Transcription factors as readers and effectors of DNA methylation

In the paper

“of the 15,557 ChIP–seq peaks identified for CEBPβ, 6,675 (42.9%) had a methylation level greater than 80%.”

In the paper

“Overall, 77,124 (29.9%) of the 258,188 DHSs detected in H1 cells had an average methylation level greater than 80% at CpG sites (FIG. 2a)”

In the paper

“Based on the analysis of KLF4 binding in ES cells, we also found that KLF4 binds to methylated enhancer regions53”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“A motif could be reconstructed with these 3,894 binding peaks, which represented the methylated motif for CEBPβ (FIG. 2c).”

In the paper

“Among the 6,675 peaks with a high methylation level, 3,894 carried a highly methylated (>80%) CpG site within the enriched motifs.”

In the paper

“indicating that a substantial fraction of their binding sites are located in highly methylated regions (FIG. 2a).”

In the paper

“The most significantly enriched motif did not contain a CpG site.”

In the paper

“The DNA methylation levels within the ChIP–seq peaks showed a bimodal distribution for all TFs except RXRα”

In the paper

“when the same analysis was applied to those peaks that have high methylation levels, a significantly enriched motif containing a CpG site at position 4 was discovered (FIG. 2b).”

In the paper

“Indeed, we observed a large overlap between known pioneer TFs and proteins that bind to methylated DNA (for example, forkhead box protein A (FOXA) and GATA families, which are the best-studied pioneer factors)106,108–110.”

What the sources say

• Not checked yetcites [106] Zaret, K. S. & Carroll, J. S. Pioneer transcription factors: establishing competence for gene expression. Genes Dev. 25, 2227–2241 (2011). source
• Not checked yetcites [108] Bossard, P. & Zaret, K. S. GATA transcription factors as potentiators of gut endoderm differentiation. Development 125, 4909–4917 (1998). source
• Not checked yetcites [109] Laverriere, A. C. et al. GATA-4/5/6, a subfamily of three transcription factors transcribed in developing heart and gut. J. Biol. Chem. 269, 23177–23184 (1994). source
• Not checked yetcites [110] Liu, J. K., DiPersio, C. M. & Zaret, K. S. Extracellular signals that regulate liver transcription factors during hepatic differentiation in vitro. Mol. Cell. Biol. 11, 773–784 (1991). source

In the paper

“recent large-scale analyses of gene expression profiles and DNA methylomes showed that a substantial portion of DNA methylation sites is positively correlated with gene expression31”

• Also stated in: § Activation or repression · plain

  “large-scale analyses of gene expression profiles and DNA methylomes have revealed that a substantial proportion of DNA methylation sites is positively correlated with gene expression31”

What the sources say

• Not checked yetcites [31] Irizarry, R. A. et al. The human colon cancer methylome shows similar hypo- and hypermethylation at conserved tissue-specific CpG island shores. Nat. Genet. 41, 178–186 (2009). source

In the paper

“the transcriptional regulator Kaiso, which contains POZ (pox virus and zinc-finger) and zinc-finger domains, was found to bind to a specific methylated sequence with its C2H2 zinc-finger domains41”

What the sources say

• Not checked yetcites [41] Prokhortchouk, A. et al. The p120 catenin partner Kaiso is a DNA methylation-dependent transcriptional repressor. Genes Dev. 15, 1613–1618 (2001). source

In the paper

“the nuclear protein MVBP (methylated VBE-binding protein) was shown to bind to a rice tungro bacilliform virus promoter region only when the promoter was methylated46”

What the sources say

• Not checked yetcites [46] He, X., Futterer, J. & Hohn, T. Sequence-specific and methylation-dependent and -independent binding of rice nuclear proteins to a rice tungro bacilliform virus vascular bundle expression element. J. Biol. Chem. 276, 2644–2651 (2001). source

In the paper

“evidence is emerging to suggest that transcription factors lacking a MBD can also interact with methylated DNA”

• Also stated in: § Readers of methylated DNA · hedged

  “Over the past 15 years, evidence has emerged that suggests that some TFs lacking MBDs are able to interact with methylated DNA23–27”

