Summary Production of bile by the liver is crucial for the absorption of lipophilic nutrients. Dysregulation of bile acid homeostasis can lead to cholestatic liver disease and ER stress. We show using global location analysis (“ChIP-on-Chip”) and cell-type specific gene ablation that the winged helix transcription factor Foxa2 is required for normal bile acid homeostasis. As suggested by the location analysis, deletion of Foxa2 in hepatocytes in Foxa2loxP/loxPAlfp.Cre mice leads to decreased transcription of genes encoding bile acid transporters on both the basolateral and canalicular membranes, resulting in intrahepatic cholestasis. Foxa2-deficient mice are strikingly sensitive to a diet containing cholic acid, which results in toxic accumulation of hepatic bile salts, ER stress, and liver injury. In addition, we demonstrate that expression of FOXA2 is dramatically decreased in liver samples from patients with different cholestatic syndromes, suggesting that reduced FOXA2 levels could exacerbate the injury.

Summary Aging is accompanied by physiological impairments, which, in insulin-responsive tissues, including the liver, predispose individuals to metabolic disease. However, the molecular mechanisms underlying these changes remain largely unknown. Here, we analyze genome-wide profiles of RNA and chromatin organization in the liver of young (3 months) and old (21 months) mice. Transcriptional changes suggest that de-repression of the nuclear receptors PPARα, PPARγ, and LXRα in aged mouse liver leads to activation of targets regulating lipid synthesis and storage, whereas age-dependent changes in nucleosome occupancy are associated with binding sites for both known regulators (forkhead factors and nuclear receptors) and for novel candidates associated with nuclear lamina (Hdac3 and Srf) implicated to govern metabolic function of aging liver. Winged-helix transcription factor Foxa2 and nuclear receptor co-repressor Hdac3 exhibit reciprocal binding pattern at PPARα targets contributing to gene expression changes that lead to steatosis in aged liver.

While the molecular mechanisms of glucocorticoid regulation of transcription have been studied in detail, the global networks regulated by the glucocorticoid receptor (GR) remain unknown. To address this question, we performed an orthogonal analysis to identify direct targets of the GR. First, we analyzed the expression profile of mouse livers in the presence or absence of exogenous glucocorticoid, resulting in over 1,300 differentially expressed genes. We then executed genome-wide location analysis on chromatin from the same livers, identifying more than 300 promoters that are bound by the GR. Intersecting the two lists yielded 53 genes whose expression is functionally dependent upon the ligand-bound GR. Further network and sequence analysis of the functional targets enabled us to suggest interactions between the GR and other transcription factors at specific target genes. Together, our results further our understanding of the GR and its targets, and provide the basis for more targeted glucocorticoid therapies.

Gene duplication is a powerful driver of evolution. Newly duplicated genes acquire new roles that are relevant to fitness, or they will be lost over time. A potential path to functional relevance is mutation of the coding sequence leading to the acquisition of novel biochemical properties, as analyzed here for the highly homologous paralogs Foxa1 and Foxa2 transcriptional regulators. We determine by genome-wide location analysis (ChIP-Seq) that, although Foxa1 and Foxa2 share a large fraction of binding sites in the liver, each protein also occupies distinct regulatory elements in vivo. Foxa1-only sites are enriched for p53 binding sites and are frequently found near genes important to cell cycle regulation, while Foxa2-restricted sites show only a limited match to the forkhead consensus and are found in genes involved in steroid and lipid metabolism. Thus, Foxa1 and Foxa2, while redundant during development, have evolved divergent roles in the adult liver, ensuring the maintenance of both genes during evolution.

Objective Type II nuclear hormone receptors, including farnesoid X receptors (FXR), liver X receptors (LXR), and peroxisome proliferator-activated receptors (PPAR), which serve as drug targets for metabolic diseases, are permanently positioned in the nucleus and thought to be bound to DNA regardless of the ligand status. However, recent genome-wide location analysis showed that LXRα and PPARα binding in the liver is largely ligand-dependent. We hypothesized that pioneer factor Foxa2 evicts nucleosomes to enable ligand-dependent binding of type II nuclear receptors and performed genome-wide studies to test this hypothesis. Methods ATAC-Seq was used to profile chromatin accessibility; ChIP-Seq was performed to assess transcription factors (Foxa2, FXR, LXRα, and PPARα) binding; and RNA-Seq analysis determined differentially expressed genes in wildtype and Foxa2 mutants treated with a ligand (GW4064 for FXR, GW3965, and T09 for LXRα). Results We reveal that chromatin accessibility, FXR binding, LXRα occupancy, and ligand-responsive activation of gene expression by FXR and LXRα require Foxa2. Unexpectedly, Foxa2 occupancy is drastically increased when either receptor, FXR or LXRα, is bound by an agonist. In addition, co-immunoprecipitation experiments demonstrate that Foxa2 interacts with either receptor in a ligand-dependent manner, suggesting that Foxa2 and the receptor, bind DNA as an interdependent complex during ligand activation. Furthermore, PPARα binding is induced in Foxa2 mutants treated with FXR and LXR ligands, leading to the activation of PPARα targets. Conclusions Our model requires pioneering activity for ligand activation that challenges the existing ligand-independent binding mechanism. We also demonstrate that Foxa2 is required to achieve activation of the proper receptor – one that binds the added ligand – by repressing the activity of a competing receptor.