJ, Scacheri PC (2011) Epigenetic signatures distinguish multiple classes of enhancers with distinct cellular functions. Genome Res 21:1273–1283 source
• ?Could not verify

  The Background discusses H3K27ac as a marker of active enhancers, not the claimed combination of H3K4me1 and H3K27me3 at poised enhancers. Searches for those marks, poised enhancers, and Polycomb found only related reference titles, not a passage in this paper stating the claim. The indexed passages have no page numbers. (LLM, conf=0.73)

  cites [138] Koenecke N, Johnston J, Gaertner B et al (2016) Genome-wide identification of Drosophila dorso-ventral enhancers by differential histone acetylation analysis. Genome Biol 17:196 sourceExperimental — accuracy not yet measured

In the paper

“Furthermore, even enhancers with extremely high evolutionary conservation may not require the exact preservation of their sequence for their correct function, as was recently shown using CRISPR-mediated mutagenesis [112].”

What the sources say

• Supported

  The Discussion reports that most tested ultraconserved enhancers retained activity after mutation, supporting the claim that their known enhancer functions do not require perfect sequence preservation. The Methods (Generation of Enhancer Knock-in Mice) also describe using CRISPR/Cas9 to replace endogenous enhancers with mutated alleles. The paper cautions that effects at untested developmental stages or on other functions remain possible. (LLM, conf=0.92)

  cites [112] Snetkova V, Ypsilanti AR, Akiyama JA et al (2021) Ultraconserved enhancer function does not require perfect sequence conservation. Nat Genet 53:521–528 source

In the paper

“This study detected multiple “waves” of a progressive recruitment of at least 46 sequence-specific TFs and cofactors, including chromatin remodelling complexes and histone acetyl-transferases, eventually resulting in a derepression of an oestrogen-response gene pS2 [87].”

What the sources say

• Partly supported

  Split into 2 atoms — 0 confirmed, 2 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [87] Métivier R, Penot G, Hübner MR et al (2003) Estrogen receptor-α directs ordered, cyclical, and combinatorial recruitment of cofactors on a natural target promoter. Cell 115:751–763 sourceExperimental — accuracy not yet measured

In the paper

“Notably, at dimeric motif sites, TFs tended to bind initially as monomers with subsequent dimerisation presumably stabilising the DNA–protein complex [119].”

What the sources say

• Supported

  The Discussion says that dimerization is not obligatory for most tested TF dimers and suggests sequential monomer binding as their binding mode. It also says TF–TF interactions stabilize binding at dimeric motifs. The citing sentence preserves the paper’s tentative framing. (LLM, conf=0.90)

  cites [119] Sönmezer C, Kleinendorst R, Imanci D et al (2021) Molecular co-occupancy identifies transcription factor binding cooperativity in vivo. Mol Cell 81:255-267.e6 source

In the paper

“In particular, Mediator and BRD4 co-localise at very high levels at “super-enhancers” (SEs): long stretches of sequences with gene regulatory activity that generally drive greater levels of gene expression than regular enhancers [69–72].”

What the sources say

• Supported

  The Introduction describes super-enhancers as enriched for BRD4 and Mediator and associated with high transcriptional activity. The Results section defines them as extended regions with high levels of both proteins, supporting the cited sentence’s description; the paper does not give a direct expression-level comparison with regular enhancers. (LLM, conf=0.86)

  cites [69] Crump NT, Ballabio E, Godfrey L et al (2021) BET inhibition disrupts transcription but retains enhancer–promoter contact. Nat Commun 12:223 source
• Partly supported

  The Results section (“Disproportionate binding of BRD4 to overloaded enhancers”; page unavailable) reports exceptionally high BRD4 loading at enhancers larger than typical enhancers. The Discussion (page unavailable) describes them as approximately 12-fold larger. Our copy does not establish co-localization with Mediator or show that these enhancers generally drive higher gene expression than regular enhancers; it reports greater sensitivity to BET inhibition instead. (LLM, conf=0.87)

  cites [70] Chapuy B, McKeown MR, Lin CY et al (2013) Discovery and characterization of super-enhancer-associated dependencies in diffuse large B cell lymphoma. Cancer Cell 24:777–790 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results sections (page unavailable) report that BRD4 and Mediator subunits localize at super-enhancers of stem-cell genes in human embryonic stem cells, and describe super-enhancers as large enhancer domains. The copy does not establish that super-enhancers generally drive higher gene expression than regular enhancers. It also reports that BRD4 peak signal intensity was similar at super-enhancers and typical enhancers in these cells, although BRD4-binding peaks were more frequent at super-enhancers. (LLM, conf=0.85)

  cites [71] Di Micco R, Fontanals-Cirera B, Low V et al (2014) Control of embryonic stem cell identity by BRD4-dependent transcriptional elongation of super-enhancer-associated pluripotency genes. Cell Rep 9:234–247 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results sections on large genomic domains and functional attributes describe super-enhancers as unusually long, highly occupied by Mediator, and associated with higher gene expression than typical enhancers. The passages have no page numbers in our copy. They do not establish that BRD4 co-localises with Mediator at super-enhancers. (LLM, conf=0.88)

  cites [72] Whyte WA, Orlando DA, Hnisz D et al (2013) Master transcription factors and mediator establish super-enhancers at key cell identity genes. Cell 153:307–319 sourceExperimental — accuracy not yet measured

In the paper

“The TF collective model was originally formulated in Drosophila, where a set of five TFs bound multitudes of enhancers required for heart development that showed no detectable similarity in their sequence organisation, beyond the presence of binding motifs for some of these TFs [91].”

What the sources say

• Partly supported

  Split into 2 atoms — 1 confirmed, 1 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [91] Junion G, Spivakov M, Girardot C et al (2012) A transcription factor collective defines cardiac cell fate and reflects lineage history. Cell 148:473–486 sourceExperimental — accuracy not yet measured

In the paper

“The stripe 2 enhancer controls eve expression in early embryogenesis via binding of tissue-specific TFs [121–123].”

What the sources say

• ?Not checked — no full text of the sourcecites [121] Stanojevic D, Small S, Levine M (1991) Regulation of a segmentation stripe by overlapping activators and repressors in the Drosophila embryo. Science 254:1385–1387 source
• ?Not checked — no full text of the sourcecites [122] Goto T, Macdonald P, Maniatis T (1989) Early and late periodic patterns of even skipped expression are controlled by distinct regulatory elements that respond to different spatial cues. Cell 57:413–422 source
• Partly supported

  The Abstract and Discussion describe the stripe 2 regulatory element as binding factors that activate or repress eve expression in early embryos. They identify these factors as a maternal morphogen and gap proteins with spatially patterned expression, but do not establish that they are tissue-specific transcription factors. (LLM, conf=0.82)

  cites [123] Small S, Kraut R, Hoey T et al (1991) Transcriptional regulation of a pair-rule stripe in Drosophila. Genes Dev 5:827–839 sourceExperimental — accuracy not yet measured

In the paper

“Consistent with this, “strengthening” TF binding sites at developmental enhancers results in aberrant gene expression patterns [125].”

What the sources say

• Supported

  The supplied passages (labelled Abstract; no page number provided) report that making all five GATA and ETS binding motifs in the Otx-a developmental enhancer optimal produced expression in ectopic tissues. The paper also states that excess optimal binding sites can lead to loss of specificity and ectopic gene expression. (LLM, conf=0.93)

  cites [125] Farley EK, Olson KM, Zhang W et al (2015) Suboptimization of developmental enhancers. Science 350:325–328 source

In the paper

“Enhancers consistent with the billboard model are likely also widespread in vertebrate genomes [55, 93].”

What the sources say

• Partly supported

  The Results section (“Reporter expression is influenced by binding site order”; page unavailable) says the tested configurations are largely consistent with the billboard model. The Results section (“Synthetic elements mimic putative liver enhancers”; page unavailable) also finds similar binding-site configurations in 78% of the putative mouse liver enhancers examined. This supports the model’s relevance to native mammalian enhancers, but does not establish that billboard-consistent enhancers are widespread across vertebrate genomes. (LLM, conf=0.84)

  cites [55] Smith RP, Taher L, Patwardhan RP et al (2013) Massively parallel decoding of mammalian regulatory sequences supports a flexible organizational model. Nat Genet 45:1021–1028 sourceExperimental — accuracy not yet measured
• Partly supported

  In “Highly conserved non-coding sequences and transcriptional regulation” (page unavailable), the paper calls many enhancers loose, variable assemblies of binding sites and identifies that structure as the billboard model. It also finds billboard-like organization in five zebrafish notochord enhancers. Those examples support the possibility, but the paper does not establish how widespread billboard-like enhancers are across vertebrate genomes. (LLM, conf=0.83)

  cites [93] Rastegar S, Hess I, Dickmeis T et al (2008) The words of the regulatory code are arranged in a variable manner in highly conserved enhancers. Dev Biol 318:366–377 sourceExperimental — accuracy not yet measured

In the paper

“The various proposed models for enhancer grammar range from very rigid to flexible, largely dependent on the species, cell types and loci in which they were formulated (reviewed in [29]).”

