SOX transcription factors direct TCF-independent WNT/β-catenin responsive transcription to govern cell fate in human pluripotent stem cells.

WNT/β-catenin signaling controls gene expression across biological contexts from development and stem cell homeostasis to diseases including cancer. How β-catenin is recruited to distinct enhancers to activate context-specific transcription is unclear, given that most WNT/ß-catenin-responsive transcription is thought to be mediated by TCF/LEF transcription factors (TFs). With time-resolved multi-omic analyses, we show that SOX TFs can direct lineage-specific WNT-responsive transcription during the differentiation of human pluripotent stem cells (hPSCs) into definitive endoderm and neuromesodermal progenitors. We demonstrate that SOX17 and SOX2 are required to recruit β-catenin to lineage-specific WNT-responsive enhancers, many of which are not occupied by TCFs. At TCF-independent enhancers, SOX TFs establish a permissive chromatin landscape and recruit a WNT-enhanceosome complex to activate SOX/ß-catenin-dependent transcription. Given that SOX TFs and the WNT pathway are critical for specification of most cell types, these results have broad mechanistic implications for the specificity of WNT responses across developmental and disease contexts.

Mukherjee S ，Luedeke DM ，McCoy L ，Iwafuchi M ，Zorn AM ... -  《Cell Reports》

GATA6 regulates WNT and BMP programs to pattern precardiac mesoderm during the earliest stages of human cardiogenesis.

Haploinsufficiency for GATA6 is associated with congenital heart disease (CHD) with variable comorbidity of pancreatic or diaphragm defects, although the etiology of disease is not well understood. Here, we used cardiac directed differentiation from human embryonic stem cells (hESCs) as a platform to study GATA6 function during early cardiogenesis. GATA6 loss-of-function hESCs had a profound impairment in cardiac progenitor cell (CPC) specification and cardiomyocyte (CM) generation due to early defects during the mesendoderm and lateral mesoderm patterning stages. Profiling by RNA-seq and CUT&RUN identified genes of the WNT and BMP programs regulated by GATA6 during early mesoderm patterning. Furthermore, interactome analysis detected GATA6 binding with developmental transcription factors and chromatin remodelers, suggesting cooperative regulation of cardiac lineage gene accessibility. We show that modulating WNT and BMP inputs during the first 48 hr of cardiac differentiation is sufficient to partially rescue CPC and CM defects in GATA6 heterozygous and homozygous mutant hESCs. This study provides evidence of the regulatory functions for GATA6 directing human precardiac mesoderm patterning during the earliest stages of cardiogenesis to further our understanding of haploinsufficiency causing CHD and the co-occurrence of cardiac and other organ defects caused by human GATA6 mutations.

Bisson JA ，Gordillo M ，Kumar R ，de Silva N ，Yang E ，Banks KM ，Shi ZD ，Lee K ，Yang D ，Chung WK ，Huangfu D ，Evans T ... -  《eLife》

Sox17 and β-catenin co-occupy Wnt-responsive enhancers to govern the endoderm gene regulatory network.

Lineage specification is governed by gene regulatory networks (GRNs) that integrate the activity of signaling effectors and transcription factors (TFs) on enhancers. Sox17 is a key transcriptional regulator of definitive endoderm development, and yet, its genomic targets remain largely uncharacterized. Here, using genomic approaches and epistasis experiments, we define the Sox17-governed endoderm GRN in Xenopus gastrulae. We show that Sox17 functionally interacts with the canonical Wnt pathway to specify and pattern the endoderm while repressing alternative mesectoderm fates. Sox17 and β-catenin co-occupy hundreds of key enhancers. In some cases, Sox17 and β-catenin synergistically activate transcription apparently independent of Tcfs, whereas on other enhancers, Sox17 represses β-catenin/Tcf-mediated transcription to spatially restrict gene expression domains. Our findings establish Sox17 as a tissue-specific modifier of Wnt responses and point to a novel paradigm where genomic specificity of Wnt/β-catenin transcription is determined through functional interactions between lineage-specific Sox TFs and β-catenin/Tcf transcriptional complexes. Given the ubiquitous nature of Sox TFs and Wnt signaling, this mechanism has important implications across a diverse range of developmental and disease contexts.

