Wnt signaling regulates hepatocyte cell division by a transcriptional repressor cascade
Open accessAbstract
Cell proliferation is tightly controlled by inhibitors that block cell cycle progression until growth signals relieve this inhibition, allowing cells to divide. In several tissues, including the liver, cell proliferation is inhibited at mitosis by the transcriptional repressors E2F7 and E2F8, leading to formation of polyploid cells. Whether growth factors promote mitosis and cell cycle progression by relieving the E2F7/E2F8-mediated inhibition is unknown. We report here on a mechanism of cell division control in the postnatal liver, in which Wnt/β-catenin signaling maintains active hepatocyte cell division through Tbx3 , a Wnt target gene. The TBX3 protein directly represses transcription of E2f7 and E2f8 , thereby promoting mitosis. This cascade of sequential transcriptional repressors, initiated by Wnt signals, provides a paradigm for exploring how commonly active developmental signals impact cell cycle completion.
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Written from the paper’s full text.
TL;DR
In mouse liver, Wnt signaling induces the transcriptional repressor Tbx3, which represses the mitotic inhibitors E2f7 and E2f8 to permit hepatocyte M-phase progression and maintain diploidy. This defines a transcriptional repressor cascade downstream of Wnt that controls hepatocyte cell division and has implications for tissue homeostasis and disease.
Key findings
- Tbx3 acts downstream of Wnt signaling in hepatocytes: CRISPR-Cas9 deletion of Apc caused ectopic Tbx3 expression, while Wls knockout led to partial loss of Tbx3.
- Tbx3 loss slowed cultured hepatocyte proliferation and increased polyploidy; Tbx3 overexpression increased proliferation even without the Wnt activator CHIR99021 and increased the percentage of diploid cells.
- In vivo, Tbx3 knockout increased polyploid nuclei (>2N: 59.8% in control vs. 92% in Tbx3 KO), while simultaneous deletion of Tbx3, E2f7, and E2f8 restored ploidy and nuclear size to control-like levels.
- TBX3 bound the E2f7 promoter and E2f8 enhancer by ChIP-seq/ChIP-qPCR; E2f7 and E2f8 transcripts increased upon Tbx3 loss and decreased upon Tbx3 overexpression, and reporter assays confirmed TBX3-mediated repression. Long-term Tbx3 knockout caused fibrosis in five of eight adult KO livers versus none of five controls.
Methods
- Mouse conditional genetics used Axin2-rtTA/TetO-Cre and Alb-CreERT2 drivers, with doxycycline from postnatal week 2 to week 4 or tamoxifen at P3; Apc was deleted by AAV-delivered CRISPR-Cas9, and Wls was deleted using Cdh5-CreERT2.
- Primary mouse hepatocytes were cultured under Wnt-dependent expansion conditions and manipulated by lentiviral Tbx3 shRNA knockdown or Sleeping Beauty-mediated Tbx3 overexpression.
- Analyses included ChIP-seq and ChIP-qPCR, mRNA in situ hybridization, qRT-PCR, luciferase reporter assays in HepG2 cells, flow cytometry for ploidy, nuclear area quantification, and Sirius Red staining for fibrosis.
Limitations
- The paper states that whether TBX3-high versus TBX3-low pericentral cells differ in Tbx3 function or proliferation rate is unclear, and whether mitosis in periportal hepatocytes is regulated by a periportal signal remains to be studied.
- The paper notes that Tbx3 is dosage-sensitive in other contexts and that loss of Tbx3 did not affect liver shape or relative mass, despite the later fibrotic phenotype.
- Evident from the text: human evidence is limited to TBX3 expression patterns and ENCODE ChIP-seq data rather than functional human liver experiments; several in vivo analyses rely on small samples (e.g., n = 3–4 mice, fibrosis in 5/8 KO vs. 0/5 control), and reporter assays were performed in HepG2 cells rather than primary hepatocytes.
Model-written summary. Check it against the paper before relying on it.
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Cites
Statistics
Claims the paper makes
38 of 63 cited sources checked · 19 supported · 1 flagged · accuracy not yet measured
- Findingcitation
Human TBX3 protein binds to E2F7 and E2F8 enhancers.
“Analysis of ChIP-Seq data from the ENCODE database (GSE105374) (33, 34) showed that human TBX3 protein also binds to E2F7 and E2F8 enhancers (SI Appendix, Fig. S3C)”
Quote in the paperSubstantial claim2 of 3 sources checked- cites GSE105374
Couldn’t be matched to a paper in Crossref, so it can’t be checked.
- cites [33] I. Dunham; ENCODE Project Consortium, An integrated encyclopedia of DNA elements in the human genome. Nature 489, 57– 74 (2012). 22955616 PMC3439153 source
The Genome-wide integration section reports transcription-factor binding at enhancers in general, but does not report TBX3 binding at E2F7 or E2F8 enhancers. Searches for TBX3, E2F7, E2F8, and GSE105374 found no passage in our copy establishing the specific claim. In the citing sentence, the binding result is described as the citing authors’ analysis of ENCODE data. (LLM, conf=0.76)
Experimental — accuracy not yet measured - cites [34] C. A. Davis, The Encyclopedia of DNA elements (ENCODE): Data portal update. Nucleic Acids Res. 46, D794– D801 (2018). 29126249 PMC5753278 source
- Findingown data
Tbx3 seems expressed in a gradient in the pericentral zone.
“Tbx3, which has been shown to act in a dosage-sensitive manner in other contexts (50), seems to be expressed in a gradient in the pericentral zone (SI Appendix, Fig. S1A).”
Quote in the paperNot a substantial claim1 of 1 sources checked- cites [50] R. Russell, A dynamic role of TBX3 in the pluripotency circuitry. Stem Cell Reports 5, 1155– 1170 (2015). 26651606 PMC4682344 source
- Premisecitation
In mouse and human liver, Wnt ligands are produced by endothelial cells of central vein branches and nearby sinusoids.
“In the mouse and human liver, Wnt ligands are produced by endothelial cells of the central vein branches and the nearby sinusoids (17–19)”
Quote in the paperSubstantial claim3 of 3 sources checked- cites [17] B. Wang, L. Zhao, M. Fish, C. Y. Logan, R. Nusse, Self-renewing diploid Axin2(+) cells fuel homeostatic renewal of the liver. Nature 524, 180– 185 (2015). 26245375 PMC4589224 source
The Discussion reports Wnt9b expression in central-vein endothelial cells and Wnt2 expression in both central-vein and sinusoidal endothelial cells. The experiments described are in mice; our copy does not establish the same finding in human liver. (LLM, conf=0.88)
Experimental — accuracy not yet measured - cites [18] L. McEnerney, Dual modulation of human hepatic zonation via canonical and non-canonical Wnt pathways. Exp. Mol. Med. 49, e413 (2017). 29244788 PMC5750478 source
The Introduction and Discussion describe rodent studies in which central-vein endothelial cells produce Wnt ligands. The Discussion says this still requires validation in human liver, and the paper does not establish production by endothelial cells of nearby sinusoids. Page numbers are unavailable in our copy. (LLM, conf=0.88)
Experimental — accuracy not yet measured - cites [19] L. Zhao, Tissue repair in the mouse liver following acute carbon tetrachloride depends on injury-induced Wnt/β-catenin signaling. Hepatology 69, 2623– 2635 (2019). 30762896 PMC7043939 source
The Results section (“Upregulation of endogenous and exogenous Wnt ligands”; page not supplied) says that Wnt2 and Wnt9b are expressed by central-vein endothelial cells and that Wnt2 is also expressed by sinusoidal endothelial cells. The paper’s experiments concern mouse liver; our copy does not establish the same claim for human liver. (LLM, conf=0.87)
Experimental — accuracy not yet measured
- Premisecitation
In the liver, E2f7 and E2f8 are responsible for cell cycle arrest that uncouples DNA synthesis from proliferation.