What the sources say

• Partly supported

  In the Introduction, under “Motif organization, enhancer grammar and models of enhancer architecture” (page unavailable), the review contrasts the rigid enhanceosome model with the more flexible billboard model and describes a spectrum between them. It gives examples from particular species and loci, but does not establish that the differences among proposed models are largely dependent on the species, cell types, and loci in which they were formulated. (LLM, conf=0.82)

  cites [29] Long HK, Prescott SL, Wysocka J (2016) Ever-changing landscapes: transcriptional enhancers in development and evolution. Cell 167:1170–1187 sourceExperimental — accuracy not yet measured

In the paper

“Interestingly, at non-dimeric sites, physical interactions between co-bound TFs were not necessary [119], consistent with evidence from Drosophila where co-occupied DNA-bound TFs were often spaced quite widely apart (> 50 bp) [118].”

What the sources say

• Partly supported

  The Results section, “Cooperative binding is common at active enhancers,” reports co-binding across spacings of 50–90 bp and significant cooperativity at distances up to 140 bp, supporting the point that Drosophila TFs can co-occupy widely separated sites. But “Many active enhancers contain multiple bound TFs” says ~50 bp was the most observed spacing; the copy does not establish that co-occupied factors were often more than 50 bp apart. (LLM, conf=0.83)

  cites [118] Rao S, Ahmad K, Ramachandran S (2021) Cooperative binding between distant transcription factors is a hallmark of active enhancers. Mol Cell 81:1651-1665.e4 sourceExperimental — accuracy not yet measured

In the paper

“For example, enhancers devoid of the classic H3K27ac mark have recently been described [163]; consistent with this, it was shown that this mark is not required for enhancer function, at least in mouse ESCs [164].”

What the sources say

• Supported

  The Abstract states that the paper identifies active functional enhancers marked by H3K122ac that lack H3K27ac. It also describes an H3K27ac-negative enhancer group in mouse embryonic stem cells, directly supporting the cited claim. The separate assertion about whether H3K27ac is required for enhancer function is attached to citation [164], not this source. (LLM, conf=0.94)

  cites [163] Pradeepa MM, Grimes GR, Kumar Y et al (2016) Histone H3 globular domain acetylation identifies a new class of enhancers. Nat Genet 48:681–686 source

In the paper

“Using these techniques with antibodies to enhancer- and promoter-associated histone modifications or those to RNA polymerase has also revealed large numbers of 3D enhancer–promoter contacts [244, 295–301].”

What the sources say

• Supported

  The Results section (‘H3K27ac HiChIP identifies functional enhancer interactions’; page unavailable) reports 4,191 chromatin loops detected using the enhancer- and promoter-associated histone mark H3K27ac to interrogate enhancer–promoter interactions. The Online Methods (‘HiChIP’; page unavailable) specify that the assay used an antibody to H3K27ac. This supports the histone-antibody route described in the sentence; the cited paper need not also demonstrate the RNA-polymerase alternative. (LLM, conf=0.91)

  cites [244] Mumbach MR, Satpathy AT, Boyle EA et al (2017) Enhancer connectome in primary human cells identifies target genes of disease-associated DNA elements. Nat Genet 49:1602–1612 source
• Partly supported

  The Results section (“Enhancer-Promoter Interactions Depend on YY1 in Living Cells”; page unavailable) reports that H3K27ac HiChIP detected enhancer–promoter interactions; H3K27ac is a modification present at both enhancers and promoters. The paper therefore supports the histone-modification part of the claim, but the passages found do not show contacts detected using antibodies to RNA polymerase. (LLM, conf=0.86)

  cites [295] Weintraub AS, Li CH, Zamudio AV et al (2017) YY1 Is a structural regulator of enhancer-promoter loops. Cell 171:1573-1588.e28 sourceExperimental — accuracy not yet measured
• Partly supported

  The Methods (HiChIP sequencing and analysis; page unavailable) describe using an antibody to the enhancer-associated H3K27ac mark to identify long-range DNA loops. The Results (HLA-DR3 and -DR15 hQTLs differentially regulate HLA genes; page unavailable) report looping between an enhancer and the HLA-DRB1 promoter. Our copy does not show that promoter-associated histone or RNA-polymerase antibodies revealed contacts, or establish the claimed large number of enhancer–promoter contacts. (LLM, conf=0.85)

  cites [296] Pelikan RC, Kelly JA, Fu Y et al (2018) Enhancer histone-QTLs are enriched on autoimmune risk haplotypes and influence gene expression within chromatin networks. Nat Commun 9:2905 sourceExperimental — accuracy not yet measured
• Supported

  The passage labeled Abstract describes PLAC-seq using H3K4me3 ChIP-seq to enrich for active promoters, then reports 219,509 significant chromatin interactions across brain cell types. This supports the cited paper as an example of a histone-modification-antibody technique revealing many 3D promoter–enhancer contacts; it does not need to have used the alternative RNA-polymerase antibody approach. (LLM, conf=0.91)

  cites [297] Nott A, Holtman IR, Coufal NG et al (2019) Brain cell type-specific enhancer–promoter interactome maps and diseaserisk association. Science 366:1134–1139 source
• Partly supported

  The Methods (§4.3) describe using an H3K27Ac antibody, and the Results (§2.1) report 66,092–449,157 chromatin loops per cell line, with 35%–40% contacting a promoter. This supports the histone-modification example and the large number of contacts, but the paper does not report using antibodies to RNA polymerase. (LLM, conf=0.84)

  cites [298] O’Mara TA, Spurdle AB, Glubb DM, Endometrial Cancer Association Consortium (2019) Analysis of promoter-associated chromatin interactions reveals biologically relevant candidate target genes at endometrial cancer risk loci. Cancers 11:1440 sourceExperimental — accuracy not yet measured
• Supported

  The Materials and Methods (HiChIP experiments) state that the authors used chromatin immunoprecipitation against the enhancer- and promoter-associated mark H3K27ac. The Results (A compendium of activity and chromatin interactions in KCs) report more than 50,000 significant interactions per HiChIP dataset, including interactions involving active enhancers and promoters. This paper supports the histone-modification example in the citing sentence; it does not need to establish the alternative RNA-polymerase approach. (LLM, conf=0.92)

  cites [299] Shi C, Ray-Jones H, Ding J et al (2021) Chromatin looping links target genes with genetic risk loci for dermatological traits. J Invest Dermatol. https://doi.org/10.1016/j.jid.2021.01.015 source
• Supported

  In Results, under “A survey of enhancer-promoter 3D looping in the intestinal epithelium” (page unavailable), the paper reports using HiChIP with antibodies to H3K4me3, a promoter-associated histone modification, to detect 3D chromatin interactions. It identifies enhancer-promoter interactions as one of the predominant interaction types observed. This paper supports the promoter-antibody example in the sentence’s group of citations. (LLM, conf=0.87)

  cites [300] Chen L, Cao W, Aita R et al (2021) Three-dimensional interactions between enhancers and promoters during intestinal differentiation depend upon HNF4. Cell Rep 34:108679 source
• Partly supported

  The Results section (page unavailable) says the authors used H3K4me3 antibodies with PLAC-seq to detect promoter-centered chromatin contacts and found thousands of contacts involving distal enhancers or promoters. This supports the histone-modification portion, but the paper does not report using RNA-polymerase antibodies or enhancer-associated histone antibodies to detect those contacts. (LLM, conf=0.86)

  cites [301] Kubo N, Ishii H, Xiong X et al (2021) Promoter-proximal CTCF binding promotes distal enhancer-dependent gene activation. Nat Struct Mol Biol 28:152–161 sourceExperimental — accuracy not yet measured

In the paper

“Many developmental enhancers are found in the poised or Polycomb-repressed state in early development [139, 142, 143], and the functional role of these states remains an area of active research.”

What the sources say

• Supported

  The Results section (page unavailable) reports that PRC2-bound regions in mouse embryonic stem cells carry a chromatin signature previously identified for poised enhancers. It also finds that many of these regions acquire enhancer-associated activity during differentiation, supporting the claim that developmental enhancers can be poised or Polycomb-repressed early in development. (LLM, conf=0.88)

  cites [139] Ngan CY, Wong CH, Tjong H et al (2020) Chromatin interaction analyses elucidate the roles of PRC2-bound silencers in mouse development. Nat Genet 52:264–272 source
• Supported

  The Introduction describes poised enhancers linked to developmental genes as bound by Polycomb complexes before activation. The Results report 3,057 poised enhancers in the early mouse E6.5 epiblast, where they carry the Polycomb-associated H3K27me3 mark and remain inactive. (LLM, conf=0.90)

  cites [142] Crispatzu G, Rehimi R, Pachano T et al (2021) The chromatin, topological and regulatory properties of pluripotency-associated poised enhancers are conserved in vivo. Nat Commun 12(1):4344 source
• ?Not checked — no full text of the sourcecites [143] Cruz-Molina S, Respuela P, Tebartz C et al (2017) PRC2 facilitates the regulatory topology required for poised enhancer function during pluripotent stem cell differentiation. Cell Stem Cell 20:689-705.e9 source

In the paper

“For example, enhancers devoid of the classic H3K27ac mark have recently been described [163]; consistent with this, it was shown that this mark is not required for enhancer function, at least in mouse ESCs [164].”