What the sources say

• Not checked yetcites [23] Li, M. et al. Sensitive digital quantification of DNA methylation in clinical samples. Nat. Biotechnol. 27, 858–863 (2009). source
• Not checked yetcites [24] Gavin, D. P. & Sharma, R. P. Histone modifications, DNA methylation, and schizophrenia. Neurosci. Biobehav. Rev. 34, 882–888 (2010). source
• Not checked yetcites [25] Jiang, Y. H. et al. A mixed epigenetic/genetic model for oligogenic inheritance of autism with a limited role for UBE3A. Am. J. Med. Genet. A 131A, 1–10 (2004). source
• Not checked yetcites [26] Nagarajan, R. P., Hogart, A. R., Gwye, Y., Martin, M. R. & LaSalle, J. M. Reduced MeCP2 expression is frequent in autism frontal cortex and correlates with aberrant MECP2 promoter methylation. Epigenetics 1, e1–e11 (2006). source
• Not checked yetcites [27] Lister, R. et al. Global epigenomic reconfiguration during mammalian brain development. Science 341, 1237905 (2013). source

In the paper

“Two amino acids, one cysteine and one serine, were found to interact with 5mC45.”

What the sources say

• Not checked yetcites [45] Karlsson, Q. H., Schelcher, C., Verrall, E., Petosa, C. & Sinclair, A. J. Methylated DNA recognition during the reversal of epigenetic silencing is regulated by cysteine and serine residues in the Epstein-Barr virus lytic switch protein. PLoS Pathog. 4, e1000005 (2008). source

In the paper

“although most of the evidence showing a positive correlation between methylation and expression seems to result from methylation downstream of the transcription start site.”

In the paper

“DNA methylation of a promoter or an enhancer has been shown to be correlated with increased transcription of a target gene47,49”

What the sources say

• Not checked yetcites [47] Bahar Halpern, K., Vana, T. & Walker, M. D. Paradoxical role of DNA methylation in activation of FoxA2 gene expression during endoderm development. J. Biol. Chem. 289, 23882–23892 (2014). source
• Not checked yetcites [49] Niesen, M. I. et al. Activation of a methylated promoter mediated by a sequence-specific DNA-binding protein, RFX. J. Biol. Chem. 280, 38914–38922 (2005). source

In the paper

“19 proteins were identified that interact preferentially with the methylated DNA probe rather than the non-methylated counterpart in mouse embryonic stem (ES) cell nuclear extracts.”

What the sources say

• Not checked yetcites [51] Spruijt, C. G. et al. Dynamic readers for 5-(hydroxy)methylcytosine and its oxidized derivatives. Cell 152, 1146–1159 (2013). This paper describes the identification of proteins that interact with mCpG sites, 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) in ES cells and neuronal progenitor cells using a MS/MS-based approach. source

In the paper

“Unlike MBD proteins, a handful of mammalian TFs were found to possess sequence-dependent mCpG-binding activity in a few studies.”

In the paper

“In total, 41 TFs and 6 cofactors were found to bind to at least one methylated sequence.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“this study suggests that CTCF binding itself is necessary to maintain differential methylation of ICRs.”

What the sources say

• Not checked yetcites [82] Schoenherr, C. J., Levorse, J. M. & Tilghman, S. M. CTCF maintains differential methylation at the Igf2/H19 locus. Nat. Genet. 33, 66–69 (2003). source

In the paper

“the factors that showed binding activity to methylated sequences were widespread among various TF subfamilies, such as zf-C2H2, homeobox, bHLH (basic helix–loop–helix), forkhead, bZIP and HMG (high-mobility group) box.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“the presence of a bulky methyl group in the major groove leads to a subtle widening of the major groove and a subtle narrowing of the minor groove.”