“In the liver, two members of the E2F family of cell cycle regulators, E2f7 and E2f8, are responsible for this form of cell cycle arrest (7, 8).”
- Also stated in: § Abstract · plain
“Therefore, in the liver, DNA synthesis is uncoupled from cell proliferation due to the activity of E2F7 and E2F8.”
Quote in the paperSubstantial claim1 of 2 sources checked - Also stated in: § Abstract · plain
- Premisecitation
Deleting E2f7 and E2f8 from hepatocytes results in completion of M phase and generation of diploid daughter cells.
“Deleting E2f7 and E2f8 from hepatocytes results in completion of M phase and the generation of diploid daughter cells (7, 8).”
Quote in the paperSubstantial claim1 of 2 sources checked
- Premisecitation
Hepatocytes in the postnatal liver expand rapidly in a Wnt/β-catenin signaling-dependent way.
“Hepatocytes in the postnatal liver expand rapidly in a Wnt/β-catenin signaling-dependent way (12–16).”
Quote in the paperSubstantial claim4 of 5 sources checked- cites [12] S. P. S. Monga, P. Pediaditakis, K. Mule, D. B. Stolz, G. K. Michalopoulos, Changes in WNT/β-catenin pathway during regulated growth in rat liver regeneration. Hepatology 33, 1098– 1109 (2001). 11343237 PMC1821078 source
The Abstract describes rapid hepatocyte proliferation after partial hepatectomy in adult rats, and the Discussion links early β-catenin nuclear translocation to that regeneration. It does not study normal postnatal liver expansion, and it suggests Wnt/β-catenin signaling may contribute to proliferation rather than establishing that expansion depends on it. The indexed passages provide no page numbers. (LLM, conf=0.88)
Experimental — accuracy not yet measured - cites [13] X. Tan, J. Behari, B. Cieply, G. K. Michalopoulos, S. P. Monga, Conditional deletion of beta-catenin reveals its role in liver growth and regeneration. Gastroenterology 131, 1561– 1572 (2006). 17101329 source
- cites [14] U. Apte, beta-Catenin is critical for early postnatal liver growth. Am. J. Physiol. Gastrointest. Liver Physiol. 292, G1578– G1585 (2007). 17332475 source
- cites [15] L. Planas-Paz, The RSPO-LGR4/5-ZNRF3/RNF43 module controls liver zonation and size. Nat. Cell Biol. 18, 467– 479 (2016). 27088858 source
- cites [16] T. Leibing, Angiocrine Wnt signaling controls liver growth and metabolic maturation in mice. Hepatology 68, 707– 722 (2018). 29059455 PMC6099291 source
The Abstract (page unavailable) describes postnatal hepatocyte growth and proliferation and reports that loss of endothelial Wnt signaling reduces liver size in mice. The Discussion (page unavailable) links reduced liver size to loss of Wnt activation. The paper does not establish that rapid postnatal hepatocyte expansion itself depends on Wnt/β-catenin signaling. (LLM, conf=0.83)
Experimental — accuracy not yet measured
- Premisecitation
Activating mutations in Wnt signaling components are common in liver cancer.
“activating mutations in Wnt signaling components are common in liver cancer (41, 42)”
Quote in the paperNot a substantial claim1 of 2 sources checked- cites [41] R. Nusse, H. Clevers, Wnt/β-catenin signaling, disease, and emerging therapeutic modalities. Cell 169, 985– 999 (2017). 28575679 source
- cites [42] W. Wang, R. Smits, H. Hao, C. He, Wnt/β-catenin signaling in liver cancers. Cancers (Basel) 11, 926 (2019). PMC6679127 31269694 source
Section 4.3 reports activating CTNNB1 mutations in 60–80% of hepatoblastomas, and section 4.1 reports them in 15–25% of hepatocellular carcinomas. But section 4.2 says mutations in related genes are less frequent in cholangiocarcinoma, so the unqualified claim across liver cancers overstates the evidence. Page numbers are unavailable in our copy. (LLM, conf=0.86)
Experimental — accuracy not yet measured
- Premisecitation
E2F7 and E2F8 inhibit expression of many genes including Aurka/b, Ccnb1, and Plk1.
“Acting as mutually redundant transcriptional repressors, E2F7 and E2F8 inhibit the expression of many genes, including Aurka/b, Ccnb1, and Plk1, which encode proteins that act during mitotic progression (7, 8, 11).”
Quote in the paperSubstantial claim2 of 3 sources checked- cites [7] H. Z. Chen, Canonical and atypical E2Fs regulate the mammalian endocycle. Nat. Cell Biol. 14, 1192– 1202 (2012). 23064266 PMC3616487 source
The Results section, “E2F7/E2F8 Regulate Endocycle-related Genes” (page unavailable), reports that loss of E2F7 or E2F8 derepresses many genes, including genes involved in G2/M progression and mitosis. Our copy does not identify Aurka, Aurkb, Ccnb1, or Plk1 as genes whose expression they inhibit, so it supports the general point but not the named examples. (LLM, conf=0.84)
Experimental — accuracy not yet measured - cites [8] S. K. Pandit, E2F8 is essential for polyploidization in mammalian cells. Nat. Cell Biol. 14, 1181– 1191 (2012). 23064264 source
- cites [11] A. de Bruin, Identification and characterization of E2F7, a novel mammalian E2F family member capable of blocking cellular proliferation. J. Biol. Chem. 278, 42041– 42049 (2003). 12893818 source
The FIG. 5 section reports that E2F7 inhibits E2F-mediated reporter activation, and the Discussion says E2F7 can repress a subset of E2F-regulated promoters (page numbers unavailable). Our copy does not establish a mutually redundant role for E2F8 or identify Aurka/b, Ccnb1, or Plk1 as repressed targets; the Discussion says the specific targets mediating growth inhibition still need to be identified. (LLM, conf=0.88)
Experimental — accuracy not yet measured
- Premisecitation
Expression of E2f7 and E2f8 is induced at weaning when the liver switches from diploid to polyploid.
“Expression of E2f7 and E2f8 is induced at the time of weaning when the tissue switches from initially containing diploid hepatocytes to becoming mostly polyploid (4, 9, 10).”
Quote in the paperSubstantial claim1 of 3 sources checked- cites [4] J.-E. Guidotti, Liver cell polyploidization: A pivotal role for binuclear hepatocytes. J. Biol. Chem. 278, 19095– 19101 (2003). 12626502 source
- cites [9] G. Gentric, S. Celton-Morizur, C. Desdouets, Polyploidy and liver proliferation. Clin. Res. Hepatol. Gastroenterol. 36, 29– 34 (2012). 21778131 source
- cites [10] S. Zhang, Y. H. Lin, B. Tarlow, H. Zhu, The origins and functions of hepatic polyploidy. Cell Cycle 18, 1302– 1315 (2019). 31096847 PMC6592246 source
- Findingown data
Tbx3 is expressed in Wnt-responsive pericentral hepatocytes during postnatal liver growth.
“Similar to the adult liver, Tbx3 is expressed in Wnt-responsive pericentral hepatocytes during postnatal liver growth (SI Appendix, Fig. S1A).”
- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
“In the adult human liver, TBX3 is specifically expressed in pericentral hepatocytes, similar to the mouse liver (SI Appendix, Fig. S3B).”
Quote in the paperSubstantial claim - Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
- Findingown data
Tbx3 represses transcription of E2f7 and E2f8.
“We found that the TBX3 protein binds to promoter and enhancer regions of E2f7 and E2f8, respectively, and verified this interaction by ChIP-qPCR (Fig. 3A and B).”
- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
“E2f7 and E2f8 transcripts were both ectopically increased in Tbx3 KO livers (Fig. 3C and D).”
- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
“TBX3 is a specific transcriptional repressor of E2f7 and E2f8”
Quote in the paperSubstantial claim - Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
- Findingown data
Tbx3 knockdown slows hepatocyte proliferation.
“This resulted in significant slowing down of hepatocyte proliferation (Fig. 1B)”
- Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
“Tbx3 knockdown (Tbx3 KD) slows down hepatocyte proliferation.”
Quote in the paperSubstantial claim - Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
- Findingown data
Wnt signaling induces Tbx3 expression in hepatocytes.
“Here, we find that Wnt induces expression of the transcription factor Tbx3”
- Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
“Using CRISPR-Cas9 gene editing, we deleted Apc, a negative regulator of Wnt signal transduction in hepatocytes, and observed ectopic expression of Tbx3 (Fig. 1A).”
- Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
“Conversely, mice carrying mutations for the Wnt secretion machinery gene Wntless (Wls) exhibited a partial loss of Tbx3 expression (Fig. 1A).”
Quote in the paperSubstantial claim - Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
- Findingown data
Tbx3 knockdown increases polyploidy in cultured hepatocytes.
“increased numbers of polyploid cells (Fig. 1C and SI Appendix, Fig. S1C and Table S1)”
- Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
“Tbx3 knockdown leads to increased numbers of polyploid hepatocytes in culture.”
Quote in the paperSubstantial claim - Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
- Findingown data
Tbx3 overexpression increases hepatocyte proliferation even without Wnt activation.
“overexpression of Tbx3 in hepatocytes was sufficient to increase proliferation (Fig. 1D and SI Appendix, Fig. S1D and E), even in the absence of CHIR99021, a GSK3 inhibitor that activates Wnt signaling (Fig. 1D).”
- Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
“Tbx3 overexpression (OE) is sufficient to increase hepatocyte proliferation, in the absence of activated Wnt signaling.”
Quote in the paperSubstantial claim - Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
- Findingown data
Loss of Tbx3 in hepatocytes causes increased polyploidy in vivo.
“nuclear DNA content analysis (Fig. 2D) of GFP-labeled hepatocytes revealed a significant increase in the proportion of polyploid nuclei in Tbx3 KO livers (>2N: 59.8% in control and 92% in Tbx3 KO)”
- Also stated in: § Deletion of Tbx3 and Changes in Ploidy. · plain
“Tbx3 KO livers were comprised of larger nuclei at all analyzed timepoints compared to control”
- Also stated in: § Deletion of Tbx3 and Changes in Ploidy. · boosted
“DNA content analysis confirmed increased ploidy in mutant tissues”
Quote in the paperSubstantial claim - Also stated in: § Deletion of Tbx3 and Changes in Ploidy. · plain
- Findingown data
Tbx3 overexpression increases the percentage of diploid cells.
“overexpression of Tbx3 increased the percentage of diploid cells (Fig. 1E and SI Appendix, Table S1).”
- Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
“Tbx3 overexpression in primary hepatocytes leads to a higher percentage of diploid cells.”
Quote in the paperSubstantial claim - Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
- Findingown data
E2f7 and E2f8 transcripts are increased in Tbx3 KO livers.
“E2f7 and E2f8 transcripts were both ectopically increased in Tbx3 KO livers (Fig. 3C and D).”
- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
“qRT-PCR analysis of Tbx3, E2f7, and E2f8 transcripts from whole livers of the same samples as in C show significantly increased levels of E2f7 and E2f8 in the absence of Tbx3.”
Quote in the paperSubstantial claim - Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
- Findingown data
Loss of Tbx3 leads to fibrosis in adult liver.
“collagen staining revealed fibrotic areas in five of eight adult Tbx3 KO livers, while none of the five control animals exhibited fibrosis (Fig. 5C).”
- Also stated in: § Discussion · hedged
“the fibrotic phenotype of Tbx3 KO mice indicates an important role for cell cycle regulation in the tissue.”
Quote in the paperSubstantial claim - Also stated in: § Discussion · hedged
- Findingown data
Loss of Tbx3 did not affect liver shape or relative mass at any timepoint.
“Loss of Tbx3 did not affect liver shape or relative mass, at any timepoint (SI Appendix, Fig. S2C and D).”
- Also stated in: § Loss of Tbx3 Leads to Fibrosis. · hedged
“we did not observe major changes in liver weight or morphology (SI Appendix, Fig. S2C and D).”
Quote in the paperNot a substantial claim - Also stated in: § Loss of Tbx3 Leads to Fibrosis. · hedged
- Interpretationown data
Tbx3 fine tunes the degree and onset of zonated polyploidy in the liver by promoting mitosis and cell division.
“Our data indicated that Tbx3, acting in the Wnt signaling network, fine tunes the degree and onset of zonated polyploidy in the liver, by promoting mitosis and cell division.”
Quote in the paperSubstantial claim - Premisecitation
Tbx3-negative hepatocytes in the periportal zone maintain low ploidy levels through the animal's lifetime.
“Moreover, Tbx3-negative hepatocytes in the periportal zone seem to maintain low ploidy levels through the course of the animal’s lifetime (51).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [51] S. Tanami, Dynamic zonation of liver polyploidy. Cell Tissue Res. 368, 405– 410 (2017). 27301446 source
- Interpretationown data
TBX3 has conserved functions in human and mouse livers.
“suggesting that TBX3 has conserved functions in both human and mouse livers.”
Quote in the paperSubstantial claim - Interpretationreasoning
Tbx3 knockdown causes hepatocytes to fail to complete M phase.
“suggesting that hepatocytes failed to complete M phase.”
Quote in the paperSubstantial claim - Interpretationown data
Tbx3 regulates polyploidization of hepatocytes in vivo and in vitro by controlling cell division.
“Taken together, we conclude that Tbx3 regulates polyploidization of hepatocytes in vivo and in vitro by its control over cell division.”
Quote in the paperSubstantial claim - Interpretationreasoning
Such a cascade of transcriptional repressors may be a general mechanism for cell division control.
“Such a cascade of transcriptional repressors may be a general mechanism for cell division control.”
Quote in the paperSubstantial claim - Study design and conductown data
Two of the five Tbx3 shRNAs tested were verified to effectively knock down Tbx3 in primary hepatocytes.
“Hepatocytes were harvested 4 d after the second transfection for qRT-PCR analysis and two out of five Tbx3 shRNA were verified to be effectively knocking down Tbx3.”
Quote in the paperSubstantial claim - Interpretationreasoning
The fibrotic phenotype of Tbx3 KO mice indicates an important role for cell cycle regulation in the tissue.
“the fibrotic phenotype of Tbx3 KO mice indicates an important role for cell cycle regulation in the tissue.”
Quote in the paperSubstantial claim - Premisecitation
In some tissues including liver, cells can enter S phase but not complete M phase, becoming polyploid.
“In some tissues, including the liver, cells can enter S phase to duplicate their genome, but do not complete M phase, thus becoming polyploid (4–6).”
Quote in the paperSubstantial claim0 of 3 sources checked- cites [4] J.-E. Guidotti, Liver cell polyploidization: A pivotal role for binuclear hepatocytes. J. Biol. Chem. 278, 19095– 19101 (2003). 12626502 source
- cites [5] J. I. Øvrebø, B. A. Edgar, Polyploidy in tissue homeostasis and regeneration. Development 145, dev156034 (2018). 30021843 PMC10682953 source
- cites [6] R. Donne, M. Saroul-Aïnama, P. Cordier, S. Celton-Morizur, C. Desdouets, Polyploidy in liver development, homeostasis and disease. Nat. Rev. Gastroenterol. Hepatol. 17, 391– 405 (2020). 32242122 source
- Classified as probably not a substantial claim. 21
- May combine two claims. 16
- The quote only partly states this claim. 1
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationTbx3 promotes hepatic progenitor proliferation likely by repressing p19ARF.