What the sources say

• Supported

  The Conclusions state that depletion of H3K27ac does not affect enhancer activity in mouse ESCs. This supports the citing sentence’s claim, including its restriction to mouse ESCs. (LLM, conf=0.94)

  cites [164] Zhang T, Zhang Z, Dong Q et al (2020) Histone H3K27 acetylation is dispensable for enhancer activity in mouse embryonic stem cells. Genome Biol 21:45 source

In the paper

“Notably, some enhancer variants that appear functionally neutral when tested under normal steady-state conditions may show “cryptic” effects under stress or upon stimulus response [107, 226, 227].”

What the sources say

• Supported

  In “The cryptic consequences of TFBS redundancy” (page not provided), the paper reports that removing two enhancer regions has no phenotype under normal conditions but causes defects at temperature extremes. The same section describes an otherwise neutral TFBS polymorphism whose effect is revealed when gonadal signalling is ablated, supporting the stimulus-response part of the claim. (LLM, conf=0.92)

  cites [107] Spivakov M (2014) Spurious transcription factor binding: non-functional or genetically redundant? BioEssays 36:798–806 source
• Partly supported

  In “How is cryptic genetic variation related to canalization?” (page unavailable), the paper says genetic effects can be suppressed under normal conditions and become visible when the environment changes. It does not specifically discuss enhancer variants, steady-state enhancer tests, or stimulus responses, so those details are not established by this source. (LLM, conf=0.84)

  cites [226] Gibson G, Reed LK (2008) Cryptic genetic variation. Curr Biol 18:R989–R990 sourceExperimental — accuracy not yet measured
• Supported

  In “How does CGV accumulate?” (page unavailable), the paper describes a cis-regulatory region that is largely superfluous under ideal conditions but necessary to preserve gene expression under thermal stress. This supports the claim that an apparently neutral enhancer can have a cryptic effect when conditions change. (LLM, conf=0.90)

  cites [227] Paaby AB, Rockman MV (2014) Cryptic genetic variation: evolution’s hidden substrate. Nat Rev Genet 15:247–258 source

In the paper

“Much of this variation has apparently little phenotypic effect due to redundancy both within enhancers (such as through multiple homotypic TF binding sites [191, 193]) and across them (such as through multiple enhancers regulating the same gene, as will be discussed below).”

What the sources say

• Supported

  The Discussion says much variation may be buffered within cis-regulatory modules by neighboring homotypic binding motifs, and the Background notes that many regulatory variants cause no apparent functional aberration. In the citing sentence, this citation is attached to redundancy within enhancers; redundancy across enhancers is a separate, uncited part of the sentence. (LLM, conf=0.92)

  cites [191] Spivakov M, Akhtar J, Kheradpour P et al (2012) Analysis of variation at transcription factor binding sites in Drosophila and humans. Genome Biol 13:R49 source
• Supported

  The paper’s introductory section identifies functional redundancy as a mechanistic advantage of homotypic clusters of transcription-factor binding sites. The citation is attached to that within-enhancer example, not to the separate claim about redundancy across enhancers. (LLM, conf=0.90)

  cites [193] Gotea V, Visel A, Westlund JM et al (2010) Homotypic clusters of transcription factor binding sites are a key component of human promoters and enhancers. Genome Res 20:565–577 source

In the paper

“These enrichment methods have detected multitudes of enhancer–promoter loops in many human and mouse cell types [243, 286–291].”

What the sources say

• Supported

  The Results section (“Promoter Capture Hi-C”; page not specified) reports that promoter capture Hi-C detected about 175,000 high-confidence interactions per cell type across 17 human primary blood cell types. The Results also identify enhancer–promoter interactions among those detected. This paper supports the human-cell-type portion of the multiply cited sentence; it does not need to establish the mouse-cell-type portion on its own. (LLM, conf=0.90)

  cites [243] Javierre BM, Burren OS, Wilder SP et al (2016) Lineage-specific genome architecture links enhancers and non-coding disease variants to target gene promoters. Cell 167:1369-1384.e19 source
• Partly supported

  The Introduction says promoter-capture Hi-C has identified thousands of enhancer–promoter contacts in different cell types. The Results report hundreds of thousands of promoter interactions in two human cell types, iPSCs and cardiomyocytes. These passages support the claim for human cells, but do not establish its mouse-cell component. (LLM, conf=0.84)

  cites [286] Montefiori LE, Sobreira DR, Sakabe NJ et al (2018) A promoter interaction map for cardiovascular disease genetics. Elife 7:e35788 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results sections report promoter capture Hi-C detection of hundreds of thousands of promoter contacts in mouse embryonic stem and trophoblast stem cells, including contacts with enhancers. The Introduction mentions studies in human cells, but this paper does not establish the claim's broader scope of multitudes of enhancer–promoter loops across many human and mouse cell types. The supplied copy gives no page numbers. (LLM, conf=0.82)

  cites [287] Schoenfelder S, Mifsud B, Senner CE et al (2018) Divergent wiring of repressive and active chromatin interactions between mouse embryonic and trophoblast lineages. Nat Commun 9:4189 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results section ('SEs Are Highly Interactive Hubs in ESCs'; page unavailable) reports that the authors used a 3C-based method to detect numerous enhancer–promoter interactions in mouse embryonic stem cells. This supports the mouse-cell example, but this paper does not establish the claim across many human and mouse cell types. (LLM, conf=0.85)

  cites [288] Novo CL, Javierre B-M, Cairns J et al (2018) Long-range enhancer interactions are prevalent in mouse embryonic stem cells and are reorganized upon pluripotent state transition. Cell Rep 22:2615–2627 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results (Summary of identified interactions; page unavailable) report thousands of Capture Hi-C interactions in two human cell lines, including some involving promoters. The paper does not establish that these are all enhancer–promoter loops, or report detection across many human and mouse cell types; its experiments used two human cell lines. (LLM, conf=0.88)

  cites [289] Martin P, McGovern A, Orozco G et al (2015) Capture Hi-C reveals novel candidate genes and complex long-range interactions with related autoimmune risk loci. Nat Commun 6:10069 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results section (page unavailable in our copy) reports 295,422 promoter interactions detected by Capture C in human osteoblasts and also describes a comparison with a human HepG2 cell line. It does not establish detection in many human and mouse cell types; the cited paper’s experiments described here involve human cells, not mouse cells. (LLM, conf=0.88)

  cites [290] Chesi A, Wagley Y, Johnson ME et al (2019) Genome-scale Capture C promoter interactions implicate effector genes at GWAS loci for bone mineral density. Nat Commun 10:1260 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results section (“Capture Hi-C identified novel gene targets in psoriasis susceptibility loci”; page unavailable) reports an average of 6,593 interactions across three capture experiments in human cell lines, with about 30% involving a transcription start site. The paper also identifies enhancer–promoter interactions using HiChIP in HaCaT cells. It does not establish the claim for many human and mouse cell types; the mouse-cell portion is unsupported by this source. (LLM, conf=0.86)

  cites [291] Ray-Jones H, Duffus K, McGovern A et al (2020) Mapping DNA interaction landscapes in psoriasis susceptibility loci highlights KLF4 as a target gene in 9q31. BMC Biol 18:47 sourceExperimental — accuracy not yet measured

In the paper

“A classic example is in obesity susceptibility, where trait-associated SNPs are located within the intron of the FTO gene, but lead to the dis-regulation of the distal IRX3 gene [210].”

What the sources say

• Partly supported

  The paper’s Abstract places obesity-associated SNPs in FTO introns and reports that they are associated with IRX3 expression in human brain. It also describes a long-range functional connection to IRX3, but treats the possibility that the variants disrupt enhancer activity and alter expression as a possibility; it does not establish the claim’s causal wording that the SNPs lead to IRX3 dysregulation. No page number is supplied in our copy. (LLM, conf=0.86)

  cites [210] Smemo S, Tena JJ, Kim K-H et al (2014) Obesity-associated variants within FTO form long-range functional connections with IRX3. Nature 507:371–375 sourceExperimental — accuracy not yet measured

In the paper

“For example, genetic variants associated with differential binding of PU.1 were found to underpin GWAS variants associated with blood cell count and autoimmune diseases [213–215].”

