GATA4 Controls Epithelial Morphogenesis in the Developing Stomach to Promote Establishment of Glandular Columnar Epithelium.

BACKGROUND & AIMS: The transcription factor GATA4 is broadly expressed in nascent foregut endoderm. As development progresses, GATA4 is lost in the domain giving rise to the stratified squamous epithelium of the esophagus and forestomach (FS), while it is maintained in the domain giving rise to the simple columnar epithelium of the hindstomach (HS). Differential GATA4 expression within these domains coincides with the onset of distinct tissue morphogenetic events, suggesting a role for GATA4 in diversifying foregut endoderm into discrete esophageal/FS and HS epithelial tissues. The goal of this study was to determine how GATA4 regulates differential morphogenesis of the mouse gastric epithelium. METHODS: We used a Gata4 conditional knockout mouse line to eliminate GATA4 in the developing HS and a Gata4 conditional knock-in mouse line to express GATA4 in the developing FS. RESULTS: We found that GATA4-deficient HS epithelium adopted a FS-like fate, and conversely, that GATA4-expressing FS epithelium adopted a HS-like fate. Underlying structural changes in these epithelia were broad changes in gene expression networks attributable to GATA4 directly activating or repressing expression of HS or FS defining transcripts. Our study implicates GATA4 as having a primary role in suppressing an esophageal/FS transcription factor network during HS development to promote columnar epithelium. Moreover, GATA4-dependent phenotypes in developmental mutants reflected changes in gene expression associated with Barrett's esophagus. CONCLUSIONS: This study demonstrates that GATA4 is necessary and sufficient to activate the development of simple columnar epithelium, rather than stratified squamous epithelium, in the embryonic stomach. Moreover, similarities between mutants and Barrett's esophagus suggest that developmental biology can provide insight into human disease mechanisms.

Chromatin establishes an immature version of neuronal protocadherin selection during the naive-to-primed conversion of pluripotent stem cells.

In the mammalian genome, the clustered protocadherin (cPCDH) locus provides a paradigm for stochastic gene expression with the potential to generate a unique cPCDH combination in every neuron. Here we report a chromatin-based mechanism that emerges during the transition from the naive to the primed states of cell pluripotency and reduces, by orders of magnitude, the combinatorial potential in the human cPCDH locus. This mechanism selectively increases the frequency of stochastic selection of a small subset of cPCDH genes after neuronal differentiation in monolayers, 10-month-old cortical organoids and engrafted cells in the spinal cords of rats. Signs of these frequent selections can be observed in the brain throughout fetal development and disappear after birth, except in conditions of delayed maturation such as Down's syndrome. We therefore propose that a pattern of limited cPCDH-gene expression diversity is maintained while human neurons still retain fetal-like levels of maturation.

GATA4 regulates epithelial cell proliferation to control intestinal growth and development in mice.

BACKGROUND & AIMS: The embryonic small intestinal epithelium is highly proliferative, and although much is known about mechanisms regulating proliferation in the adult intestine, the mechanisms controlling epithelial cell proliferation in the developing intestine are less clear. GATA4, a transcription factor that regulates proliferation in other developing tissues, is first expressed early in the developing gut in midgut endoderm. GATA4 function within midgut endoderm and the early intestinal epithelium has not been investigated. METHODS: Using Sonic Hedgehog Cre to eliminate GATA4 in the midgut endoderm of mouse embryos, we determined the impact of loss of GATA4 on intestinal development, including epithelial cell proliferation, between E9.5-E18.5. RESULTS: We found that intestinal length and width were decreased in GATA4 mutants compared with controls. GATA4-deficient intestinal epithelium contained fewer cells, and epithelial girth was decreased. We further observed a decreased proportion of proliferating cells at E10.5 and E11.5 in GATA4 mutants. We demonstrated that GATA4 binds to chromatin containing GATA4 consensus binding sites within Cyclin D2 (Ccnd2), Cyclin dependent kinase 6 (Cdk6), and Frizzled 5 (Fzd5). Moreover, Ccnd2, Cdk6, and Fzd5 transcripts were reduced at E11.5 in GATA4 mutant tissue. Villus morphogenesis was delayed, and villus structure was abnormal in GATA4 mutant intestine. CONCLUSIONS: Our data identify GATA4 as an essential regulator of early intestinal epithelial cell proliferation. We propose that GATA4 controls proliferation in part by directly regulating transcription of cell cycle mediators. Our data further suggest that GATA4 affects proliferation through transcriptional regulation of Fzd5, perhaps by influencing the response of the epithelium to WNT signaling.