“Tbx3 is required for embryonic liver development and promotes hepatic progenitor proliferation likely by repressing the p19ARF(Cdkn2a) cell cycle inhibitor (23).”
Quote in the paperNot a substantial claim1 of 1 sources checked- cites [23] T. H. Lüdtke, V. M. Christoffels, M. Petry, A. Kispert, Tbx3 promotes liver bud expansion during mouse development by suppression of cholangiocyte differentiation. Hepatology 49, 969– 978 (2009). 19140222 source
The paper supports a role for Tbx3 in embryonic liver development, but disputes the proposed p19ARF mechanism. In “Proliferation and Migration Defects in the Tbx3-Deficient Liver Bud Are Secondary to Cell Fate Changes” (page not provided), it says p19ARF up-regulation is likely a late consequence of altered cell fate, not the cause of the proliferation defect. (LLM, conf=0.88)
Checked by a reasoning model — may be wrong.
Experimental — accuracy not yet measured
May combine two claims.
PremisecitationIn the absence of a capacity for cell division, fibrosis becomes a mode of tissue repair and leads to tissue scarring.
“In the absence of a capacity for cell division, fibrosis becomes a mode of tissue repair and leads to tissue scarring (52, 53).”
Quote in the paperSubstantial claim1 of 2 sources checked- cites [52] M. R. Dewhurst, Loss of hepatocyte cell division leads to liver inflammation and fibrosis. PLoS Genet. 16, e1009084 (2020). 33147210 PMC7641358 source
The Results (page not provided) report elevated liver fibrosis in mice whose hepatocytes cannot divide, and the Discussion (page not provided) describes a pathway from impaired division to fibrosis. The paper does not establish that fibrosis serves as a mode of tissue repair when cells cannot divide generally, or explicitly describe the resulting fibrosis as tissue scarring. (LLM, conf=0.84)
Experimental — accuracy not yet measured - cites [53] T. Kisseleva, D. Brenner, Molecular and cellular mechanisms of liver fibrosis and its regression. Nat. Rev. Gastroenterol. Hepatol. 18, 151– 166 (2021). 33128017 source
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationE2f7 and E2f8, as well as polyploidy, suppress tumorigenesis.
“E2f7 and E2f8, as well as polyploidy, are known to suppress tumorigenesis (43–46).”
Quote in the paperNot a substantial claim4 of 4 sources checked- cites [43] L. N. Kent, E2f8 mediates tumor suppression in postnatal liver development. J. Clin. Invest. 126, 2955– 2969 (2016). 27454291 PMC4966321 source
The Discussion reports that loss of the E2F7/8 repressors leads to hepatocellular carcinoma in mice, and “Tumor suppressive function of E2F8” specifically identifies E2F8 as suppressing cancer initiation in developing livers. The paper also links E2F7/8 loss to reduced hepatocyte ploidy, but the passages do not establish that polyploidy itself suppresses tumorigenesis. Page numbers were not available in our copy. (LLM, conf=0.86)
Experimental — accuracy not yet measured - cites [44] S. Zhang, The polyploid state plays a tumor-suppressive role in the liver. Dev. Cell 44, 447– 459.e5 (2018). 29429824 PMC5828993 source
The Results section ('E2f8 knockout and Anln shRNA mice confirmed the protective effects of polyploidy'; page unavailable) reports more liver tumors in E2f8-deficient mice, but attributes the difference chiefly to their lower ploidy rather than to E2f8 itself. The Introduction (page unavailable) mentions E2f7/E2f8 double-knockout livers but does not establish a tumor-suppressive role for E2f7. The source supports polyploidy's protective effect in the liver, not the full claim that both genes and polyploidy each suppress tumorigenesis. (LLM, conf=0.88)
Experimental — accuracy not yet measured - cites [45] P. D. Wilkinson, Polyploid hepatocytes facilitate adaptation and regeneration to chronic liver injury. Am. J. Pathol. 189, 1241– 1255 (2019). 30928253 PMC6547059 source
- cites [46] E. Moreno, E2F7 is a potent inhibitor of liver tumor growth in adult mice. Hepatology 73, 303– 317 (2021). 32259305 PMC7898887 source
The Results section ('E2F7/8 Transgene Induction in Liver Tumors Causes Replication Stress and Inhibits Neoplastic Growth'; page unavailable) says E2f7 and E2f8 overexpression inhibits tumorigenesis in mouse liver tumors. Our copy does not say that polyploidy suppresses tumorigenesis; the References section only lists a study about E2F8's role in polyploidization. (LLM, conf=0.88)
Experimental — accuracy not yet measured
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationWnt target genes and key metabolic enzymes have a zonated expression pattern in pericentral hepatocytes.
“creating a zonated expression pattern of Wnt target genes and key metabolic enzymes in pericentral hepatocytes (20–22).”
Quote in the paperNot a substantial claim2 of 3 sources checked- cites [20] S. Benhamouche, Apc tumor suppressor gene is the “zonation-keeper” of mouse liver. Dev. Cell 10, 759– 770 (2006). 16740478 source
- cites [21] K. B. Halpern, Single-cell spatial reconstruction reveals global division of labour in the mammalian liver. Nature 542, 352– 356 (2017). 28166538 PMC5321580 source
- cites [22] K. B. Halpern, Paired-cell sequencing enables spatial gene expression mapping of liver endothelial cells. Nat. Biotechnol. 36, 962– 970 (2018). 30222169 PMC6546596 source
The Discussion (page unavailable) describes pericentrally zonated xenobiotic-metabolism enzymes in hepatocytes, supporting the metabolic-enzyme part of the claim. The Results section on pericentral LECs (page unavailable) says these cells secrete Wnt ligands that maintain hepatocyte zonation, but it does not establish that Wnt target genes themselves have zonated expression in pericentral hepatocytes. (LLM, conf=0.84)
Experimental — accuracy not yet measured
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationA role for Wnt signaling during M phase has been suggested based on high Wnt/β-catenin signaling activity during cell division.
“A role for Wnt signaling during M phase has also been suggested based on high Wnt/β-catenin signaling activity during cell division, through an unknown mechanism (39, 40).”
Quote in the paperNot a substantial claim2 of 2 sources checked- cites [39] C. Niehrs, S. P. Acebron, Mitotic and mitogenic Wnt signalling. EMBO J. 31, 2705– 2713 (2012). 22617425 PMC3380213 source
- cites [40] B. W. Benham-Pyle, J. Y. Sim, K. C. Hart, B. L. Pruitt, W. J. Nelson, Increasing β-catenin/Wnt3A activity levels drive mechanical strain-induced cell cycle progression through mitosis. eLife 5, e19799 (2016). 27782880 PMC5104517 source
The Discussion reports that higher Wnt3A/β-catenin activity drove mechanically strained cells from G2 into mitosis, supporting a suggested role in mitotic progression. It does not establish that signaling activity was high during M phase itself; the reported activity concerns progression into mitosis. (LLM, conf=0.83)
Experimental — accuracy not yet measured
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationE2f7 and E2f8 are expressed in several other tissues, such as placenta and pancreas, where they are implicated in arresting the cell cycle.
“These mitotic inhibitors are expressed in several other tissues, such as the placenta and pancreas, where they are also implicated in arresting the cell cycle (8, 47–49).”