What the sources say

• ?Not checked — no full text of the sourcecites [213] Waszak SM, Delaneau O, Gschwind AR et al (2015) Population variation and genetic control of modular chromatin architecture in humans. Cell 162:1039–1050 source
• Partly supported

  In Results, under “bQTLs provide candidate molecular mechanisms for thousands of GWAS-implicated variants” (page unavailable), the paper identifies PU.1 binding QTLs linked to Crohn’s disease, inflammatory bowel disease, atopic dermatitis, and asthma GWAS loci. Our copy does not identify a PU.1 binding variant linked to blood cell count, and presents the disease mechanisms as candidates rather than established causes. (LLM, conf=0.85)

  cites [214] Tehranchi AK, Myrthil M, Martin T et al (2016) Pooled ChIP-Seq links variation in transcription factor binding to complex disease risk. Cell 165:730–741 sourceExperimental — accuracy not yet measured
• Supported

  The Introduction states that variants associated with differential PU.1 binding are enriched for genetic associations with cell count and susceptibility to autoimmune and inflammatory diseases. The Results section, “PU.1 differential binding underpins disease-associated variants,” reports PU.1-binding QTLs sharing genetic signals with blood-cell-trait and autoimmune-disease GWAS loci. (LLM, conf=0.94)

  cites [215] Watt S, Vasquez L, Walter K et al (2021) Genetic perturbation of PU.1 binding and chromatin looping at neutrophil enhancers associates with autoimmune disease. Nat Commun 12:2298 source

In the paper

“Finally, evidence from multiple sources including reporter assays [169, 175, 239, 240], CRISPR [240–242], chromatin mapping [243, 244] and population genetics analysis [207, 245] suggests that certain promoters, termed ‘epromoters’, can also function as enhancers (reviewed in [246]).”

What the sources say

• ?Not checked — no full text of the sourcecites [169] Arnold CD, Gerlach D, Stelzer C et al (2013) Genome-wide quantitative enhancer activity maps identified by STARR-seq. Science 339:1074–1077 source
• Supported

  The Abstract reports that the same DNA sequences typically encode both enhancer and promoter activities, and that gene promoters generated enhancer activity in the reporter assays. This supports the attributed point that some promoters can function as enhancers, even though the passages do not use the term ‘epromoters’. (LLM, conf=0.93)

  cites [175] Nguyen TA, Jones RD, Snavely AR et al (2016) High-throughput functional comparison of promoter and enhancer activities. Genome Res 26:1023–1033 source
• Supported

  In Results, under “Interacting Promoters Possess Combinatorial Regulatory Functions” (page not provided), the paper reports reporter-assay evidence that an INTS1 promoter fragment enhanced activity from the MAFK promoter. It explicitly concludes that a promoter can act as an enhancer, supporting the claim despite not using the term ‘epromoters.’ (LLM, conf=0.93)

  cites [239] Li G, Ruan X, Auerbach RK et al (2012) Extensive promoter-centered chromatin interactions provide a topological basis for transcription regulation. Cell 148:84–98 source
• ?Not checked — no full text of the sourcecites [240] Dao LTM, Galindo-Albarrán AO, Castro-Mondragon JA et al (2017) Genome-wide characterization of mammalian promoters with distal enhancer functions. Nat Genet 49:1073–1081 source
• Supported

  The paper states that promoter-proximal sequences activated neighboring genes at five of six examined loci and suggests that these promoters act through enhancer-like mechanisms (main text, presented under Abstract in our copy). Extended Data Fig. 7 explicitly describes promoters of lncRNAs and mRNAs as having enhancer-like functions. This supports the claim that certain promoters can function as enhancers. (LLM, conf=0.91)

  cites [241] Engreitz JM, Haines JE, Perez EM et al (2016) Local regulation of gene expression by lncRNA promoters, transcription and splicing. Nature 539:452–455 source
• Supported

  The Results section (‘Promoters acting as distal enhancers’) reports that core promoter fragments showed significant enhancer activity in a reporter assay. The Discussion also states that promoters of other genes can act as distal enhancers of POU5F1. (LLM, conf=0.93)

  cites [242] Diao Y, Fang R, Li B et al (2017) A tiling-deletion-based genetic screen for cis-regulatory element identification in mammalian cells. Nat Methods 14:629–635 source
• Supported

  Section 2, “Genome-wide identification of promoters with enhancer activity,” reports that 2–3% of tested human promoters displayed enhancer activity and says the authors termed these elements “Epromoters.” The Introduction also describes promoters that function as enhancers. (LLM, conf=0.94)

  cites [246] Dao LTM, Spicuglia S (2018) Transcriptional regulation by promoters with enhancer function. Transcription 9:307–314 source

In the paper

“Contrary to the initial assumption that enhancers and promoters were biologically distinguishable elements, it is becoming clear that they share many properties (reviewed in [233]).”

What the sources say

• ?Could not verify

  The available passages identify a review about enhancer and promoter activities, but contain no discussion of whether the two share properties. Searches for shared properties, functional similarities, and a blurred distinction returned only bibliographic text, so the claim cannot be verified from our copy. (LLM, conf=0.70)

  cites [233] Andersson R, Sandelin A (2020) Determinants of enhancer and promoter activities of regulatory elements. Nat Rev Genet 21:71–87 sourceExperimental — accuracy not yet measured

In the paper

“These models were initially applied to the mammalian β-globin locus, in which the developmental β-globin genes are regulated by a powerful tissue-specific enhancer known as a locus control region (LCR) [254–257].”

What the sources say

• ?Not checked — no full text of the sourcecites [254] Tuan D, Solomon W, Li Q, London IM (1985) The “beta-like-globin” gene domain in human erythroid cells. Proc Natl Acad Sci USA 82:6384–6388 source
• Supported

  The paper describes the human β-globin genes as developmentally regulated and erythroid-specific (introductory passage, page not provided). Its Discussion identifies an erythroid-specific enhancer activity in the LCR and says that activity is crucial for transcription of the β-globin genes. (LLM, conf=0.93)

  cites [255] Reik A, Telling A, Zitnik G et al (1998) The locus control region is necessary for gene expression in the human beta-globin locus but not the maintenance of an open chromatin structure in erythroid cells. Mol Cell Biol 18:5992–6000 source
• Supported

  The Abstract reports that deleting the LCR severely reduces expression of all mouse β-like globin genes, supporting its role in regulating their expression. The Results clarify that deletion does not alter developmental timing or tissue specificity: the LCR promotes high-level expression but is not required to establish those patterns. (LLM, conf=0.88)

  cites [256] Bender MA, Bulger M, Close J, Groudine M (2000) Beta-globin gene switching and DNase I sensitivity of the endogenous beta-globin locus in mice do not require the locus control region. Mol Cell 5:387–393 source
• Supported

  The introductory passage states that the human β-type globin genes are regulated by the locus control region (LCR), whose components include tissue-specific sites. The same section reports that individual LCR elements confer high-level, tissue-specific transcription and help control developmental gene expression. No page number is provided in our copy. (LLM, conf=0.92)

  cites [257] Bungert J, Davé U, Lim KC et al (1995) Synergistic regulation of human beta-globin gene switching by locus control region elements HS3 and HS4. Genes Dev 9:3083–3096 source

In the paper

“DNA FISH has been used to characterise enhancer–promoter loops for several developmental genes such as Shh and HoxD [303, 304].”

What the sources say

• ?Not checked — no full text of the sourcecites [303] Amano T, Sagai T, Tanabe H et al (2009) Chromosomal dynamics at the Shh locus: limb bud-specific differential regulation of competence and active transcription. Dev Cell 16:47–57 source
• Supported

  The FISH methods section describes the DNA FISH approach. In “Enhancer-promoter looping restricted to the posterior distal region of the limb bud,” the authors report FISH-based proximity and colocalisation of the GCR enhancer region with Hoxd13, consistent with a chromatin loop. This supports the HoxD example attributed to this paper. (LLM, conf=0.93)

  cites [304] Williamson I, Eskeland R, Lettice LA et al (2012) Anterior-posterior differences in HoxD chromatin topology in limb development. Development 139:3157–3167 source

In the paper

“The ENCODE project, for instance, found that an element might be classified as a promoter in the cell type-agnostic setting, but as a proximal enhancer-like element in a specific cell type [131].”

What the sources say

• Supported

  The Extended Data Fig. 3 caption explicitly describes a switch from cell-type-agnostic promoter-like (PLS) classification to cell-type-specific proximal enhancer-like (pELS) classification. The Methods section, ‘Identifying and classifying cCREs,’ also explains that the paper assigns both cell-type-agnostic and biosample-specific classifications. (LLM, conf=0.94)

  cites [131] ENCODE Project Consortium, Moore JE, Purcaro MJ et al (2020) Expanded encyclopaedias of DNA elements in the human and mouse genomes. Nature 583:699–710 source

In the paper

“In another example, a systemic lupus erythematosus-associated variant (termed TT > A) situated within an enhancer downstream of TNFAIP3 has been shown to abrogate NF-kB binding, affecting TNFAIP3 expression [211, 212].”

