Members of the Mad family of bHLH-Zip proteins heterodimerize with Max to repress transcription in a sequence-specific manner. Transcriptional repression by Mad:Max heterodimers is mediated by ternary complex formation with either of the corepressors mSin3A or mSin3B. We report here that mSin3A is an in vivo component of large, heterogeneous multiprotein complexes and is tightly and specifically associated with at least seven polypeptides. Two of the mSin3A-associated proteins, p50 and p55, are highly related to the histone deacetylase HDAC1. The mSin3A immunocomplexes possess histone deacetylase activity that is sensitive to the specific deacetylase inhibitor trapoxin. mSin3A-targeted repression of a reporter gene is reduced by trapoxin treatment, suggesting that histone deacetylation mediates transcriptional repression through Mad-Max-mSin3A multimeric complexes.

The nuclear bile acid receptor FXR has been proposed to play a central role in the feedback repression of the gene encoding cholesterol 7 alpha-hydroxylase (CYP7A1), the first and rate-limiting step in the biosynthesis of bile acids. We demonstrate that FXR directly regulates expression of fibroblast growth factor-19 (FGF-19), a secreted growth factor that signals through the FGFR4 cell-surface receptor tyrosine kinase. In turn, FGF-19 strongly suppresses expression of CYP7A1 in primary cultures of human hepatocytes and mouse liver through a c-Jun N-terminal kinase (JNK)-dependent pathway. This signaling cascade defines a novel mechanism for feedback repression of bile acid biosynthesis and underscores the vital role of FXR in the regulation of multiple pathways of cholesterol catabolism in the liver.

Glucose is a fundamental metabolite, yet how cells sense and respond to changes in extracellular glucose concentration is not completely understood. We recently reported that the MondoA: Mlx dimeric transcription factor directly regulates glycolysis. In this article, we consider whether MondoA:Mlx complexes have a broader role in sensing and responding to glucose status. In their latent state, MondoA:Mlx complexes localize to the outer mitochondrial membrane, yet shuttle between the mitochondria and the nucleus. We show that MondoA:Mlx complexes accumulate in the nucleus in response to glucose and 2-deoxyglucose (2-DG). Furthermore, nuclear localization of MondoA:Mlx depends on the enzymatic activity of hexokinases. These enzymes catalyze conversion of glucose to glucose-6-phosphate (G6P), which is the first step in the glycolytic pathway. Together, these findings suggest that MondoA:Mlx monitors intracellular G6P concentration and translocates to the nucleus when levels of this key metabolite increase. Transcriptional profiling experiments demonstrate that MondoA is required for >75% of the 2-DG-induced transcription signature. We identify thioredoxin-interacting protein (TXNIP) as a direct and glucose-regulated MondoA:Mlx transcriptional target. Furthermore, MondoA:Mlx complexes, via their regulation of TXNIP, are potent negative regulators of glucose uptake. These studies suggest a key role for MondoA:Mlx complexes in the adaptive transcriptional response to changes in extracellular glucose concentration and peripheral glucose uptake. metabolism ͉ mitochondria ͉ transcription