What the sources say

• Not checked yetcites [68] Liu, Y. et al. Structural basis for Klf4 recognition of methylated DNA. Nucleic Acids Res. 42, 4859–4867 (2014). This study determined the crystal structure of the KLF4-methylated DNA complex and provided the structural basis for mCpG–TF interactions. source
• Not checked yetcites [72] Buck-Koehntop, B. A. et al. Molecular basis for recognition of methylated and specific DNA sequences by the zinc finger protein Kaiso. Proc. Natl Acad. Sci. USA 109, 15229–15234 (2012). source
• Not checked yetcites [73] Tippin, D. B. & Sundaralingam, M. Nine polymorphic crystal structures of d(CCGGGCCCGG), d(CCGGGCCm5CGG), d(Cm5CGGGCCm5CGG) and d(CCGGGCC(Br)5CGG) in three different conformations: effects of spermine binding and methylation on the bending and condensation of A-DNA. J. Mol. Biol. 267, 1171–1185 (1997). source

In the paper

“suggesting that many methylated CpG sites are accessible to TFs.”

In the paper

“regions with a low level of methylation, ranging from 10% to 50%, often occur at distal regulatory regions78; that is, regions that are enriched for enhancer marks, including high levels of histone H3 lysine 4 monomethylation (H3K4me1) as well as binding sites for p300 histone acetyltransferase and other regulatory factors.”

What the sources say

• Not checked yetcites [78] Stadler, M. B. et al. DNA-binding factors shape the mouse methylome at distal regulatory regions. Nature 480, 490–495 (2011). This paper demonstrates that some proteins, such as CTCF and REST, can reduce DNA methylation levels at the genomic regions near their binding regions. source

In the paper

“If the methylation status of these two sequences is exchanged, KLF4 loses the ability to bind to either sequence53.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“This competition assay ensures that the identified interactions are indeed methylation-dependent, rather than due to CpG-flanking sequences.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“a different study discovered that Kaiso was not associated with highly methylated promoters in GM12878 lympho-blastoid cells or in K562 human myeloid leukaemia cell lines64.”

What the sources say

• Not checked yetcites [64] Blattler, A. et al. ZBTB33 binds unmethylated regions of the genome associated with actively expressed genes. Epigenetics Chromatin 6, 13 (2013). source

In the paper

“ZFP57 and its cofactor KAP1 were shown to bind selectively to nine DNA-methylated alleles of imprinting control regions (ICRs) in ES cells43.”

What the sources say

• Not checked yetcites [43] Quenneville, S. et al. In embryonic stem cells, ZFP57/KAP1 recognize a methylated hexanucleotide to affect chromatin and DNA methylation of imprinting control regions. Mol. Cell 44, 361–372 (2011). This paper demonstrates that ZFP57 and its cofactor KAP1 affect chromatin by interacting with methylated ICRs in embryonic stem cells. source

In the paper

“Kaiso was shown to bind to the methylated promoter of the MTA2 gene in HeLa cells61.”

What the sources say

• Not checked yetcites [61] Yoon, H. G., Chan, D. W., Reynolds, A. B., Qin, J. & Wong, J. N-CoR mediates DNA methylation-dependent repression through a methyl CpG binding protein Kaiso. Mol. Cell 12, 723–734 (2003). source

In the paper

“CTCF binds to the unmethylated ICRs in maternal alleles, which prevents distal enhancers from activating downstream genes81.”

What the sources say

• Not checked yetcites [81] Bell, A. C. & Felsenfeld, G. Methylation of a CTCF-dependent boundary controls imprinted expression of the Igf2 gene. Nature 405, 482–485 (2000). source

In the paper

“the above analysis suggests that many TFs shown to bind methylated DNA in vitro are also able to interact with methylated DNA in vivo”

In the paper

“Kaiso binds to the methylated promoters of CDKN2A, MGMT and HIC1 in both HCT116 and Colo320 human colon cancer cell lines62.”

What the sources say

• Not checked yetcites [62] Lopes, E. C. et al. Kaiso contributes to DNA methylation-dependent silencing of tumor suppressor genes in colon cancer cell lines. Cancer Res. 68, 7258–7263 (2008). source

In the paper

“a few proteins such as KLF4 and homeobox A5 (HOXA5) were shown to bind mCpG sites in both species”

What the sources say

• Not checked yetcites [51] Spruijt, C. G. et al. Dynamic readers for 5-(hydroxy)methylcytosine and its oxidized derivatives. Cell 152, 1146–1159 (2013). This paper describes the identification of proteins that interact with mCpG sites, 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) in ES cells and neuronal progenitor cells using a MS/MS-based approach. source
• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“which may suggest that sequence-specific mCpG-binding proteins interact preferentially with distal enhancer regions.”