What the sources say

• Partly supported

  The Abstract identifies the SLE-associated TT>A variant downstream of TNFAIP3 and reports reduced binding by a complex containing NF-κB subunits, along with reduced TNFAIP3 expression on the variant-carrying haplotype. It does not show that NF-κB binding was abrogated: the binding-assay passage reports a difference in binding intensity, while the discussion says the precise mechanism linking the variant to expression remains uncertain. (LLM, conf=0.88)

  cites [211] Adrianto I, Wen F, Templeton A et al (2011) Association of a functional variant downstream of TNFAIP3 with systemic lupus erythematosus. Nat Genet 43:253–258 sourceExperimental — accuracy not yet measured
• Partly supported

  The Abstract identifies TT>A as an SLE-associated variant in a downstream enhancer and reports reduced A20 expression, the protein encoded by TNFAIP3. The Results section, “The TT>A risk alleles bind NF-κB subunits with reduced affinity” (page unavailable), finds impaired NF-κB binding—not that binding is abrogated. The claim overstates that effect. (LLM, conf=0.90)

  cites [212] Wang S, Wen F, Wiley GB et al (2013) An enhancer element harboring variants associated with systemic lupus erythematosus engages the TNFAIP3 promoter to influence A20 expression. PLoS Genet 9:e1003750 sourceExperimental — accuracy not yet measured

In the paper

“Recently, it has been proposed that cis-regulatory elements exist on a spectrum of enhancer/promoter ability [233, 248].”

What the sources say

• ?Could not verify

  The supplied passages and indexed matches contain bibliographic information, not the paper’s discussion of regulatory elements. Searches for an enhancer–promoter spectrum, a continuum of activity, and elements functioning as both enhancers and promoters found no substantive passage, so this copy cannot verify the claim. (LLM, conf=0.74)

  cites [233] Andersson R, Sandelin A (2020) Determinants of enhancer and promoter activities of regulatory elements. Nat Rev Genet 21:71–87 sourceExperimental — accuracy not yet measured
• Supported

  The Discussion describes a continuum of cis-regulatory functions, from elements acting mainly as promoters through elements with both promoter and enhancer activity to elements acting mainly as enhancers. The abstract also explicitly calls this a spectrum of activities. (LLM, conf=0.95)

  cites [248] Mikhaylichenko O, Bondarenko V, Harnett D et al (2018) The degree of enhancer or promoter activity is reflected by the levels and directionality of eRNA transcription. Genes Dev 32:42–57 source

In the paper

“Promoters are also sometimes repurposed into enhancers, but at a 13-fold lower rate than enhancer-to-promoter [100].”

What the sources say

• Partly supported

  The Results section, ‘Enhancers are the main source of regulatory repurposing’ (page not specified), reports promoter-to-enhancer repurposing and an approximately 13-fold enrichment in the opposite direction after accounting for enhancer loss. It does not report a 13-fold difference in the raw repurposing rates: that difference was approximately 26-fold in the primate liver analysis. The comparable enrichment in glires was approximately sixfold and not statistically significant. (LLM, conf=0.91)

  cites [100] Carelli FN, Liechti A, Halbert J et al (2018) Repurposing of promoters and enhancers during mammalian evolution. Nat Commun 9:4066 sourceExperimental — accuracy not yet measured

In the paper

“For instance, eQTL analysis in monocytes stimulated with interferon-γ or lipopolysaccharide has revealed thousands of new variants affecting gene expression, which overlapped nearly 250 variants associated with response to infection and susceptibility to renal and immunological disorders [228].”

What the sources say

• Partly supported

  Split into 2 atoms — 1 confirmed, 1 partially supported, 0 disputed, 0 misattributed, 0 unverifiable. Expand the claim to see per-atom verdicts.

  cites [228] Fairfax BP, Humburg P, Makino S et al (2014) Innate immune activity conditions the effect of regulatory variants upon monocyte gene expression. Science 343:1246949 sourceExperimental — accuracy not yet measured

In the paper

“The “grey area” between enhancers and promoters is supported by evolutionary evidence that some enhancers have undergone genetic mutations allowing them to be repurposed into gene promoters in mammals [100].”

What the sources say

• Supported

  The Introduction reports that putative ancestral enhancers were converted to promoters during mammalian evolution and proposes that inheritable sequence mutations could facilitate this change. The Discussion says the authors traced sequence changes accompanying regulatory repurposing and identifies changes in U1-site distribution as a possible mechanism. (LLM, conf=0.90)

  cites [100] Carelli FN, Liechti A, Halbert J et al (2018) Repurposing of promoters and enhancers during mammalian evolution. Nat Commun 9:4066 source

In the paper

“Likewise, variants associated with type 1 diabetes were strongly enriched at promoter-connected active enhancers in activated, compared with resting, CD4+ T cells [229].”

What the sources say

• Partly supported

  The Results section (‘PCHi-C-facilitated mapping of candidate disease-causal genes’; page unavailable) reports overlap between fine-mapped autoimmune-disease variants and expressed enhancer RNAs at promoter-interacting regions. The paper also compares activated and non-activated CD4+ T cells, but the passages available do not establish that type 1 diabetes variants were strongly enriched at those enhancers in activated cells relative to resting cells. (LLM, conf=0.74)

  cites [229] Burren OS, Rubio García A, Javierre B-M et al (2017) Chromosome contacts in activated T cells identify autoimmune disease candidate genes. Genome Biol 18:165 sourceExperimental — accuracy not yet measured

In the paper

“One key feature determining the position of a regulatory element on this spectrum is likely to be the directionality of its transcriptional output.”

What the sources say

• Supported

  The Abstract says the spectrum of enhancer and promoter activities is highly correlated with transcriptional directionality. The Discussion section 'A subset of promoters can function as developmental enhancers' likewise describes a continuum of regulatory functions that mirrors a continuum of eRNA directionality. The claim's 'likely' is consistent with this evidence. (LLM, conf=0.90)

  cites [248] Mikhaylichenko O, Bondarenko V, Harnett D et al (2018) The degree of enhancer or promoter activity is reflected by the levels and directionality of eRNA transcription. Genes Dev 32:42–57 source

In the paper

“However, knockouts of both PRC1 and PRC2 components affect chromosomal contacts between Polycomb-bound loci [280, 360].”

What the sources say

• Supported

  The paper reports that RING1A/B knockout disrupts the Hox spatial network and reduces contacts within Hox clusters (Abstract; page unavailable). It also reports that selected Hox-cluster interactions remain detectable after knockout of EED, a PRC2 component, but occur at reduced frequencies in some cases (Abstract; page unavailable). Thus both knockouts affect contacts, though the reported PRC2 effect is more limited. (LLM, conf=0.90)

  cites [280] Schoenfelder S, Sugar R, Dimond A et al (2015) Polycomb repressive complex PRC1 spatially constrains the mouse embryonic stem cell genome. Nat Genet 47:1179–1186 source
• Partly supported

  The Results section, “The preferential co-localization of Polycomb-enriched genomic regions is Eed-dependent” (page unavailable), reports that loss of Eed, a PRC2 component, reduces contacts between Polycomb-occupied regions in embryonic stem cells. The passages do not report a knockout of a PRC1 component, so this source supports only the PRC2 part of the claim. (LLM, conf=0.87)

  cites [360] Denholtz M, Bonora G, Chronis C et al (2013) Long-range chromatin contacts in embryonic stem cells reveal a role for pluripotency factors and polycomb proteins in genome organization. Cell Stem Cell 13:602–616 sourceExperimental — accuracy not yet measured

In the paper

“Surprisingly however, deletion or degradation of cohesin or CTCF in differentiated cells has only mild effects on gene expression [344–349].”

What the sources say

• Partly supported

  The Results (page unavailable) report that RAD21 deletion depleted cohesin in noncycling thymocytes and that 1,153 of 17,849 assayed genes were differentially expressed. This supports a limited effect by the proportion of genes affected, but the paper also reports expression changes across the full expression spectrum. It does not test CTCF deletion or degradation, so it cannot support that part of the claim. (LLM, conf=0.84)

  cites [344] Seitan VC, Faure AJ, Zhan Y et al (2013) Cohesin-based chromatin interactions enable regulated gene expression within preexisting architectural compartments. Genome Res 23:2066–2077 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [345] Zuin J, Dixon JR, van der Reijden MIJA et al (2014) Cohesin and CTCF differentially affect chromatin architecture and gene expression in human cells. Proc Natl Acad Sci USA 111:996–1001 source
• Partly supported

