Conditional Gata4 deletion in mice induces bile acid absorption in the proximal small intestine.

BACKGROUND AND AIMS: The transcription factor GATA4 is expressed throughout most of the small intestine except distal ileum, and restricts expression of the apical sodium-dependent bile acid transporter (ASBT), the rate-limiting intestinal bile acid transporter, to distal ileum. The hypothesis was tested that reduction of GATA4 activity in mouse small intestine results in an induction of bile acid transport in proximal small intestine sufficient to restore bile acid absorption and homeostasis after ileocaecal resection (ICR). METHODS: Bile acid homeostasis was characterised in non-surgical, sham or ICR mice using two recombinant Gata4 models in which Asbt expression is induced to different levels. RESULTS: Reduction of intestinal GATA4 activity resulted in an induction of ASBT expression, bile acid absorption and expression of bile acid-responsive genes in proximal small intestine, and a reduction of luminal bile acids in distal small intestine. While faecal bile acid excretion and bile acid pool size remained unchanged, the bile acid pool became more hydrophilic due to a relative increase in tauro-beta-muricholate absorption. Furthermore, proximal induction of Asbt in both Gata4 mutant models corrected ICR-associated bile acid malabsorption, reversing the decrease in bile acid pool size and increase in faecal bile acid excretion and hepatic cholesterol 7alpha-hydroxylase expression. CONCLUSIONS: Reduction of intestinal GATA4 activity induces bile acid absorption in proximal small intestine without inducing major changes in bile acid homeostasis. This induction is sufficient to correct bile acid malabsorption caused by ICR in mice.

GATA4 mediates gene repression in the mature mouse small intestine through interactions with friend of GATA (FOG) cofactors.

GATA4, a transcription factor expressed in the proximal small intestine but not in the distal ileum, maintains proximal-distal distinctions by multiple processes involving gene repression, gene activation, and cell fate determination. Friend of GATA (FOG) is an evolutionarily conserved family of cofactors whose members physically associate with GATA factors and mediate GATA-regulated repression in multiple tissues. Using a novel, inducible, intestine-specific Gata4 knock-in model in mice, in which wild-type GATA4 is specifically inactivated in the small intestine, but a GATA4 mutant that does not bind FOG cofactors (GATA4ki) continues to be expressed, we found that ileal-specific genes were significantly induced in the proximal small intestine (P<0.01); in contrast, genes restricted to proximal small intestine and cell lineage markers were unaffected, indicating that GATA4-FOG interactions contribute specifically to the repression function of GATA4 within this organ. Fog1 mRNA displayed a proximal-distal pattern that parallels that of Gata4, and FOG1 protein was co-expressed with GATA4 in intestinal epithelial cells, implicating FOG1 as the likely mediator of GATA4 function in the small intestine. Our data are the first to indicate FOG function and expression in the mammalian small intestine.

Gata4 and Hnf1alpha are partially required for the expression of specific intestinal genes during development.

The terminal differentiation phases of intestinal development in mice occur during cytodifferentiation and the weaning transition. Lactase-phlorizin hydrolase (LPH), liver fatty acid binding protein (Fabp1), and sucrase-isomaltase (SI) are well-characterized markers of these transitions. With the use of gene inactivation models in mature mouse jejunum, we have previously shown that a member of the zinc finger transcription factor family (Gata4) and hepatocyte nuclear factor-1alpha (Hnf1alpha) are each indispensable for LPH and Fabp1 gene expression but are both dispensable for SI gene expression. In the present study, we used these models to test the hypothesis that Gata4 and Hnf1alpha regulate LPH, Fabp1, and SI gene expression during development, specifically focusing on cytodifferentiation and the weaning transition. Inactivation of Gata4 had no effect on LPH gene expression during either cytodifferentiation or suckling, whereas inactivation of Hnf1alpha resulted in a 50% reduction in LPH gene expression during these same time intervals. Inactivation of Gata4 or Hnf1alpha had a partial effect ( approximately 50% reduction) on Fabp1 gene expression during cytodifferentiation and suckling but no effect on SI gene expression at any time during development. Throughout the suckling period, we found a surprising and dramatic reduction in Gata4 and Hnf1alpha protein in the nuclei of absorptive enterocytes of the jejunum despite high levels of their mRNAs. Finally, we show that neither Gata4 nor Hnf1alpha mediates the glucocorticoid-induced precocious maturation of the intestine but rather are downstream targets of this process. Together, these data demonstrate that specific intestinal genes have differential requirements for Gata4 and Hnf1alpha that are dependent on the developmental time frame in which they are expressed.

Gata4 is essential for the maintenance of jejunal-ileal identities in the adult mouse small intestine.

Gata4, a member of the zinc finger family of GATA transcription factors, is highly expressed in duodenum and jejunum but is nearly undetectable in distal ileum of adult mice. We show here that the caudal reduction of Gata4 is conserved in humans. To test the hypothesis that the regional expression of Gata4 is critical for the maintenance of jejunal-ileal homeostasis in the adult small intestine in vivo, we established an inducible, intestine-specific model that results in the synthesis of a transcriptionally inactive Gata4 mutant. Synthesis of mutant Gata4 in jejuna of 6- to 8-week-old mice resulted in an attenuation of absorptive enterocyte genes normally expressed in jejunum but not in ileum, including those for the anticipated targets liver fatty acid binding protein (Fabp1) and lactase-phlorizin hydrolase (LPH), and a surprising induction of genes normally silent in jejunum but highly expressed in ileum, specifically those involved in bile acid transport. Inactivation of Gata4 resulted in an increase in the goblet cell population and a redistribution of the enteroendocrine subpopulations, all toward an ileal phenotype. The gene encoding Math1, a known activator of the secretory cell fate, was induced approximately 75% (P < 0.05). Gata4 is thus an important positional signal required for the maintenance of jejunal-ileal identities in the adult mouse small intestine.

Hepatocyte nuclear factor-1alpha is required for expression but dispensable for histone acetylation of the lactase-phlorizin hydrolase gene in vivo.

Hepatocyte nuclear factor-1alpha (HNF-1alpha) is a modified homeodomain-containing transcription factor that has been implicated in the regulation of intestinal genes. To define the importance and underlying mechanism of HNF-1alpha for the regulation of intestinal gene expression in vivo, we analyzed the expression of the intestinal differentiation markers and putative HNF-1alpha targets lactase-phlorizin hydrolase (LPH) and sucrase-isomaltase (SI) in hnf1alpha null mice. We found that in adult jejunum, LPH mRNA in hnf1alpha(-/-) mice was reduced 95% compared with wild-type controls (P < 0.01, n = 4), whereas SI mRNA was virtually identical to that in wild-type mice. Furthermore, SI mRNA abundance was unchanged in the absence of HNF-1alpha along the length of the adult mouse small intestine as well as in newborn jejunum. We found that HNF-1alpha occupies the promoters of both the LPH and SI genes in vivo. However, in contrast to liver and pancreas, where HNF-1alpha regulates target genes by recruitment of histone acetyl transferase activity to the promoter, the histone acetylation state of the LPH and SI promoters was not affected by the presence or absence of HNF-1alpha. Finally, we showed that a subset of hypothesized intestinal target genes is regulated by HNF-1alpha in vivo and that this regulation occurs in a defined tissue-specific and developmental context. These data indicate that HNF-1alpha is an activator of a subset of intestinal genes and induces these genes through an alternative mechanism in which it is dispensable for chromatin remodeling.