In the paper

“ZFP57 binding sites were mapped in hybrid ES cells, and ZFP57 was found to interact with the methylated parental-origin allele60.”

What the sources say

• Not checked yetcites [60] Strogantsev, R. et al. Allele-specific binding of ZFP57 in the epigenetic regulation of imprinted and non-imprinted monoallelic expression. Genome Biol. 16, 112 (2015). source

In the paper

“many TFs (such as MHC class II regulatory factor RFX1, zinc-finger homeobox 3 (ZFHX3), lysine-specific histone demethylase 1A (LSD1), zinc-finger and BTB domain-containing protein 44 (ZBT44) and thymocyte nuclear protein 1 (THYN1; also known as THY28), and the Krüppel-like factors (for example, KLF2, KLF4 and KLF5)), were identified as new mCpG-binding proteins (TABLE 2).”

What the sources say

• Not checked yetcites [51] Spruijt, C. G. et al. Dynamic readers for 5-(hydroxy)methylcytosine and its oxidized derivatives. Cell 152, 1146–1159 (2013). This paper describes the identification of proteins that interact with mCpG sites, 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) in ES cells and neuronal progenitor cells using a MS/MS-based approach. source

In the paper

“A global survey of methylated-DNA-binding proteins suggests that many other protein domains might also be able to interact with mCpG sites, including homeobox, HLH and E2F domains53.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“TFs identified using the other methods may not necessarily recognize mCpGs in vivo, and further studies are needed to dissect the functionality of the interactions.”

In the paper

“Consequently, 5mC can affect the access of a given TF to the affected motifs in both major and minor grooves in genomic DNA and thus change the sequence specificity of protein–DNA interactions.”

In the paper

“for many newly discovered methylated DNA-binding proteins, the methylated motifs differ from the non-methylated motifs53.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“extensive DNA methylation was found to coexist with active H3K27 acetylation (H3K27ac) marks in a large number of enhancers79.”

What the sources say

• Not checked yetcites [79] Charlet, J. et al. Bivalent regions of cytosine methylation and H3K27 acetylation suggest an active role for DNA methylation at enhancers. Mol. Cell 62, 422–431 (2016). source

In the paper

“Of the 154 methylated motifs examined, 150 showed strong binding signals to at least one protein on the microarray.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“Among the eight bZIP proteins, CEBPα and CEBPβ were found to specifically bind to a methylated sequence56.”

What the sources say

• Not checked yetcites [56] Mann, I. K. et al. CG methylated microarrays identify a novel methylated sequence bound by the CEBPB|ATF4 heterodimer that is active in vivo. Genome Res. 23, 988–997 (2013). This paper describes the use of DNA microarrays to identify proteins that interact with methylated DNA. source

In the paper

“A structural comparison of MeCP2 and KLF4 indeed showed a common structural feature involving one arginine and one asparagine53.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“Both in vitro DNase I digestion assays and structural studies indicate that methylation has a profound impact on DNA structure and shape”

What the sources say

• Not checked yetcites [69] Dantas Machado, A. C. et al. Evolving insights on how cytosine methylation affects protein–DNA binding. Brief. Funct. Genom. 14, 61–73 (2015). source

In the paper

“suggesting that MBD-lacking TFs bind to methylated DNA motifs nearly as strongly as MBD2b.”

In the paper

“The deduced Kd values of ZMYM3, AP2α and KLF4 were determined as 460 nM, 399 nM and 479 nM, respectively.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“Note that this is a conservative estimate because we used a stringent definition of highly methylated sites (that is, >80%).”

In the paper

“MBD family proteins tend to bind to regions with a high methylation density (that is, high methylation level and high CpG density)76”

What the sources say

• Not checked yetcites [76] Baubec, T., Ivanek, R., Lienert, F. & Schubeler, D. Methylation-dependent and -independent genomic targeting principles of the MBD protein family. Cell 153, 480–492 (2013). source

In the paper

“it suggests that methylation-dependent TF–DNA interactions may be cell type-specific.”

In the paper

“Adding a methyl group to the cytosine could affect the local DNA shape, as evidenced by the altered DNase I digestion rate and patterns70,71.”