Quote in the paperNot a substantial claim3 of 4 sources checked- cites [8] S. K. Pandit, E2F8 is essential for polyploidization in mammalian cells. Nat. Cell Biol. 14, 1181– 1191 (2012). 23064264 source
- cites [47] Q. R. Qi, Involvement of atypical transcription factor E2F8 in the polyploidization during mouse and human decidualization. Cell Cycle 14, 1842– 1858 (2015). 25892397 PMC4614110 source
The Introduction (page not provided) links E2F7 and E2F8 to trophoblast giant cells and describes them as transcriptional repressors. The section “E2F8 initiates the endoreplication of mouse decidual cells” (page not provided) says E2F8 represses mitosis and cytokinesis in decidual cells. Our copy does not establish expression in the pancreas or cell-cycle arrest in the placenta and pancreas. (LLM, conf=0.84)
Experimental — accuracy not yet measured - cites [48] R. B. Matondo, Atypical E2f functions are critical for pancreas polyploidization. PLoS One 13, e0190899 (2018). 29329320 PMC5766144 source
- cites [49] M. Mizuno, The role of E2F8 in the human placenta. Mol. Med. Rep. 19, 293– 301 (2019). 30387815 PMC6297733 source
The Results (page unavailable) show E2F8 expression in the human placenta, but the study did not investigate E2F7 expression there or address expression in the pancreas. Its Results also report that E2F8 silencing did not significantly affect the cell cycle or proliferation in the placental cell line tested, so this paper does not support the attributed role in cell-cycle arrest. (LLM, conf=0.88)
Experimental — accuracy not yet measured
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationTbx3 is among the genes highly expressed in the pericentral zone.
“Among the genes highly expressed in the pericentral zone is the transcriptional repressor Tbx3 (17, 19).”
Quote in the paperNot a substantial claim2 of 2 sources checked- cites [17] B. Wang, L. Zhao, M. Fish, C. Y. Logan, R. Nusse, Self-renewing diploid Axin2(+) cells fuel homeostatic renewal of the liver. Nature 524, 180– 185 (2015). 26245375 PMC4589224 source
- cites [19] L. Zhao, Tissue repair in the mouse liver following acute carbon tetrachloride depends on injury-induced Wnt/β-catenin signaling. Hepatology 69, 2623– 2635 (2019). 30762896 PMC7043939 source
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationTbx3 maintains expression of hepatocyte-lineage genes and represses cholangiocyte fate.
“Tbx3 also maintains expression of hepatocyte-lineage genes, such as Cebpα and Hnf4α, and represses the cholangiocyte fate (23, 24).”
Quote in the paperNot a substantial claim2 of 2 sources checked- cites [23] T. H. Lüdtke, V. M. Christoffels, M. Petry, A. Kispert, Tbx3 promotes liver bud expansion during mouse development by suppression of cholangiocyte differentiation. Hepatology 49, 969– 978 (2009). 19140222 source
- cites [24] A. Suzuki, S. Sekiya, D. Büscher, J. C. Izpisúa Belmonte, H. Taniguchi, Tbx3 controls the fate of hepatic progenitor cells in liver development by suppressing p19ARF expression. Development 135, 1589– 1595 (2008). 18356246 source
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationTbx3 is required for embryonic liver development.
“Tbx3 is required for embryonic liver development and promotes hepatic progenitor proliferation likely by repressing the p19ARF(Cdkn2a) cell cycle inhibitor (23).”
Quote in the paperNot a substantial claim1 of 1 sources checked- cites [23] T. H. Lüdtke, V. M. Christoffels, M. Petry, A. Kispert, Tbx3 promotes liver bud expansion during mouse development by suppression of cholangiocyte differentiation. Hepatology 49, 969– 978 (2009). 19140222 source
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationTbx3 acts as a target of Wnts in other contexts.
“These findings agree with Tbx3 acting as a target of Wnts in other contexts (25, 27–30).”
Quote in the paperNot a substantial claim3 of 5 sources checked- cites [25] C. A. Renard, Tbx3 is a downstream target of the Wnt/beta-catenin pathway and a critical mediator of beta-catenin survival functions in liver cancer. Cancer Res. 67, 901– 910 (2007). 17283120 source
- cites [27] N. Aydoğdu, TBX2 and TBX3 act downstream of canonical WNT signaling in patterning and differentiation of the mouse ureteric mesenchyme. Development 145, dev171827 (2018). 30478225 source
- cites [28] A. Waghray, Tbx3 controls Dppa3 levels and exit from pluripotency toward mesoderm. Stem Cell Reports 5, 97– 110 (2015). 26095607 PMC4618439 source
- cites [29] M. C. Eblaghie, Interactions between FGF and Wnt signals and Tbx3 gene expression in mammary gland initiation in mouse embryos. J. Anat. 205, 1– 13 (2004). 15255957 PMC1571327 source
- cites [30] D. Zimmerli, TBX3 acts as tissue-specific component of the Wnt/β-catenin transcriptional complex. eLife 9, e58123 (2020). 32808927 PMC7434441 source
The Results and discussion describe TBX3 as a regulator of Wnt target genes, but do not say that TBX3 itself is a target of Wnts in other contexts. Searches for TBX3 as a Wnt target and for Wnt-dependent TBX3 expression found no passage addressing that direction of regulation. (LLM, conf=0.76)
Experimental — accuracy not yet measured
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationTbx3 acts in a dosage-sensitive manner in other contexts.
“Tbx3, which has been shown to act in a dosage-sensitive manner in other contexts (50)”
Quote in the paperNot a substantial claim1 of 1 sources checked- cites [50] R. Russell, A dynamic role of TBX3 in the pluripotency circuitry. Stem Cell Reports 5, 1155– 1170 (2015). 26651606 PMC4682344 source
May combine two claims.
Findingown dataTbx3 knockdown increases and overexpression decreases E2f7/E2f8 transcripts in cultured hepatocytes.
“Knockdown and overexpression of Tbx3 in cultured primary hepatocytes further corroborated the interactions with E2f7 and E2f8 (Fig. 3E and F).”
- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
“qRT-PCR analysis shows increased E2f7 and E2f8 transcription in Tbx3 KD cultured mouse primary hepatocytes.”
- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
“E2f7 and E2f8 are down-regulated upon overexpression of Tbx3.”
Quote in the paperSubstantial claim- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
May combine two claims.
Findingown dataTBX3 represses E2f7/E2f8 via binding to their regulatory regions, confirmed by luciferase reporter.
“Addition of the mouse E2f7 or E2f8 enhancer regions containing the TBX3 binding sites to an HSV-TK reporter construct led to significant repression of the luciferase reporter activity compared to control vector with HSV-TK promoter only (Fig. 3G).”
- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · boosted
“reporter activity was increased when TBX3 was knocked down or T-box binding motifs were mutated or deleted from the enhancer regions, confirming that TBX3 is a specific transcriptional repressor of E2f7 and E2f8 (Fig. 3G and SI Appendix, Fig. S3A).”
Quote in the paperSubstantial claim- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · boosted
May combine two claims. The quote only partly states this claim.
Findingown dataTbx3 promotes hepatocyte proliferation and mitotic progression.
“This resulted in significant slowing down of hepatocyte proliferation (Fig. 1B) and increased numbers of polyploid cells (Fig. 1C and SI Appendix, Fig. S1C and Table S1), suggesting that hepatocytes failed to complete M phase.”
- Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
“overexpression of Tbx3 in hepatocytes was sufficient to increase proliferation (Fig. 1D and SI Appendix, Fig. S1D and E), even in the absence of CHIR99021”
- Also stated in: § Deletion of Tbx3 and Changes in Ploidy. · plain
“Tbx3 regulates polyploidization of hepatocytes in vivo and in vitro by its control over cell division.”
Quote in the paperSubstantial claim- Also stated in: § Tbx3 Promotes Hepatocyte Proliferation Downstream of Wnt Signaling. · plain
May combine two claims.
Findingown dataSimultaneous deletion of Tbx3, E2f7, and E2f8 restores normal ploidy and nuclear size.
“GFP-labeled nuclei in Tbx3-E2f7-E2f8 TKO livers were indistinguishable from controls in ploidy and size (Fig. 4B–D and SI Appendix, Table S1).”
- Also stated in: § Epistatic Relationships between Tbx3 and E2f7/E2f8. · plain
“simultaneous removal of E2f7, E2f8, and Tbx3 resolves the polyploid phenotype caused by removal of Tbx3 alone.”