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Acetylation of histones triggers association with bromodomaincontaining proteins that regulate diverse chromatin-related processes. Although acetylation of transcription factors has been appreciated for some time, the mechanistic consequences are less well understood. The hematopoietic transcription factor GATA1 is acetylated at conserved lysines that are required for its stable association with chromatin. We show that the BET family protein Brd3 binds via its first bromodomain (BD1) to GATA1 in an acetylation-dependent manner in vitro and in vivo. Mutation of a single residue in BD1 that is involved in acetyl-lysine binding abrogated recruitment of Brd3 by GATA1, demonstrating that acetylation of GATA1 is essential for Brd3 association with chromatin. Notably, Brd3 is recruited by GATA1 to both active and repressed target genes in a fashion seemingly independent of histone acetylation. Anti-Brd3 ChIP followed by massively parallel sequencing in GATA1-deficient erythroid precursor cells and those that are GATA1 replete revealed that GATA1 is a major determinant of Brd3 recruitment to genomic targets within chromatin. A pharmacologic compound that occupies the acetyl-lysine binding pockets of Brd3 bromodomains disrupts the Brd3-GATA1 interaction, diminishes the chromatin occupancy of both proteins, and inhibits erythroid maturation. Together these findings provide a mechanism for GATA1 acetylation and suggest that Brd3 "reads" acetyl marks on nuclear factors to promote their stable association with chromatin.hematopoiesis | posttranslational modifications | gene regulation P osttranslational acetylation of lysine residues is a widely used cellular signaling mechanism involving histone and nonhistone proteins. Typically, acetylated lysines (acK) serve as docking sites for bromodomains that are found in numerous chromatinrelated proteins (1, 2). Thus, bromodomains serve as "readers" of acK marks in a cascade of signaling events that influence diverse processes within chromatin. The hematopoietic zinc finger protein GATA1 was one of the earliest reported acetylated transcription factors (3, 4). GATA1 is a pivotal regulator of the erythroid, megakaryocyte, and mast cell lineages (5-9). GATA1 activates all known erythroid-specific genes, and mice lacking GATA1 succumb to anemia due to failed maturation and apoptosis of erythroid precursor cells (5, 10, 11). GATA1 also functions as a repressor, down-regulating genes associated with the proliferative, immature state. Mutations in the zinc finger region of GATA1 underlie congenital anemias and thrombocytopenias (12), whereas alterations at the N terminus of GATA1 are associated with megakaryoblastic leukemias in patients with Down syndrome (13). Previously, we found that GATA1 interacts with the acetyltransferase CREB binding protein (CBP) (14) to stimulate histone acetylation at GATA1 target genes (15, 16). CBP and its paralog p300 also acetylate GATA1 at two lysine-rich clusters C-terminal to each of its zinc fingers (Fig. 1A) (3, 4). CBPmediated acetylation of GATA1 c...

Recent evidence suggests that certain LEF/TCF family members act as repressors in the absence of Wnt signaling. We show here that repression by LEF1 requires histone deacetylase (HDAC) activity. Further, LEF1 associates in vivo with HDAC1, and transcription of a model LEF1-dependent target gene is modulated by the ratio of HDAC1 to ␤-catenin, implying that repression by LEF1 is mediated by promoter-targeted HDAC. Consistent with this hypothesis, under repression conditions the promoter region of a LEF1 target gene is hypoacetylated. By contrast, when the reporter is activated, its promoter becomes hyperacetylated. Coexpression of ␤-catenin with LEF1 and HDAC1 results in the formation of a ␤-catenin/HDAC1 complex. Surprisingly, the enzymatic activity of HDAC1 associated with ␤-catenin is attenuated. Together, these findings imply that activation of LEF1-dependent genes by ␤-catenin involves a two-step mechanism. First, HDAC1 is dissociated from LEF1 and its enzymatic activity is attenuated. This first step yields a promoter that is inactive but poised for activation. Second, once HDAC1-dependent repression has been overridden, ␤-catenin binds LEF1 and the ␤-catenin-LEF1 complex is competent to activate the expression of downstream target genes.The LEF1 transcription factor and its homologs (TCF1, TCF3, TCF4, dTCF, and pop1) transduce Wnt signals during development and the genesis of colon cancer (8,14,18,28,29,38,47). Wnt-stimulated transcriptional activation by the LEF/ TCF family is mediated by a bipartite transcriptional activator composed of a LEF/TCF family member and ␤-catenin. The rate-limiting step in the formation of this dimeric transcription factor appears to be the nuclear accumulation of ␤-catenin. In the absence of Wnt signal, ␤-catenin is localized to the cytoplasm, where it is phosphorylated by glycogen synthase kinase 3␤ (GSK3␤) and rapidly degraded. Phosphorylation of ␤-catenin by GSK3␤ is thought to occur within a multiprotein complex containing the adenomatous polyposis coli tumor suppressor protein and axin. Wnt signaling regulates ␤-catenin turnover by inactivating cytoplasmic GSK3␤, resulting in the stabilization of ␤-catenin. Stabilized ␤-catenin accumulates and translocates to the nucleus, where it interacts with an N-terminal region of members of the LEF/TCF family.LEF/TCF proteins were originally identified as transcriptional activators. However, a growing body of evidence indicates that LEF/TCF proteins also function as transcriptional repressors in the absence of Wnt signals (4). For example, in the early Xenopus laevis embryo, XTCF3 represses transcription of the Wnt-responsive homeobox gene siamois when Wnt signals are not present and activates siamois expression in cells receiving Wnt signals (7). Genetic studies of the Drosophila melanogaster LEF/TCF homologue dTCF (pangolin) and the Caenorhabditis elegans LEF/TCF homolog pop1 suggest that these transcription factors also repress the transcription of downstream target genes in the absence of Wnt signals. Thus, this feature of LEF/...