What the sources say

• Not checked yetcites [70] He, H. H. et al. Refined DNase-seq protocol and data analysis reveals intrinsic bias in transcription factor footprint identification. Nat. Methods 11, 73–78 (2014). source
• Not checked yetcites [71] Lazarovici, A. et al. Probing DNA shape and methylation state on a genomic scale with DNase I. Proc. Natl Acad. Sci. USA 110, 6376–6381 (2013). source

In the paper

“no obvious affinity could be detected when these tested motifs were unmethylated.”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“ChIP–BS-seq is the only approach that does not use naked DNA fragments to identify the TFs that bind to methylated DNA.”

In the paper

“Specifically, hypermethylation is often associated with a depletion of TF binding.”

In the paper

“Single-gene studies have also demonstrated that DNA methylation can activate gene expression47–49.”

What the sources say

• Not checked yetcites [47] Bahar Halpern, K., Vana, T. & Walker, M. D. Paradoxical role of DNA methylation in activation of FoxA2 gene expression during endoderm development. J. Biol. Chem. 289, 23882–23892 (2014). source
• Not checked yetcites [48] Hantusch, B., Kalt, R., Krieger, S., Puri, C. & Kerjaschki, D. Sp1/Sp3 and DNA-methylation contribute to basal transcriptional activation of human podoplanin in MG63 versus Saos-2 osteoblastic cells. BMC Mol. Biol. 8, 20 (2007). source
• Not checked yetcites [49] Niesen, M. I. et al. Activation of a methylated promoter mediated by a sequence-specific DNA-binding protein, RFX. J. Biol. Chem. 280, 38914–38922 (2005). source

In the paper

“The manipulation of the methylation status of DNA sequences has been shown (mostly in in vitro studies) to result in the differential binding of TFs, including E2F, AP2α, MYC and MYN88–95.”

What the sources say

• Not checked yetcites [88] Bednarik, D. P. et al. DNA CpG methylation inhibits binding of NF-kappa B proteins to the HIV-1 long terminal repeat cognate DNA motifs. New Biol. 3, 969–976 (1991).
• Not checked yetcites [89] Comb, M. & Goodman, H. M. CpG methylation inhibits proenkephalin gene expression and binding of the transcription factor AP-2. Nucleic Acids Res. 18, 3975–3982 (1990). source
• Not checked yetcites [90] Ehrlich, K. C., Cary, J. W. & Ehrlich, M. A broad bean cDNA clone encoding a DNA-binding protein resembling mammalian CREB in its sequence specificity and DNA methylation sensitivity. Gene 117, 169–178 (1992). source
• Not checked yetcites [91] Falzon, M. & Kuff, E. L. Binding of the transcription factor EBP-80 mediates the methylation response of an intracisternal A-particle long terminal repeat promoter. Mol. Cell. Biol. 11, 117–125 (1991). source
• Not checked yetcites [92] Iguchi-Ariga, S. M. & Schaffner, W. CpG methylation of the cAMP-responsive enhancer/promoter sequence TGACGTCA abolishes specific factor binding as well as transcriptional activation. Genes Dev. 3, 612–619 (1989). source
• Not checked yetcites [93] Inamdar, N. M., Ehrlich, K. C. & Ehrlich, M. CpG methylation inhibits binding of several sequence-specific DNA-binding proteins from pea, wheat, soybean and cauliflower. Plant Mol. Biol. 17, 111–123 (1991). source
• Not checked yetcites [94] Kovesdi, I., Reichel, R. & Nevins, J. R. Role of an adenovirus E2 promoter binding factor in E1A-mediated coordinate gene control. Proc. Natl Acad. Sci. USA 84, 2180–2184 (1987). source
• Not checked yetcites [95] Prendergast, G. C., Lawe, D. & Ziff, E. B. Association of Myn, the murine homolog of max, with c-Myc stimulates methylation-sensitive DNA binding and ras cotransformation. Cell 65, 395–407 (1991). source

In the paper

“Of note, these two mechanisms are not mutually exclusive.”

In the paper

“which means that DNA modification could influence RNA splicing.”

In the paper

“when the sequences of binding motifs were altered, overall methylation levels decreased83.”