Quote in the paperSubstantial claim- Also stated in: § Epistatic Relationships between Tbx3 and E2f7/E2f8. · plain
May combine two claims.
Findingown dataTBX3 binds to the E2f7 promoter and the E2f8 enhancer.
“We found that the TBX3 protein binds to promoter and enhancer regions of E2f7 and E2f8, respectively, and verified this interaction by ChIP-qPCR (Fig. 3A and B).”
- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
“TBX3 binds to E2f7 promoter and E2f8 enhancer, shown by ChIP-Seq binding profiles”
Quote in the paperSubstantial claim- Also stated in: § TBX3 Represses Transcription of E2f7 and E2f8. · plain
May combine two claims.
Findingown dataCHIR99021 alone enhances hepatocyte proliferation but less than Tbx3 overexpression.
“As expected, CHIR99021 alone enhanced proliferation of hepatocytes albeit not to the same degree as Tbx3 overexpression (Fig. 1D).”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataTbx3 KO livers have cytoplasmic vacuoles and anomalies in the pericentral zone.
“Upon histological analysis, we observed cytoplasmic vacuoles and anomalies in the pericentral zone (Fig. 5B).”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S4 (based).
Findingnone givenIn the growing and adult liver, Tbx3 is expressed primarily in the pericentral zone.
“in the growing and adult liver, Tbx3 is expressed primarily in the pericentral zone”
Quote in the paperNot a substantial claimMay combine two claims.
Interpretationown dataLoss of Tbx3 is not compatible with long-term homeostatic tissue renewal, likely due to failure in cell division leading to compensatory fibrotic repair.
“Appearance of fibrotic areas in Tbx3 KO livers suggests that loss of Tbx3 is not compatible with long-term homeostatic tissue renewal, likely due to failure in cell division, which leads to compensatory fibrotic repair.”
Quote in the paperSubstantial claimMay combine two claims.
PremisecitationIn hepatic tumor cells with activating β-catenin mutations, Tbx3 is overexpressed and mediates cell proliferation and survival.
“In hepatic tumor cells with activating β-catenin mutations, Tbx3 is overexpressed and mediates cell proliferation and survival (25).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [25] C. A. Renard, Tbx3 is a downstream target of the Wnt/beta-catenin pathway and a critical mediator of beta-catenin survival functions in liver cancer. Cancer Res. 67, 901– 910 (2007). 17283120 source
May combine two claims.
Interpretationown dataIn the absence of Tbx3, hepatocytes complete S phase but not M phase.
“The decrease in diploidy and the concomitant increase in polyploidy indicated that in the absence of Tbx3, hepatocytes were able to complete S phase but not M phase.”
Quote in the paperSubstantial claimMay combine two claims.
Interpretationown dataGenetic interactions imply a linear pathway: Wnt activates Tbx3, which represses E2f7/E2f8, relieving cell cycle arrest and maintaining diploid dividing state.
“These genetic interactions imply a linear pathway whereby Wnt activates Tbx3, which represses the mitotic inhibitors E2f7 and E2f8; this therefore relieves cell cycle arrest and maintains hepatocytes in a diploid, dividing state.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationWnt/β-catenin signaling controls the G1/S transition.
“The Wnt/β-catenin signaling pathway is one of the mechanisms known to control G1/S transition (35–38).”
Quote in the paperNot a substantial claim0 of 4 sources checked- cites [35] T. C. He, Identification of c-MYC as a target of the APC pathway. Science 281, 1509– 1512 (1998). 9727977 source
- cites [36] M. Shtutman, The cyclin D1 gene is a target of the beta-catenin/LEF-1 pathway. Proc. Natl. Acad. Sci. U.S.A. 96, 5522– 5527 (1999). 10318916 PMC21892 source
- cites [37] O. Tetsu, F. McCormick, Beta-catenin regulates expression of cyclin D1 in colon carcinoma cells. Nature 398, 422– 426 (1999). 10201372 source
- cites [38] C. Torre, The transforming growth factor-α and cyclin D1 genes are direct targets of β-catenin signaling in hepatocyte proliferation. J. Hepatol. 55, 86– 95 (2011). 21145869 source
Probably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductnone givenAutomatically detected hepatocyte nuclei were validated or corrected manually when necessary.
“Hepatocyte nuclei were detected automatically with the “analyze particles” feature of Fiji (ImageJ) software to encircle GFP- or HNF4α-positive nuclei and selected regions were validated or corrected manually when necessary.”
Quote in the paperNot a substantial claimClassified as probably not a substantial claim.
Study design and conductnone givenFirefly luciferase activity was normalized to Renilla luciferase in each sample.
“Firefly luciferase activity was normalized to Renilla luciferase in each sample.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationQuiescence is commonly imposed by inhibitors that prevent cells from entering S phase.
“Quiescence is commonly imposed by inhibitors that prevent cells from entering S phase (2).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [2] B. D. Manning, L. C. Cantley, AKT/PKB signaling: Navigating downstream. Cell 129, 1261– 1274 (2007). 17604717 PMC2756685 source
Probably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductnone givenDoublets were excluded from flow cytometry analysis by forward-scatter width/height and side-scatter width/height gating.
“Doublets were excluded by forward-scatter width (FSC-W)/ forward-scatter height (FSC-H) and side-scatter width (SSC-W) / side-scatter height (SSC-H) analysis.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationCells in adult tissues remain mostly quiescent unless activated by growth factors.
“Cells in adult tissues remain mostly quiescent unless they become activated by growth factors to enter the cell cycle and proliferate (1).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [1] S. M. Jones, A. Kazlauskas, Growth factor-dependent signaling and cell cycle progression. FEBS Lett. 490, 110– 116 (2001). 11223025 source
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
PremisecitationGrowth factors repress inhibitors and activate the G1/S transition to allow cell cycle entry.
“To allow cell cycle entry, growth factors repress these inhibitors and activate the G1/S transition (3).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites [3] H. A. Coller, What’s taking so long? S-phase entry from quiescence versus proliferation. Nat. Rev. Mol. Cell Biol. 8, 667– 670 (2007). 17637736 source
Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.
Study design and conductnone givenSingle-stained channels were used for compensation and a fluorophore-minus-one control was used for gating in flow cytometry.
“Single-stained channels were used for compensation and fluorophore minus one control was used for gating.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductnone givenA rat IgG1 isotype control was used as the IgG control in chromatin immunoprecipitation.
“Rat IgG1 isotype control (Thermo Fisher MA1-90035) was used for IgG control.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductown dataGapdh promoter primers were used as internal controls in ChIP-qPCR.
“Forward: 5′-AAGGTCATCCCAGAGCTGAA-3′ and reverse: 5′-CTGCTTCACCACCTTCTTGA-3′ primers detecting Gapdh promoter area were used as internal controls.”
Quote in the paperNot a substantial claim
The authors state this as an assumption.
Whether mechanisms analogous to S-phase repressors operate in M phase was unknown.
“It was unknown whether similar mechanisms operate in the M phase.”
“Whether growth factors can suppress the activity of E2f7 and E2f8 and thereby promote cell division is currently not known.”
The paper relies on this without support.
CHIR99021 activates Wnt signaling by inhibiting GSK3.
“CHIR99021, a GSK3 inhibitor that activates Wnt signaling”
Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
Each contradiction between two passages listed here was confirmed by an independent check.
E2f8 TBX3-binding site is called both an enhancer and a promoter
MinorThe results identify the tested E2f8 TBX3-binding site as an enhancer, whereas the ChIP-qPCR methods call the E2f8 site a promoter area.
“We found that the TBX3 protein binds to promoter and enhancer regions of E2f7 and E2f8, respectively, and verified this interaction by ChIP-qPCR (Fig. 3A and B).”