  The Discussion (‘TAD Insulation and Transcriptional Regulation’; page unavailable) describes the immediate transcriptional effects of CTCF depletion as ‘rather limited’ and ‘initially mild.’ But the experiments were in mouse embryonic stem cells, not differentiated cells; the Results also report substantially more differentially expressed genes after four days. This paper does not establish the claimed mild gene-expression effects of cohesin loss in differentiated cells. (LLM, conf=0.90)

  cites [346] Nora EP, Goloborodko A, Valton A-L et al (2017) Targeted degradation of CTCF decouples local insulation of chromosome domains from genomic compartmentalization. Cell 169:930–944.e22 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results report that 87% of expressed genes changed in transcription by less than 30% after cohesin degradation, supporting the claim of generally mild effects. The study used HCT-116 colorectal carcinoma cells and did not test CTCF deletion or degradation for its effects on gene expression; it therefore does not establish the full claim about cohesin or CTCF in differentiated cells. Page numbers are not available in our copy. (LLM, conf=0.88)

  cites [347] Rao SSP, Huang S-C, Glenn St Hilaire B et al (2017) Cohesin loss eliminates all loop domains. Cell 171:305-320.e24 sourceExperimental — accuracy not yet measured
• Partly supported

  In “Transcriptional changes upon TAD loss” (page unavailable), the paper reports that Nipbl deletion in differentiated hepatocytes significantly altered expression of about a thousand genes, while the cells’ regulatory potential was mostly unperturbed. This supports a limited overall effect, but the paper does not establish that the expression changes were uniformly mild or report gene-expression effects from CTCF deletion or degradation. (LLM, conf=0.83)

  cites [348] Schwarzer W, Abdennur N, Goloborodko A et al (2017) Two independent modes of chromatin organization revealed by cohesin removal. Nature 551:51–56 sourceExperimental — accuracy not yet measured
• Partly supported

  The Introduction says steady-state gene expression is less affected by architectural-protein depletion, and the Results report that most genes were relatively unperturbed after cohesin depletion. But the Introduction also says cohesin depletion impairs activation of inducible genes, so the unqualified claim about gene expression is broader than the paper supports. The paper’s experiments used HeLa Kyoto cells and protein degradation, not deletion. (LLM, conf=0.83)

  cites [349] Thiecke MJ, Wutz G, Muhar M et al (2020) Cohesin-dependent and -independent mechanisms mediate chromosomal contacts between promoters and enhancers. Cell Rep 32:107929 sourceExperimental — accuracy not yet measured

In the paper

“Recent studies have confirmed its global role in connecting cell type-specific active enhancers and promoters in a range of mammalian cell types including mESCs [295] and neural progenitor cells [365].”

What the sources say

• Supported

  The Results section reports that most YY1-associated interactions in mESCs connect active regulatory elements, including enhancers and promoters. The Discussion describes YY1 as a general regulator of enhancer–promoter interactions across mammalian cell types. The sentence cites a separate source for neural progenitor cells; this paper also mentions that finding as a report from Beagan et al. (2017). (LLM, conf=0.92)

  cites [295] Weintraub AS, Li CH, Zamudio AV et al (2017) YY1 Is a structural regulator of enhancer-promoter loops. Cell 171:1573-1588.e28 source
• Partly supported

  The Results section reports YY1 enrichment at looping NPC-specific enhancers and genes, and loss of several such loops after Yy1 knockdown. But the paper describes YY1 as a candidate architectural protein and does not establish the claimed global role; it also says it cannot conclusively define YY1’s looping role in ES cells. In the citing sentence, the ES-cell example is attached to a different citation. (LLM, conf=0.86)

  cites [365] Beagan JA, Duong MT, Titus KR et al (2017) YY1 and CTCF orchestrate a 3D chromatin looping switch during early neural lineage commitment. Genome Res 27:1139–1152 sourceExperimental — accuracy not yet measured

In the paper

“Moreover, PRC1 was recently found to mediate chromatin interactions between a subset of active developmental enhancers and promoters in Drosophila [361].”

What the sources say

• Supported

  The Results section on enhancer-promoter 3D contacts reports stronger contacts between PRC1-bound active enhancers and promoters in Drosophila eye discs than between matched elements without PRC1 peaks. The Discussion describes these as PRC1-dependent enhancer-promoter contacts, consistent with the citing sentence. (LLM, conf=0.92)

  cites [361] Loubiere V, Papadopoulos GL, Szabo Q et al (2020) Widespread activation of developmental gene expression characterized by PRC1-dependent chromatin looping. Sci Adv 6:eaax4001 source

In the paper

“In contrast, DNA FISH gives access to the full distribution of inter-locus distances across all analysed cells [307].”

What the sources say

• Supported

  The Results section (‘3C and FISH probe different aspects of spatial organization’) says that imaging a labeled pair of loci in a population of cells allows measurement of the probability density function of their spatial distances. This supports access to a distance distribution across the cells analyzed; it does not imply distributions for every possible pair of loci. (LLM, conf=0.92)

  cites [307] Fudenberg G, Imakaev M (2017) FISH-ing for captured contacts: towards reconciling FISH and 3C. Nat Methods 14:673–678 source

In the paper

“and it was found that divergent gene pairs separated by CTCF sites had lower than expected correlation of gene expression genome-wide, confirming the insulatory role of CTCF on the global scale [327].”

What the sources say

• Supported

  The Results (page not provided) report that divergent gene pairs separated by predicted CTCF sites had expression correlations close to those of random gene pairs, unlike pairs without intervening CTCF sites. The authors describe this genome-wide analysis as strong evidence that most predicted CTCF sites function as insulators. (LLM, conf=0.93)

  cites [327] Xie X, Mikkelsen TS, Gnirke A et al (2007) Systematic discovery of regulatory motifs in conserved regions of the human genome, including thousands of CTCF insulator sites. Proc Natl Acad Sci USA 104:7145–7150 source

In the paper

“YY1 and ZNF143 are both zinc finger TFs with DNA binding domains that have recently been implicated as looping factors mediating interactions between active regulatory elements [295, 362–365].”

What the sources say

• Partly supported

  The Introduction and Discussion (page numbers unavailable) identify YY1 as a DNA-binding zinc-finger factor that facilitates enhancer–promoter loops. Our copy does not establish the corresponding claim about ZNF143; a search for ZNF143 returned no relevant passage. (LLM, conf=0.88)

  cites [295] Weintraub AS, Li CH, Zamudio AV et al (2017) YY1 Is a structural regulator of enhancer-promoter loops. Cell 171:1573-1588.e28 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results section identifies ZNF143 as a zinc finger protein enriched at interacting loci, and the Discussion says the findings implicate it in long-range chromatin interactions. The passages do not establish the same claim for YY1, and the Discussion says ZNF143’s specific role in establishing or maintaining chromatin structure remains to be determined. No page numbers were available in our copy. (LLM, conf=0.87)

  cites [362] Heidari N, Phanstiel DH, He C et al (2014) Genome-wide map of regulatory interactions in the human genome. Genome Res 24:1905–1917 sourceExperimental — accuracy not yet measured
• Partly supported

  The Introduction and Discussion identify ZNF143 as a zinc-finger protein that binds a DNA recognition sequence and helps secure interactions between promoters and distal regulatory elements. The cited paper does not establish the corresponding claim about YY1; a search for YY1 found no relevant passage in our copy. Page numbers were not available. (LLM, conf=0.87)

  cites [363] Bailey SD, Zhang X, Desai K et al (2015) ZNF143 provides sequence specificity to secure chromatin interactions at gene promoters. Nat Commun 2:6186 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [364] Ye B-Y, Shen W-L, Wang D et al (2016) ZNF143 is involved in CTCF-mediated chromatin interactions by cooperation with cohesin and other partners. Mol Biol 50:496–503 source
• Partly supported

  The Results section identifies YY1 as a zinc-finger protein enriched at enhancer–gene looping interactions, and the Discussion describes it as a candidate architectural protein. Our copy does not address ZNF143, so it supports the YY1 part of the claim but not the assertion about both factors. (LLM, conf=0.88)

  cites [365] Beagan JA, Duong MT, Titus KR et al (2017) YY1 and CTCF orchestrate a 3D chromatin looping switch during early neural lineage commitment. Genome Res 27:1139–1152 sourceExperimental — accuracy not yet measured

In the paper

“LDB1 was first identified as a looping factor in the mouse β-globin locus [317, 371].”

What the sources say

• ?Not checked — no full text of the sourcecites [317] Song S-H, Hou C, Dean A (2007) A positive role for NLI/Ldb1 in long-range beta-globin locus control region function. Mol Cell 28:810–822 source
• Partly supported

  The Introduction identifies Ldb1 as a factor involved in LCR–β-globin interactions, and the Results show that tethering Ldb1 can produce an LCR–β-globin loop. Our copy does not establish that the mouse β-globin locus was where Ldb1 was first identified as a looping factor; the Introduction cites earlier work on its role there. (LLM, conf=0.83)

  cites [371] Deng W, Lee J, Wang H et al (2012) Controlling long-range genomic interactions at a native locus by targeted tethering of a looping factor. Cell 149:1233–1244 sourceExperimental — accuracy not yet measured

In the paper

“Notably, some contacts between PcG-associated regions, including poised promoters and enhancers in ESCs, are retained after these regions are activated during cell differentiation [141, 143].”