What the sources say

• Not checked yetcites [83] Krebs, A. R., Dessus-Babus, S., Burger, L. & Schubeler, D. High-throughput engineering of a mammalian genome reveals building principles of methylation states at CG rich regions. eLife 3, e04094 (2014). source

In the paper

“pioneer TFs must possess special features to enable protein–DNA interactions.”

In the paper

“A large number of novel binding sites for the TF NRF1 were created as a result of the triple knockout (TKO).”

What the sources say

• Not checked yetcites [97] Domcke, S. et al. Competition between DNA methylation and transcription factors determines binding of NRF1. Nature 528, 575–579 (2015). This paper shows that the removal of DNA methylation would create novel binding sites for NRF1 and thus affect the NRF1–DNA interactions in vivo , whereas other studies showed that DNA methylation could affect TF–DNA interactions in vitro. source

In the paper

“Similarly, the mCpG-binding protein MeCP2 was found to play a part in regulating exon splicing113.”

What the sources say

• Not checked yetcites [113] Maunakea, A. K., Chepelev, I., Cui, K. & Zhao, K. Intragenic DNA methylation modulates alternative splicing by recruiting MeCP2 to promote exon recognition. Cell Res. 23, 1256–1269 (2013). This study demonstrates that MeCP2 affects splicing events through its interaction with methylated DNA in vivo. source

In the paper

“this protein was previously shown to bind to methylated DNA47,51.”

What the sources say

• Not checked yetcites [47] Bahar Halpern, K., Vana, T. & Walker, M. D. Paradoxical role of DNA methylation in activation of FoxA2 gene expression during endoderm development. J. Biol. Chem. 289, 23882–23892 (2014). source
• Not checked yetcites [51] Spruijt, C. G. et al. Dynamic readers for 5-(hydroxy)methylcytosine and its oxidized derivatives. Cell 152, 1146–1159 (2013). This paper describes the identification of proteins that interact with mCpG sites, 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC) in ES cells and neuronal progenitor cells using a MS/MS-based approach. source

In the paper

“More importantly, a distinct set of sequence motifs was discovered in these regions”

What the sources say

• Not checked yetcites [101] Wan, J. et al. Characterization of tissue-specific differential DNA methylation suggests distinct modes of positive and negative gene expression regulation. BMC Genomics 16, 49 (2015). source

In the paper

“suggesting that some TFs bind preferentially to these regions101”

What the sources say

• Not checked yetcites [101] Wan, J. et al. Characterization of tissue-specific differential DNA methylation suggests distinct modes of positive and negative gene expression regulation. BMC Genomics 16, 49 (2015). source

In the paper

“They found that CpG density showed a negative correlation with methylation level”

What the sources say

• Not checked yetcites [83] Krebs, A. R., Dessus-Babus, S., Burger, L. & Schubeler, D. High-throughput engineering of a mammalian genome reveals building principles of methylation states at CG rich regions. eLife 3, e04094 (2014). source

In the paper

“Changes in DNA methylation are often associated with chromatin status, resulting in increased or decreased DNA accessibility 96,97.”

What the sources say

• Not checked yetcites [96] Maurano, M. T. et al. Role of DNA methylation in modulating transcription factor occupancy. Cell Rep. 12, 1184–1195 (2015). source
• Not checked yetcites [97] Domcke, S. et al. Competition between DNA methylation and transcription factors determines binding of NRF1. Nature 528, 575–579 (2015). This paper shows that the removal of DNA methylation would create novel binding sites for NRF1 and thus affect the NRF1–DNA interactions in vivo , whereas other studies showed that DNA methylation could affect TF–DNA interactions in vitro. source

In the paper

“suggesting that these sites are likely to be functional101”

What the sources say

• Not checked yetcites [101] Wan, J. et al. Characterization of tissue-specific differential DNA methylation suggests distinct modes of positive and negative gene expression regulation. BMC Genomics 16, 49 (2015). source

In the paper

“A single-nucleotide mutation in the CTCF binding motif had no effect on the DNA methylation level.”