“E2f7 forward: 5′-CGCCAGAGGAGGTGTTTTAG-3′ and reverse: 5′-CGGGCAGAGCGTGTAGAT-3′ or E2f8 forward: 5′-GAACACTTGGGTGACCCTGA-3′ and reverse: 5′-CAAAGGGAATGCACACTGG-3′ were used to detect the TBX3-bound E2f7 or E2f8 promoter areas, respectively.”
Cited works that were retracted
Works this paper cites that were later retracted, from the retraction notices publishers deposit. Major when the notice came out before this paper was published.
Cites a retracted work: 10.3727/096504017x15145624664031
InfoThe cited work was retracted on 2026-09-14 (notice 10.32604/or.2026.092122), not before this paper was published (2022-07-22).
Cited work 10.3727/096504017x15145624664031 · retracted 2026-09-14 · notice 10.32604/or.2026.092122 · this paper published 2022-07-22
“To verify that repression of E2f7 and E2f8 by TBX3 occurs on the regulatory sequences of these genes, we employed a luciferase reporter assay in human hepatoblastoma HepG2 cells, which express TBX3 at high levels (32).”
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“To verify that repression of E2f7 and E2f8 by TBX3 occurs on the regulatory sequences of these genes, we employed a luciferase reporter assay in human hepatoblastoma HepG2 cells, which express TBX3 at high levels (32).”
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BackgroundThe Encyclopedia of DNA elements (ENCODE): data portal update
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BackgroundMitotic and mitogenic Wnt signalling
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Synergistic Function of E2F7 and E2F8 Is Essential for Cell Survival and Embryonic Development
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“E2f7; E2f8 were gifted by Alain de Bruin (Utrecht University, Utrecht, The Netherlands) (60) and were rederived at the Stanford Transgenic Facility.”
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BackgroundPolyploidy in liver development, homeostasis and disease
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The Polyploid State Plays a Tumor-Suppressive Role in the Liver
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Efficient temporally controlled targeted somatic mutagenesis in hepatocytes of the mouse
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“Alb-CreERT2 mice were a gift from Julien Sage (Stanford University, Stanford, CA) (59).”
MethodPolyploidy in tissue homeostasis and regeneration
Øvrebø, J. I., & Edgar, B. A. (2018) · Development · doi:10.1242/dev.156034 · Cited by 171 · see citing works
E2F8 is essential for polyploidization in mammalian cells
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“Deleting E2f7 and E2f8 from hepatocytes results in completion of M phase and the generation of diploid daughter cells (7, 8).”
BackgroundCanonical and atypical E2Fs regulate the mammalian endocycle
Chen, H.-Z., Ouseph, M. M., Li, J., Pécot, T., Chokshi, V., Kent, L., Bae, S., Byrne, M., Duran, C., Comstock, G., et al. (2012) · Nature Cell Biology · doi:10.1038/ncb2595 · Cited by 161 · cited 3× · see citing works
“Deleting E2f7 and E2f8 from hepatocytes results in completion of M phase and the generation of diploid daughter cells (7, 8).”
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“In hepatic tumor cells with activating β-catenin mutations, Tbx3 is overexpressed and mediates cell proliferation and survival (25).”
BackgroundThe Glucocorticoid Receptor Controls Hepatic Dyslipidemia through Hes1
Lemke, U., Krones, A., Díaz, M. B., Narvekar, P., Ziegler, A., Vegiopoulos, A., Cato, A. C. B., Böhl, S., Klingmüller, U., Screaton, R. A., et al. (2008) · Cell Metabolism · doi:10.1016/j.cmet.2008.08.001 · Cited by 151 · cited 2× · see citing works
“Hepatocyte nuclei preparation method was developed by modifying the chromatin preparation protocols described previously (63, 64).”
MethodGeneration of mice with a conditional null allele for Wntless
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“1Lan/J) (57) strains were obtained from The Jackson Laboratory (JAX).”
MethodWnt/β-Catenin Signaling in Liver Cancers
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“Interestingly, activating mutations in Wnt signaling components are common in liver cancer (41, 42), while E2f7 and E2f8, as well as polyploidy, are known to suppress tumorigenesis (43–46).”
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“Tbx3 also maintains expression of hepatocyte-lineage genes, such as Cebpα and Hnf4α, and represses the cholangiocyte fate (23, 24).”
BackgroundGrowth factor‐dependent signaling and cell cycle progression
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β-Catenin is critical for early postnatal liver growth
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“To allow cell cycle entry, growth factors repress these inhibitors and activate the G1/S transition (3).”
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“Tbx3-expressing cells may proliferate at different rates or capacities in the context of injury when additional cell cycle inducers promote tissue repair (19).”
BackgroundSelf-citationE2f8 mediates tumor suppression in postnatal liver development
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Dual modulation of human hepatic zonation via canonical and non-canonical Wnt pathways
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Dynamic zonation of liver polyploidy
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“Moreover, Tbx3negative hepatocytes in the periportal zone seem to maintain low ploidy levels through the course of the animal’s lifetime (51).”
BackgroundPolyploidy and liver proliferation
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A Dynamic Role of TBX3 in the Pluripotency Circuitry
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“Tbx3, which has been shown to act in a dosage-sensitive manner in other contexts (50), seems to be expressed in a gradient in the pericentral zone (SI Appendix, Fig.”
BackgroundImpaired neural development caused by inducible expression of Axin in transgenic mice
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Polyploid Hepatocytes Facilitate Adaptation and Regeneration to Chronic Liver Injury
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Tbx3 Controls Dppa3 Levels and Exit from Pluripotency toward Mesoderm
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TBX3 acts as tissue-specific component of the Wnt/β-catenin transcriptional complex
Zimmerli, D., Borrelli, C., Jauregi-Miguel, A., Söderholm, S., Brütsch, S., Doumpas, N., Reichmuth, J., Murphy-Seiler, F., Aguet, M., Basler, K., et al. (2020) · eLife · doi:10.7554/elife.58123 · Cited by 47 · see citing works
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“A role for Wnt signaling during M phase has also been suggested based on high Wnt/β-catenin signaling activity during cell division, through an unknown mechanism (39, 40).”
BackgroundLoss of hepatocyte cell division leads to liver inflammation and fibrosis
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“In the absence of a capacity for cell division, fibrosis becomes a mode of tissue repair and leads to tissue scarring (52, 53).”
BackgroundAydoğdu, N., Rudat, C., Trowe, M.-O., Kaiser, M., Lüdtke, T. H., Taketo, M. M., Christoffels, V. M., Moon, A., & Kispert, A. (2018) · Development · doi:10.1242/dev.171827 · Cited by 41 · see citing works
Mouse Tbx3 Mutants Suggest Novel Molecular Mechanisms for Ulnar-Mammary Syndrome
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“Tbx3 mice were a gift from Anne Moon (Geisinger Clinic, Danville, PA) (58).”
MethodThe origins and functions of hepatic polyploidy
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“Expression of E2f7 and E2f8 is induced at the time of weaning when the tissue switches from initially containing diploid hepatocytes to becoming mostly polyploid (4, 9, 10).”
BackgroundE2F7 Is a Potent Inhibitor of Liver Tumor Growth in Adult Mice
Moreno, E., Toussaint, M. J. M., van Essen, S. C., Bongiovanni, L., van Liere, E. A., Koster, M. H., Yuan, R., van Deursen, J. M., Westendorp, B., & de Bruin, A. (2021) · Hepatology · doi:10.1002/hep.31259 · Cited by 37 · see citing works
Analyzing Transcription Factor Occupancy During Embryo Development Using ChIP-seq
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“Hepatocyte nuclei preparation method was developed by modifying the chromatin preparation protocols described previously (63, 64).”