What the sources say

• Partly supported

  The Results section on rewiring and recolouring (Figure 6E–F; page not available) reports that some promoter interactions persist during ESC-to-NEC differentiation, including an IRX3 interaction whose promoter-interacting region changes from repressed to active. The Results section on regulatory elements also reports contacts between poised or Polycomb-associated promoters and similarly marked regions. The copy does not establish that the retained contacts specifically join poised promoters and enhancers that both become activated. (LLM, conf=0.82)

  cites [141] Freire-Pritchett P, Schoenfelder S, Várnai C et al (2017) Global reorganisation of regulatory units upon lineage commitment of human embryonic stem cells. Elife 6:e21926 sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [143] Cruz-Molina S, Respuela P, Tebartz C et al (2017) PRC2 facilitates the regulatory topology required for poised enhancer function during pluripotent stem cell differentiation. Cell Stem Cell 20:689-705.e9 source

In the paper

“Transgene assays of the human β-globin locus then showed that two insulator sites binding CTCF could form a chromatin loop [332].”

What the sources say

• Supported

  The Abstract reports that an extra HS5 copy on a human β-globin transgene formed a new loop with endogenous HS5, and that loop formation depended on an intact CTCF site. The Results and Discussion section confirms CTCF binding at both HS5 sites in the transgene assays. Page numbers are not provided in our copy. (LLM, conf=0.94)

  cites [332] Hou C, Zhao H, Tanimoto K, Dean A (2008) CTCF-dependent enhancer-blocking by alternative chromatin loop formation. Proc Natl Acad Sci U S A 105:20398–20403 source

In the paper

“In mammals, PcG-associated promoters and enhancers also engage in multitudes of looping contacts, particularly in early development [139, 141, 276, 357].”

What the sources say

• Partly supported

  The Results section on PRC2-mediated chromatin interactions reports numerous loops involving promoters and distal regions at developmentally regulated genes in mouse embryonic stem cells (page unavailable). The section on PRC2-associated silencers identifies some distal anchors as having poised-enhancer signatures, but says their enhancer activity can emerge during differentiation. It does not establish that active PcG-associated enhancers generally engage in many loops throughout early mammalian development. (LLM, conf=0.84)

  cites [139] Ngan CY, Wong CH, Tjong H et al (2020) Chromatin interaction analyses elucidate the roles of PRC2-bound silencers in mouse development. Nat Genet 52:264–272 sourceExperimental — accuracy not yet measured
• Supported

  The Results section, “Identification of putative regulatory elements and their associated gene promoters” (page unavailable), reports that Polycomb-associated promoters preferentially contact poised or Polycomb-associated regions. The Discussion section, “A comprehensive atlas of promoter interactions” (page unavailable), describes extensive promoter connectivity to those regions in human pluripotent and early lineage-committed cells, supporting the claim’s early-development context. (LLM, conf=0.86)

  cites [141] Freire-Pritchett P, Schoenfelder S, Várnai C et al (2017) Global reorganisation of regulatory units upon lineage commitment of human embryonic stem cells. Elife 6:e21926 source
• Partly supported

  The Results section ('Transcription factors mediate chromatin architecture within TADs'; page unavailable) reports that Polycomb-associated, H3K27me3-rich regions form nested sets of chromatin contacts, supporting the claim’s general point about multiple contacts. It does not establish that the interacting regions are specifically PcG-associated promoters and enhancers, or that these contacts occur particularly in early development. (LLM, conf=0.84)

  cites [276] Hsieh T-HS, Cattoglio C, Slobodyanyuk E et al (2020) Resolving the 3D landscape of transcription-linked mammalian chromatin folding. Mol Cell 78:539-553.e8 sourceExperimental — accuracy not yet measured
• Partly supported

  The Results (‘Cohesin-Independent Chromosomal Interactions Exist in ESCs’; page unavailable) report numerous Polycomb-associated interactions in embryonic stem cells, but say the interacting domains were mostly associated with promoters. They note that Polycomb domains can also be associated with enhancers without establishing that PcG-associated enhancers engage in multitudes of looping contacts. The cell-type-specific Results further find much weaker or no obvious interactions in the other mammalian cell types examined, so the broad mammalian wording exceeds this evidence. (LLM, conf=0.85)

  cites [357] Rhodes JDP, Feldmann A, Hernández-Rodríguez B et al (2020) Cohesin disrupts polycomb-dependent chromosome interactions in embryonic stem cells. Cell Rep 30:820-835.e10 sourceExperimental — accuracy not yet measured

In the paper

“More unusually, it emerged that CTCF binding has an insulating capacity, effectively blocking signals between enhancers and their target genes in the β-globin and H19/Igf2 loci [323–326].”

What the sources say

• ?Not checked — no full text of the sourcecites [323] Bell AC, West AG, Felsenfeld G (1999) The protein CTCF is required for the enhancer blocking activity of vertebrate insulators. Cell 98:387–396 source
• Supported

  The Discussion reports that CTCF-binding elements at both chicken β-globin boundaries block enhancer action. Figure 6 states that the 3′ CTCF-binding site is necessary and sufficient for enhancer-blocking activity. This supports the β-globin portion of the sentence; the paper is not offered as evidence for the H19/Igf2 portion. (LLM, conf=0.90)

  cites [324] Saitoh N, Bell AC, Recillas-Targa F et al (2000) Structural and functional conservation at the boundaries of the chicken beta-globin domain. EMBO J 19:2315–2322 source
• ?Not checked — no full text of the sourcecites [325] Hark AT, Schoenherr CJ, Katz DJ et al (2000) CTCF mediates methylation-sensitive enhancer-blocking activity at the H19/Igf2 locus. Nature 405:486–489 source
• ?Not checked — no full text of the sourcecites [326] Kanduri C, Pant V, Loukinov D et al (2000) Functional association of CTCF with the insulator upstream of the H19 gene is parent of origin-specific and methylation-sensitive. Curr Biol 10:853–856 source

In the paper

“A key insight into how CTCF mediates chromosomal looping was the discovery of its association with the cohesin complex [132, 334, 335].”

What the sources say

• ?Not checked — no full text of the sourcecites [132] Parelho V, Hadjur S, Spivakov M et al (2008) Cohesins functionally associate with CTCF on mammalian chromosome arms. Cell 132:422–433 source
• Supported

  The paper states that the cohesin subunit Scc3/SA1 is an interacting partner of CTCF, and the Discussion explicitly describes the finding as CTCF interacting with cohesin. Page numbers are not supplied in our copy. (LLM, conf=0.94)

  cites [334] Rubio ED, Reiss DJ, Welcsh PL et al (2008) CTCF physically links cohesin to chromatin. Proc Natl Acad Sci USA 105:8309–8314 source
• ?Not checked — no full text of the sourcecites [335] Wendt KS, Yoshida K, Itoh T et al (2008) Cohesin mediates transcriptional insulation by CCCTC-binding factor. Nature 451:796–801 source

In the paper

“Following the discovery of chromatin looping in the β-globin locus, it was ascertained that several TFs were required for looping between the LCR and the distal genes, including GATA-1, FOG-1, EKLF and NLI/Ldb1 [315–317].”

What the sources say

• Supported

  The Abstract states that GATA-1 and FOG-1 are required for physical interaction between the LCR and the β-major globin promoter. That supports this paper’s contribution to the sentence’s multi-citation list. (LLM, conf=0.92)

  cites [315] Vakoc CR, Letting DL, Gheldof N et al (2005) Proximity among distant regulatory elements at the beta-globin locus requires GATA-1 and FOG-1. Mol Cell 17:453–462 source
• Supported

  This paper supports the EKLF part of the claim. In Results and Discussion (page unavailable), EKLF knockout strongly reduces interactions between an LCR element and the βmaj promoter, and the authors conclude that EKLF is necessary for their participation in the active chromatin hub. The other named factors are not the part of this multi-citation sentence supported by this paper. (LLM, conf=0.90)

  cites [316] Drissen R, Palstra R-J, Gillemans N et al (2004) The active spatial organization of the beta-globin locus requires the transcription factor EKLF. Genes Dev 18:2485–2490 source
• ?Not checked — no full text of the sourcecites [317] Song S-H, Hou C, Dean A (2007) A positive role for NLI/Ldb1 in long-range beta-globin locus control region function. Mol Cell 28:810–822 source

In the paper

“Forced looping between β-globin and its LCR was shown to rescue β-globin expression in K562 erythroid cells (which instead aberrantly express γ-globin) in a reversible manner [313].”

What the sources say

• Supported

  The Representative Results report that K562 cells express high levels of γ-globin rather than β-globin, and that forced β-globin promoter–LCR looping increased β-globin expression. The Figure 2 caption and accompanying results describe this induction as reversible after short-term treatment. The same section notes that expression remained elevated after longer-term treatment and washout, so reversibility is demonstrated for the short-term condition. (LLM, conf=0.91)

  cites [313] Morgan SL, Chang EY, Mariano NC et al (2018) CRISPR-mediated reorganization of chromatin loop structure. J Vis Exp. https://doi.org/10.3791/57457 source

In the paper

“However, Promoter Capture Hi-C-based studies did not observe a significant change in enhancer–promoter contacts upon RNAP inhibition, including in cells rapidly depleted of cohesin or CTCF [349, 380].”