What the sources say

• Not checked yetcites [78] Stadler, M. B. et al. DNA-binding factors shape the mouse methylome at distal regulatory regions. Nature 480, 490–495 (2011). This paper demonstrates that some proteins, such as CTCF and REST, can reduce DNA methylation levels at the genomic regions near their binding regions. source

In the paper

“the sequence-specific DNA-binding protein RFX activates a methylated promoter49.”

What the sources say

• Not checked yetcites [49] Niesen, M. I. et al. Activation of a methylated promoter mediated by a sequence-specific DNA-binding protein, RFX. J. Biol. Chem. 280, 38914–38922 (2005). source

In the paper

“Importantly, the binding of CTCF to the exon or intron was dependent on DNA methylation”

What the sources say

• Not checked yetcites [112] Shukla, S. et al. CTCF-promoted RNA polymerase II pausing links DNA methylation to splicing. Nature 479, 74–79 (2011). source

In the paper

“Among the differentially methylated regions located within 4 kb upstream of transcriptional start sites, approximately 47% showed a positive correlation with the expression of their putative target genes.”

What the sources say

• Not checked yetcites [101] Wan, J. et al. Characterization of tissue-specific differential DNA methylation suggests distinct modes of positive and negative gene expression regulation. BMC Genomics 16, 49 (2015). source

In the paper

“the nuclear receptor PPARγ (peroxisome proliferator-activated receptor-γ) recruits TET1, resulting in a reduced methylation level around its binding sites through the interaction with TET1 (REF. 85).”

What the sources say

• Not checked yetcites [85] Fujiki, K. et al. PPARγ-induced PARylation promotes local DNA demethylation by production of 5-hydroxymethylcytosine. Nat. Commun. 4, 2262 (2013). source

In the paper

“It is well known that DNA methylation can affect the binding of some TFs87.”

What the sources say

• Not checked yetcites [87] Tate, P. H. & Bird, A. P. Effects of DNA methylation on DNA-binding proteins and gene expression. Curr. Opin. Genet. Dev. 3, 226–231 (1993). source

In the paper

“For example, a handful pioneer TFs (such as OCT4, SOX2 and KLF4) were shown to bind only partial motifs displayed on the nucleosome surface107.”

What the sources say

• Not checked yetcites [107] Soufi, A. et al. Pioneer transcription factors target partial DNA motifs on nucleosomes to initiate reprogramming. Cell 161, 555–568 (2015). source

In the paper

“suggesting that the methylation status surrounding the spliced exons could affect the inclusion level of these exons.”

What the sources say

• Not checked yetcites [112] Shukla, S. et al. CTCF-promoted RNA polymerase II pausing links DNA methylation to splicing. Nature 479, 74–79 (2011). source

In the paper

“Notably, as methylation-binding proteins participate in multiple biological processes, including gene regulation and splicing regulation, not all methylation-binding proteins are likely to be pioneer factors.”

In the paper

“This work suggests that protein binding has a general role in reducing DNA methylation levels”

In the paper

“these results support a model whereby the binding of certain proteins can directly affect DNA methylation levels (FIG. 3).”

In the paper

“methylation at the 3′ end of the CpG island confers tissue-specific transcriptional activation during human ES cell differentiation100.”

What the sources say

• Not checked yetcites [100] Yu, D. H. et al. Developmentally programmed 3′ CpG island methylation confers tissue- and cell-type-specific transcriptional activation. Mol. Cell. Biol. 33, 1845–1858 (2013). source

In the paper

“Interestingly, several of their members showed the ability to bind to methylated DNA, including HOXA5, HOXA9, GATA3 and GATA4 (REF. 53).”

What the sources say

• Not checked yetcites [53] Hu, S. et al. DNA methylation presents distinct binding sites for human transcription factors. eLife 2, e00726 (2013). This study identifies the transcription factors that preferentially bind to methylated DNA using a protein microarray-based approach and verified that endogenous KLF4 binds to methylated DNA in human ES cells. source

In the paper

“Taken together, the notion that TFs may act as DNA methylation readers is an emerging concept supported by predominantly in vitro but also by emerging in vivo evidence.”

In the paper

“For example, profiling the DNA methylation status in the promoters of 272 glioblastoma tumours showed that a distinct subset of samples displayed hypermethylation at a large number of loci, a phenotype termed ‘CpG island methylator phenotype’ (REF. 116).”