MethodAtypical E2f functions are critical for pancreas polyploidization
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The role of E2F8 in the human placenta
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“E2f8 (gene ID: 108961) is located in the cell nucleus, and participates in the regulation of hepatocytes [32] and anti-tumor CD4+ T helper cells [33].”
BackgroundThe Time-Resolved Genomic Impact of WNT/ β-Catenin Signaling
Pagella, P., Söderholm, S., Nordin, A., Zambanini, G., Ghezzi, V., Jauregi‐Miguel, A., & Cantù, C. (2022) · Cell Systems · doi:10.1016/j.cels.2023.06.004 · Cited by 26 · see citing works
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“Previous studies have demonstrated that Tbx3 regulates division of hepatocytes during liver development by repressing cell cycle inhibitors(Jin et al., 2022; Suzuki, Sekiya, Buscher, Izpisua Belmonte, & Taniguchi, 2008).”
BackgroundSelf-citationThe G 1 -S transition is promoted by Rb degradation via the E3 ligase UBR5
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Design principles and therapeutic applications of novel synthetic WNT signaling agonists
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“44,47–49 Genetic pathway inactivation in hepatocytes decreases liver size in mice and significantly delays hepatocyte proliferation and liver regeneration after partial hepatectomy (PH).”
BackgroundBinucleated human hepatocytes arise through late cytokinetic regression during endomitosis M phase
Darmasaputra, G. S., Geerlings, C. C., Lopes, S. M. C. D. S., Clevers, H., & Galli, M. (2024) · Journal of Cell Biology · doi:10.1083/jcb.202403020 · Cited by 12 · see citing works
“In line with previous research in rodents (Wang et al., 2015; Jin et al., 2022), these findings suggest that E2F7 and E2F8 promote binucleation downstream of WNT signaling in human Hep-Orgs.”
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“While TCF7L1 is closely related to TCF7L2 and is a member of the TCF/LEF family of transcription factors, TBX3 also is a downstream target of Wnt signaling, and liver-speci fi c deletion of Tbx3 promotes hepatic fi brosis 52 .”
Wu, P. V., Fish, M., Hazard, F. K., Zhu, C., Vennam, S., Walton, H., Wagh, D. A., Coller, J., Przybył, J., Morri, M., et al. (2024) · Nature Communications · doi:10.1038/s41467-024-53802-4 · Cited by 10 · see citing works
“In the normal liver, Wnt signals from endothelial cells in the central vein act on neighboring hepatocytes to promote mitosis 28 .”
Self-citationYang, J., Davidoff, A. M., & Murphy, A. J. (2025) · Molecular Cancer · doi:10.1186/s12943-025-02405-8 · Cited by 9 · see citing works
“Second, WNT released from the endothelial cells of the liver central vein promotes hepatocyte mitosis by inhibiting E2F7/ E2F8 via the WNT target TBX3 [50–52].”
Hong, X., Zeng, S., Wang, M., Wang, Y., Cao, Y., Qu, Y., Huang, X., Li, X., Qiao, Q., & Zhang, J. (2025) · Cancer Letters · doi:10.1016/j.canlet.2025.218193 · Cited by 6 · see citing works
Ferrando-Marco, M., & Barkoulas, M. (2025) · Development · doi:10.1242/dev.204546 · Cited by 3 · see citing works
Somatic mutations in TBX3 promote hepatic clonal expansion by accelerating VLDL secretion
Mannino, G., Quinn, G., Zhu, M., Wang, Z., Wang, X., Li, B., Hsieh, M.-H., Mathews, T., Zacharias, L., Gu, W., et al. (2025) · Journal of Clinical Investigation · doi:10.1172/jci191855 · Cited by 4 · see citing works
“Within the liver, Tbx3 is expressed in zone 3 hepatocytes, downstream of canonical Wnt signaling (16).”
Hua, Q., Zhang, X.-C., Tian, R.-F., She, Z.-G., & Huang, Z. (2025) · Journal of Zhejiang University SCIENCE B · doi:10.1631/jzus.b2400127 · Cited by 4 · see citing works
“In addition, wingless-type mouse mam‐ mary tumor virus (MMTV) integration site family (WNT) signaling inhibits binucleation and polyploidi‐ zation of hepatocytes in an E2F7/8-dependent manner both in a mouse model and a human hepatocyte or‐ ganoid model (Jin et al., 2022; Darmasaputra et al., 2024).”
PRDM14 extinction enables the initiation of trophoblast stem cell formation
Xu, C., Zhao, W., Peng, L., Yin, T., Guo, J., Liu, L., Liu, L., Yang, J., Du, M., & Du, M. (2024) · Cellular and Molecular Life Sciences · doi:10.1007/s00018-024-05237-9 · Cited by 4 · see citing works
“Given that Wnt/β-catenin signaling can cause the expression of some repressors, which in turn can exert repressive effects on other genes [31], it is proposed that Wnt/β-catenin signaling might reduce PRDM14 expression in an indirect manner.”
BackgroundLiang, Z.-X., Li, S., Wang, Z.-Y., Zhou, J., Huang, Z.-Y., Li, J.-H., Bao, H., Yam, J., & Xu, Y. (2025) · Journal of Clinical and Translational Hepatology · doi:10.14218/jcth.2024.00401 · Cited by 11 · see citing works
“104,105 Invasion and metastasis: The increased malignant biological behavior of HCC patients is a major cause of their high mortality rate.”
GSK3β/NF-κB -dependent transcriptional regulation of homeostatic hepatocyte Tnf production
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Light-manipulated binary droplet transport on a high-energy surface
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“…of LGR5+ cell in the intestinal epithelium (Sato et al., 2011) or the proliferative potential of cells close to the central vein in the liver (Jin et al., 2022), could generate a short-term, elastic response that drives the expression of groups of genes required to sustain that specific…”
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Vascularization of neonatal liver lobules presages adult liver size
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Self-citationRole of T-box transcription factor 3 in gastric cancers
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“Moreover, as downstream effectors of the Wnt/ β -catenin signaling pathway, E2f7 and E2f8 also govern hepatocyte mitosis and proliferation (Jin et al., 2022).”
Molecular regulation of plant endoreduplication: core mechanisms and their upstream control
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- Authors
- Jin, Y., Anbarchian, T., Wu, P., Sarkar, A., Fish, M., Peng, W. C., & Nusse, R.
- Affiliations
- Howard Hughes Medical Institute (US)
- Department of Developmental Biology, Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305
- Venue
- Proceedings of the National Academy of Sciences
- Publisher
- National Academy of Sciences
- Published
- 2022-07-22
- Volume
- 119
- Issue
- 30
- Pages
- e2203849119-e2203849119
- ISSN
- 0027-8424 (ISSN-L)
- 1091-6490
- PMC
- PMC9335208
- OpenAlex
- W4286587461
- Semantic Scholar
- 1f637c3de202ec366ffe49662f098189211fb7e8
- Cited by
- 26
- References
- 73
- Citations by year
- 2026: 3 · 2025: 8 · 2024: 6 · 2023: 4 · 2022: 5
- FWCI
- 1.59
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Proceedings of the National Academy of Sciences (publishedVersion, cc-by)
- PubMed (publishedVersion)
- PubMed Central (submittedVersion, cc-by)
- Funders
- Howard Hughes Medical Institute (no number)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Wnt/β-catenin signaling in development and cancer
- Health Sciences › Medicine › Hepatology › Liver physiology and pathology
- Health Sciences › Medicine › Oncology › Cancer-related Molecular Pathways
- Keywords
- Wnt/β-catenin signaling, TBX3, E2F7, E2F8, transcriptional repression, cell cycle progression, polyploidy, postnatal liver development, mitosis
- MeSH terms
- Animals, Mitosis, Repressor Proteins, T-Box Domain Proteins, Hepatocytes, beta Catenin, Mice, Wnt Signaling Pathway
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-22.
Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-22.
Cited sources are checked by gpt-6-sol.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
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