Von Willebrand factor is reversibly decreased during torpor in 13-lined ground squirrels.

During torpor in a hibernating mammal, decreased blood flow increases the risk of blood clots such as deep vein thrombi (DVT). In other animal models platelets, neutrophils, monocytes and von Willebrand factor (VWF) have been found in DVT. Previous research has shown that hibernating mammals decrease their levels of platelets and clotting factors VIII (FVIII) and IX (FIX), increasing both bleeding time and activated partial thromboplastin time. In this study, FVIII, FIX and VWF activities and mRNA levels were measured in torpid and non-hibernating ground squirrels (Ictidomys tridecemlineatus). Here, we show that VWF high molecular weight multimers, collagen-binding activity, lung mRNA and promoter activity decrease during torpor. The VWF multimers reappear in plasma within 2 h of arousal in the spring. Similarly, FIX activity and liver mRNA both dropped threefold during torpor. In contrast, FVIII liver mRNA levels increased twofold while its activity dropped threefold, consistent with a post-transcriptional decrease in FVIII stability in the plasma due to decreased VWF levels. Finally, both neutrophils and monocytes are decreased eightfold during torpor which could slow the formation of DVT. In addition to providing insight in how blood clotting can be regulated to allow mammals to survive in extreme environments, hibernating ground squirrels provide an interesting model for studying.

Characterization of the developing small intestine in the absence of either GATA4 or GATA6.

BACKGROUND: Studies of adult mice lacking either GATA4 or GATA6 in the small intestine demonstrate roles for these factors in small intestinal biology. Deletion of Gata4 in the adult mouse intestine revealed an essential role for GATA4 in jejunal function. Deletion of Gata6 in the adult mouse ileum alters epithelial cell types and ileal enterocyte gene expression. The effect of deletion of Gata4 or Gata6 alone during embryonic small intestinal development, however, has not been examined. We recently demonstrated that loss of both factors in double conditional knockout embryos causes severe defects in jejunal development. Therefore, the goal of this study is to provide phenotypic analysis of the small intestine of single Gata4 and Gata6 conditional knockout embryos. RESULTS: Villin-Cre was used to delete Gata4 or Gata6 in the developing intestinal epithelium. Elimination of either GATA4 or GATA6 in the jejunum, where these factors are co-expressed, caused changes in enterocyte and enteroendocrine cell gene expression. Ectopic expression of markers of the ileal-specific bile acid metabolism pathway was induced in GATA4-deficient jejunum but not in GATA6-deficient jejunum. A subtle increase in goblet cells was also identified in jejunum of both mutants. In GATA6-deficient embryonic ileum, villus length was altered, and enterocyte gene expression was perturbed including ectopic expression of the colon marker Car1. Goblet cells were increased, and enteroendocrine cells were decreased. CONCLUSIONS: Overall, we show that aspects of the phenotypes observed in the small intestine of adult Gata4 and Gata6 conditional knockout mice emerge during development. The effect of eliminating GATA6 from the developing ileum was greater than that of eliminating either GATA4 or GATA6 from the developing jejunum likely reflecting functional redundancy between these factors in the jejunum. Although GATA4 and GATA6 functions overlap, our data also suggest unique functions for GATA4 and GATA6 within the developing intestine. GATA4 likely operates independently of GATA6 within the jejunum to regulate jejunal versus ileal enterocyte identity and consequently jejunal physiology. GATA6 likely regulates enteroendocrine cell differentiation cell autonomously whereas GATA4 affects this population indirectly.