What the sources say

• Not checked yetcites [116] Noushmehr, H. et al. Identification of a CpG island methylator phenotype that defines a distinct subgroup of glioma. Cancer Cell 17, 510–522 (2010). source

In the paper

“indicating that other mCpG-binding proteins are involved in regulating the splicing process.”

What the sources say

• Not checked yetcites [115] Wan, J. et al. Integrative analysis of tissue-specific methylation and alternative splicing identifies conserved transcription factor binding motifs. Nucleic Acids Res. 41, 8503–8514 (2013). source

In the paper

“Specifically, owing to the interference with TET family proteins, the mutation causes the CpG island methylator phenotype116,123.”

What the sources say

• Not checked yetcites [116] Noushmehr, H. et al. Identification of a CpG island methylator phenotype that defines a distinct subgroup of glioma. Cancer Cell 17, 510–522 (2010). source
• Not checked yetcites [123] Turcan, S. et al. IDH1 mutation is sufficient to establish the glioma hypermethylator phenotype. Nature 483, 479–483 (2012). source

In the paper

“Mutant IDH protein is a competitive inhibitor of hydroxylases, including the TET family of 5mC hydroxylases120–122.”

What the sources say

• Not checked yetcites [120] Cairns, R. A. & Mak, T. W. Oncogenic isocitrate dehydrogenase mutations: mechanisms, models, and clinical opportunities. Cancer Discov. 3, 730–741 (2013). source
• Not checked yetcites [121] Dang, L. et al. Cancer-associated IDH1 mutations produce 2-hydroxyglutarate. Nature 462, 739–744 (2009). source
• Not checked yetcites [122] Xu, W. et al. Oncometabolite 2-hydroxyglutarate is a competitive inhibitor of α-ketoglutarate-dependent dioxygenases. Cancer Cell 19, 17–30 (2011). source

In the paper

“This observation is consistent with a global correlation analysis of DNA methylation and differential splicing events between the brain and the retina115.”

What the sources say

• Not checked yetcites [115] Wan, J. et al. Integrative analysis of tissue-specific methylation and alternative splicing identifies conserved transcription factor binding motifs. Nucleic Acids Res. 41, 8503–8514 (2013). source

In the paper

“As a result, the IDH mutation leads to a remodelling of DNA methylation profiles.”

In the paper

“suggesting that mCpG-binding proteins might play a part in splicing regulation.”

What the sources say

• Not checked yetcites [114] Lyko, F. et al. The honey bee epigenomes: differential methylation of brain DNA in queens and workers. PLoS Biol. 8, e1000506 (2010). source

In the paper

“As we demonstrated above, superimposing ChIP–seq and DNA methylome data sets can be an effective approach to validate mCpG-dependent DNA–TF interactions in vivo.”

In the paper

“Therefore, simply knocking down a methylation reader will not help reveal its role.”

In the paper

“Interestingly, the methylation levels in some of the regions were positively associated with the inclusion level of the spliced exons”

What the sources say

• Not checked yetcites [115] Wan, J. et al. Integrative analysis of tissue-specific methylation and alternative splicing identifies conserved transcription factor binding motifs. Nucleic Acids Res. 41, 8503–8514 (2013). source

In the paper

“In this case, a high methylation level led to MeCP2 binding to alternatively spliced exons, which resulted in exon inclusion (FIG. 4).”

What the sources say

• Not checked yetcites [113] Maunakea, A. K., Chepelev, I., Cui, K. & Zhao, K. Intragenic DNA methylation modulates alternative splicing by recruiting MeCP2 to promote exon recognition. Cell Res. 23, 1256–1269 (2013). This study demonstrates that MeCP2 affects splicing events through its interaction with methylated DNA in vivo. source

In the paper

“suggesting that other TFs might also participate in splicing regulation.”

What the sources say

• Not checked yetcites [115] Wan, J. et al. Integrative analysis of tissue-specific methylation and alternative splicing identifies conserved transcription factor binding motifs. Nucleic Acids Res. 41, 8503–8514 (2013). source

In the paper

“Instead, these two scenarios may well coexist in cells.”