Mukherjee S ，Chaturvedi P ，Rankin SA ，Fish MB ，Wlizla M ，Paraiso KD ，MacDonald M ，Chen X ，Weirauch MT ，Blitz IL ，Cho KW ，Zorn AM ... -  《eLife》

Evaluating the transcriptional regulators of arterial gene expression via a catalogue of characterized arterial enhancers.

The establishment and growth of the arterial endothelium require the coordinated expression of numerous genes. However, regulation of this process is not yet fully understood. Here, we combined in silico analysis with transgenic mice and zebrafish models to characterize arterial-specific enhancers associated with eight key arterial identity genes (Acvrl1/Alk1, Cxcr4, Cxcl12, Efnb2, Gja4/Cx37, Gja5/Cx40, Nrp1, and Unc5b). Next, to elucidate the regulatory pathways upstream of arterial gene transcription, we investigated the transcription factors binding each arterial enhancer compared to a similar assessment of non-arterial endothelial enhancers. These results found that binding of SOXF and ETS factors was a common occurrence at both arterial and pan-endothelial enhancers, suggesting neither are sufficient to direct arterial specificity. Conversely, FOX motifs independent of ETS motifs were over-represented at arterial enhancers. Further, MEF2 and RBPJ binding was enriched but not ubiquitous at arterial enhancers, potentially linked to specific patterns of behaviour within the arterial endothelium. Lastly, there was no shared or arterial-specific signature for WNT-associated TCF/LEF, TGFβ/BMP-associated SMAD1/5 and SMAD2/3, shear stress-associated KLF4, or venous-enriched NR2F2. This cohort of well-characterized and in vivo-verified enhancers can now provide a platform for future studies into the interaction of different transcriptional and signaling pathways with arterial gene expression.

Nornes S ，Bruche S ，Adak N ，McCracken IR ，De Val S ... -  《eLife》

Osteoclastic ATP6AP2 maintains β-catenin levels to prevent hyper-osteoclastic activation and trabecular bone-loss.

Osteoclast (OC) formation and bone resorption are regulated by several factors, including V-ATPase, Wnt/β-catenin, and RANKL/RANK signaling. ATP6AP2, also known as the prorenin receptor (PRR), is an accessory subunit of V-ATPase and a regulator of Wnt/β-catenin signaling. While the V-ATPase subunit ATP6AP1 is essential for OC formation and function, the role of ATP6AP2 in OC-lineage cells is less clear. Here, we provide evidence that ATP6AP2 plays a negative role in osteoclastogenesis and function, contrasting with the positive role of ATP6AP1. Mice with conditional KO (cKO) of ATP6AP2 in OCs (Atp6ap2LysM) exhibit trabecular bone loss, likely due to the increased osteoclastogenesis and activity, since bone formation rates (BFRs) are comparable to control mice. In vitro assays using bone marrow macrophages (BMMs) show that Atp6ap2LysM cultures have more RANKL-induced TRAP+ OC-like cells and increased bone resorptive activity. Further studies reveal that while RANKL signaling and V-ATPase activity are normal, ATP6AP2 KO OCs, but not BMMs, have reduced basal levels of Wnt/β-catenin pathway proteins, such as LRP5/6 and β-catenin, compared to controls. Wnt3A treatment induces β-catenin and suppresses OC formation in both control and ATP6AP2 KO OC-lineage cells, indicating that Wnt/β-catenin signaling negatively regulates OC-formation and operates independently of ATP6AP2. Overall, these results suggest that ATP6AP2 is critical for maintaining basal levels of LRP5/6 receptors and β-catenin in OCs, thus acting as a negative regulator of osteoclastogenesis and activation.

Chen L ，Xiong L ，Guo H ，Feng X ，Zhu X ，Xiong WC ... -  《-》