What the sources say

• Partly supported

  The Results section on cohesin-dependent and -independent promoter interactions reports that the RNA polymerase II inhibitor triptolide had only a mild effect on promoter interaction profiles. It does not say the effect was statistically insignificant or report that inhibition was tested in cells simultaneously depleted of cohesin or CTCF. A separate Results section reports substantial rewiring of contacts after architectural-protein depletion alone. Page numbers are unavailable in our copy. (LLM, conf=0.86)

  cites [349] Thiecke MJ, Wutz G, Muhar M et al (2020) Cohesin-dependent and -independent mechanisms mediate chromosomal contacts between promoters and enhancers. Cell Rep 32:107929 sourceExperimental — accuracy not yet measured
• Partly supported

  In Results, ‘The role of Mediator in promoter-enhancer interactions’ (page unavailable), promoter capture Hi-C showed little or no change in enhancer–promoter contacts after PolII and Mediator depletion, and no further change when they were depleted alongside cohesin. The paper reports rapid cohesin depletion, but not rapid CTCF depletion; loss of CTCF-dependent loops after cohesin depletion is not the same experiment. (LLM, conf=0.87)

  cites [380] El Khattabi L, Zhao H, Kalchschmidt J et al (2019) A pliable mediator acts as a functional rather than an architectural bridge between promoters and enhancers. Cell 178:1145-1158.e20 sourceExperimental — accuracy not yet measured

In the paper

“For example, transcription at stimulus–response genes was shown to disrupt chromatin loops [387, 388], potentially through destabilising cohesin association with the chromatin fibre by RNAP machinery [388].”

What the sources say

• Partly supported

  The Results report chromatin decompaction at glucocorticoid-responsive loci, but the Discussion says dissolution of preformed loops cannot be excluded—not that transcription was shown to disrupt loops. The Results and Discussion also say decompaction can occur without transcription. Our copy does not identify destabilisation of cohesin by RNAP machinery as a mechanism (searched: cohesin). (LLM, conf=0.88)

  cites [387] Jubb AW, Boyle S, Hume DA, Bickmore WA (2017) Glucocorticoid receptor binding induces rapid and prolonged large-scale chromatin decompaction at multiple target loci. Cell Rep 21:3022–3031 sourceExperimental — accuracy not yet measured
• Supported

  The Results section, “Genic transcription affects 3D genome structure by eliminating cohesin-mediated loops,” reports that stimulus-induced transcription weakened CTCF-anchored loops at target genes. The Discussion section, “Elongating RNAPII disrupts 3D structure by displacing cohesin and CTCF,” attributes loop dissociation to elongating RNAPII displacing cohesin, supporting the proposed mechanism. (LLM, conf=0.94)

  cites [388] Heinz S, Texari L, Hayes MGB et al (2018) Transcription elongation can affect genome 3D structure. Cell 174:1522-1536.e22 source

In the paper

“For example, neither Hi-C nor Micro-C detected a widespread enrichment for enhancer–promoter interactions in early Drosophila embryos irrespective of the activity of these elements [393].”

What the sources say

• Supported

  The Discussion reports no evidence for widespread enrichment of enhancer–promoter interactions, including in tissues where the enhancers are active. The Results report no significant increase in Hi-C or Micro-C interaction frequency when the enhancers are active in the studied Drosophila embryos. (LLM, conf=0.94)

  cites [393] Ing-Simmons E, Vaid R, Bing XY et al (2021) Independence of chromatin conformation and gene regulation during Drosophila dorsoventral patterning. Nat Genet 53:487–499 source

In the paper

“Whilst it is established that transcription helps shape up higher-level genome conformation (reviewed in [386]), there is also evidence that it can either disrupt or stabilise specific enhancer–promoter loops.”

What the sources say

• Supported

  The citation is attached to the statement that transcription helps shape higher-level genome conformation, not to the subsequent claim about specific enhancer–promoter loops. The review’s section on transcription perturbation studies reports that active transcription helps maintain TAD organisation in Drosophila; its section on chromatin compartments also describes transcription-dependent relocation of a locus. (LLM, conf=0.91)

  cites [386] van Steensel B, Furlong EEM (2019) The role of transcription in shaping the spatial organization of the genome. Nat Rev Mol Cell Biol 20:327–337 source

In the paper

“Nonetheless, removing BRD4 (and, consequently, Mediator) from chromatin using chemical inhibitors did not perturb the majority of enhancer–promoter contacts, despite major changes in transcription [69].”

What the sources say

• Partly supported

  The Introduction and Results report major transcriptional changes while enhancer–promoter contacts remained largely intact after BET inhibitor treatment. However, the Discussion says the drugs reduced BRD4 and Mediator binding but left detectable binding, so describing both proteins as removed from chromatin overstates the treatment’s effect. (LLM, conf=0.86)

  cites [69] Crump NT, Ballabio E, Godfrey L et al (2021) BET inhibition disrupts transcription but retains enhancer–promoter contact. Nat Commun 12:223 sourceExperimental — accuracy not yet measured

In the paper

“Whilst LLPS in general has been demonstrated both in vitro and in vivo [421, 422], evidence for chromatin LLPS and its functional significance in gene control is still incomplete.”

What the sources say

• Supported

  The citation is attached to the first clause, which says LLPS has been demonstrated in vitro and in vivo; the statement about incomplete evidence for chromatin LLPS is not attributed to this source. The Results section reports Ddx4 phase separation in live cells and comparable droplets formed in vitro. (LLM, conf=0.94)

  cites [421] Nott TJ, Petsalaki E, Farber P et al (2015) Phase transition of a disordered nuage protein generates environmentally responsive membraneless organelles. Mol Cell 57:936–947 source
• Supported

  The citation is attached to the statement that LLPS has been demonstrated in vitro and in vivo, not to the citing authors’ assessment of chromatin LLPS. The Results report LAF-1 droplet formation in vitro and P-granule dissolution after LAF-1 depletion in embryos, which the paper interprets as evidence that LAF-1 promotes LLPS in vivo. The paper does not address chromatin LLPS or gene control. (LLM, conf=0.87)

  cites [422] Elbaum-Garfinkle S, Kim Y, Szczepaniak K et al (2015) The disordered P granule protein LAF-1 drives phase separation into droplets with tunable viscosity and dynamics. Proc Natl Acad Sci USA 112:7189–7194 source

In the paper

“However, it was shown recently that rapid removal of chromatin-bound Mediator and RNAP does not significantly affect chromosomal architecture, including enhancer–promoter interactions, in spite of triggering widespread transcriptional changes and cell cycle arrest [380].”

What the sources say

• Supported

  In Results, “The role of Mediator in promoter-enhancer interactions” (page unavailable), the paper reports rapid depletion of chromatin-bound PolII and Mediator with cell-cycle arrest, while promoter–enhancer contacts showed little or no change. Results, “Functional analysis of mMED core subunits” (page unavailable), reports a global, approximately sevenfold transcriptome downregulation after MED14 depletion. The transcription measurement used a slower depletion protocol than the rapid-depletion contact experiment. (LLM, conf=0.88)

  cites [380] El Khattabi L, Zhao H, Kalchschmidt J et al (2019) A pliable mediator acts as a functional rather than an architectural bridge between promoters and enhancers. Cell 178:1145-1158.e20 source

In the paper

“an explanation has eventually been provided by the model of chromosomal loop extrusion [342, 435].”

What the sources say

• ?Not checked — no full text of the sourcecites [342] Sanborn AL, Rao SSP, Huang S-C et al (2015) Chromatin extrusion explains key features of loop and domain formation in wild-type and engineered genomes. Proc Natl Acad Sci USA 112:E6456–E6465 source
• Supported

  The Introduction says the authors propose loop extrusion as a mechanism for TAD formation and that their simulations produce TADs consistent with Hi-C data. The Discussion likewise describes the model as forming TADs and explaining observed features. This supports the citing sentence’s attribution of an explanation to the model. (LLM, conf=0.93)

  cites [435] Fudenberg G, Imakaev M, Lu C et al (2016) Formation of chromosomal domains by loop extrusion. Cell Rep 15:2038–2049 source

In the paper

“In contrast, a proximal enhancer of the Shh gene moves further away from the Shh promoter upon activation, according to both 3C-derived methods and imaging [407].”

What the sources say

• Supported

  The Results report that Shh activation increased the measured distance between Shh and SBE6, an enhancer 100 kb upstream, in 3D-FISH imaging. The Discussion states that analyses using both FISH and 5C found decreased enhancer–promoter proximity in Shh-expressing cells. (LLM, conf=0.93)

  cites [407] Benabdallah NS, Williamson I, Illingworth RS et al (2019) Decreased enhancer-promoter proximity accompanying enhancer activation. Mol Cell 76:473-484.e7 source