Background and Aims We evaluated the safety and efficacy of cilofexor (formerly GS‐9674), a small‐molecule nonsteroidal agonist of farnesoid X receptor, in patients with nonalcoholic steatohepatitis (NASH). Approach and Results In this double‐blind, placebo‐controlled, phase 2 trial, 140 patients with noncirrhotic NASH, diagnosed by magnetic resonance imaging–proton density fat fraction (MRI‐PDFF) ≥8% and liver stiffness ≥2.5 kPa by magnetic resonance elastography (MRE) or historical liver biopsy, were randomized to receive cilofexor 100 mg (n = 56), 30 mg (n = 56), or placebo (n = 28) orally once daily for 24 weeks. MRI‐PDFF, liver stiffness by MRE and transient elastography, and serum markers of fibrosis were measured at baseline and week 24. At baseline, median MRI‐PDFF was 16.3% and MRE‐stiffness was 3.27 kPa. At week 24, patients receiving cilofexor 100 mg had a median relative decrease in MRI‐PDFF of −22.7%, compared with an increase of 1.9% in those receiving placebo (P = 0.003); the 30‐mg group had a relative decrease of −1.8% (P = 0.17 vs. placebo). Declines in MRI‐PDFF of ≥30% were experienced by 39% of patients receiving cilofexor 100 mg (P = 0.011 vs. placebo), 14% of those receiving cilofexor 30 mg (P = 0.87 vs. placebo), and 13% of those receiving placebo. Serum gamma‐glutamyltransferase, C4, and primary bile acids decreased significantly at week 24 in both cilofexor treatment groups, whereas significant changes in Enhanced Liver Fibrosis scores and liver stiffness were not observed. Cilofexor was generally well‐tolerated. Moderate to severe pruritus was more common in patients receiving cilofexor 100 mg (14%) than in those receiving cilofexor 30 mg (4%) and placebo (4%). Conclusions Cilofexor for 24 weeks was well‐tolerated and provided significant reductions in hepatic steatosis, liver biochemistry, and serum bile acids in patients with NASH. ClinicalTrials.gov No. NCT02854605.

There is considerable debate whether peroxisome proliferatoractivated receptor B/D (PPARB/D) ligands potentiate or suppress colon carcinogenesis. Whereas administration of a PPARB ligand causes increased small intestinal tumorigenesis in Apc min/+ mice, PPARB-null (Pparb À/À ) mice exhibit increased colon polyp multiplicity in colon cancer bioassays, suggesting that ligand activation of this receptor will inhibit colon carcinogenesis. This hypothesis was examined by treating wild-type (Pparb +/+ ) and Pparb À/À with azoxymethane, coupled with a highly specific PPARB ligand, GW0742. Ligand activation of PPARB in Pparb +/+ mice caused an increase in the expression of mRNA encoding adipocyte differentiationrelated protein, fatty acid-binding protein, and cathepsin E. These findings are indicative of colonocyte differentiation, which was confirmed by immunohistochemical analysis. No PPARB-dependent differences in replicative DNA synthesis or expression of phosphatase and tensin homologue, phosphoinositide-dependent kinase, integrin-linked kinase, or phosphoAkt were detected in ligand-treated mouse colonic epithelial cells although increased apoptosis was found in GW0742-treated Pparb +/+ mice. Consistent with increased colonocyte differentiation and apoptosis, inhibition of colon polyp multiplicity was also found in ligand-treated Pparb +/+ mice, and all of these effects were not found in Pparb À/À mice. In contrast to previous reports suggesting that activation of PPARB potentiates intestinal tumorigenesis, here we show that ligand activation of PPARB attenuates chemically induced colon carcinogenesis and that PPARB-dependent induction of cathepsin E could explain the reported disparity in the literature about the effect of ligand activation of PPARB in the intestine. (Cancer Res 2006; 66(8): 4394-401)