Liver X receptors (LXRs) are nuclear hormone receptors that regulate cholesterol and fatty acid metabolism in liver tissue and in macrophages. Although LXR activation enhances lipogenesis, it is not well understood whether LXRs are involved in adipocyte differentiation. Here, we show that LXR activation stimulated the execution of adipogenesis, as determined by lipid droplet accumulation and adipocyte-specific gene expression in vivo and in vitro. In adipocytes, LXR activation with T0901317 primarily enhanced the expression of lipogenic genes such as the ADD1/SREBP1c and FAS genes and substantially increased the expression of the adipocyte-specific genes encoding PPAR␥ (peroxisome proliferator-activated receptor ␥) and aP2. Administration of the LXR agonist T0901317 to lean mice promoted the expression of most lipogenic and adipogenic genes in fat and liver tissues. It is of interest that the PPAR␥ gene is a novel target gene of LXR, since the PPAR␥ promoter contains the conserved binding site of LXR and was transactivated by the expression of LXR␣. Moreover, activated LXR␣ exhibited an increase of DNA binding to its target gene promoters, such as ADD1/SREBP1c and PPAR␥, which appeared to be closely associated with hyperacetylation of histone H3 in the promoter regions of those genes. Furthermore, the suppression of LXR␣ by small interfering RNA attenuated adipocyte differentiation. Taken together, these results suggest that LXR plays a role in the execution of adipocyte differentiation by regulation of lipogenesis and adipocyte-specific gene expression.Adipocyte differentiation, called adipogenesis, is a complex process accompanied by coordinated changes in morphology, hormone sensitivity, and gene expression. These changes are regulated by several transcription factors, including C/EBPs, peroxisome proliferator-activated receptor ␥ (PPAR␥), and ADD1/SREBP1c (44, 67). These transcription factors interact with each other to execute adipocyte differentiation, including lipogenesis and adipocyte-specific gene expression, which are pivotal for metabolism in adipocytes. Expression of C/EBP␤ and C/EBP␦ occurs at a very early stage of adipocyte differentiation, and overexpression of C/EBP␣ or C/EBP␤ promotes adipogenesis through cooperation with PPAR␥ (15, 32, 68, 69). PPAR␥, a member of the nuclear hormone receptor family, is predominantly expressed in brown and white adipose tissue (58, 59). PPAR␥ is activated by fatty acid-derived molecules such as prostaglandin J2 and synthetic thiazolidinediones (TZDs), novel drugs used in type II diabetes treatment (14,25,29). Recent studies involving PPAR␥ knockout mice indicated that the major roles of PPAR␥ are adipocyte differentiation and insulin sensitization (3,26,43). ADD1/SREBP1c, which also appears to be involved in adipocyte differentiation, is highly expressed in adipose tissue and liver and is also expressed early in adipocyte differentiation (22,60). ADD1/ SREBP1c stimulates the expression of several lipogenic genes, including FAS, LPL, ACC, SCD-1, and SCD-2 (22, 5...

A study of gene silencing within the mating-type region of fission yeast defines two distinct pathways responsible for the establishment of heterochromatin assembly. One is RNA interference-dependent and acts on centromere-homologous repeats (cenH). The other is a stochastic Swi6 (the fission yeast HP1 homolog)-dependent mechanism that is not fully understood. Here we find that activating transcription factor (Atf1) and Pcr1, the fission yeast bZIP transcription factors homologous to human ATF-2, are crucial for proper histone deacetylation of both H3 and H4. This deacetylation is a prerequisite for subsequent H3 lysine 9 methylation and Swi6-dependent heterochromatin assembly across the rest of the silent mating-type (mat) region lacking the RNA interference-dependent cenH repeat. Moreover, Atf1 and Pcr1 can form complexes with both a histone deacetylase, Clr6, and Swi6, and clr6 mutations affected the H3/H4 acetylation patterns, similar to the atf1 and pcr1 deletion mutant phenotypes at the endogenous mat loci and at the ctt1 ؉ promoter region surrounding ATF/CREbinding site. These data suggest that Atf1 and Pcr1 participate in an early step essential for heterochromatin assembly at the mat locus and silencing of transcriptional targets of Atf1. Furthermore, a phosphorylation event catalyzed by the conserved mitogen-activated protein kinase pathway is important for regulation of heterochromatin silencing by Atf1 and Pcr1. These findings suggest a role for the mitogen-activated protein kinase pathway and histone deacetylase in Swi6-based heterochromatin assembly.Methylation of histone H3 lysine 9 (H3 Lys-9) by the conserved H3 Lys-9-specific methyltransferase, Su(var)3-9 in flies, SUV39H1 in human, and Clr4 in the fission yeast Schizosaccharomyces pombe (1-4) correlates with heterochromatin assembly. The methylated Lys-9 residue recruits another conserved heterochromatin protein, which is called Swi6 in S. pombe and HP1 (heterochromatin protein 1) in higher eukaryotes (5, 6), leading to regional silencing of chromatin. In the fission yeast, recent studies (6 -8) addressing the silencing of the mating-type region provide insights for understanding the regulation of heterochromatin assembly in eukaryotes. Of particular interest, previous work (9) has defined sequential requirements for the establishment and maintenance of regional heterochromatic domains.Heterochromatin assembly at the mating-type region containing the mat2 and mat3 silent donor loci and an 11-kb interval (K region) between them requires several cis-acting DNA sequences as well as trans-acting factors (8, 10 -13). Heterochromatin formation at the centromeres and within the silent mat2/3 interval requires many of the same silencing factors, including Clr3 and Clr6 (H3/H4-specific histone deacetylases), the Clr4-Rik1 complexes, and Swi6 (2, 14 -19). The DNA elements involved in silencing within the entire 20 kb of the mat2/3 silent mating-type interval include REII (20), the mat3-M element including putative ATF 1 /CREB-binding sites (21), and the ...

Adiponectin is exclusively expressed in differentiated adipocytes and plays an important role in regulating energy homeostasis, including the glucose and lipid metabolism associated with increased insulin sensitivity. However, the control of adiponectin gene expression in adipocytes is poorly understood. We show here that levels of adiponectin mRNA and protein are reduced in the white adipose tissue of ob/ob and db/db mice and that there is a concomitant reduction of the adipocyte determination-and differentiation-dependent factor 1 (ADD1)/sterol regulatory element-binding protein 1c (SREBP1c) transcription factor. To determine whether ADD1/SREBP1c regulates adiponectin gene expression, we isolated and characterized the mouse adiponectin promoter. Analysis of the adiponectin promoter revealed putative binding sites for the adipogenic transcription factors ADD1/SREBP1c, peroxisomal proliferator-activated receptor ␥ and CCAAT enhancerbinding proteins. DNase I footprinting and chromatin immunoprecipitation analyses revealed that ADD1/ SREBP1c binds in vitro and in vivo to the proximal promoter containing sterol regulatory element (SRE) motifs. A luciferase reporter containing the promoter was transactivated by ADD1/SREBP1c, and introduction of SRE mutations into the construct abolished transactivation. Adenoviral overexpression of ADD1/SREBP1c also elevated adiponectin mRNA and protein levels in 3T3-L1 adipocytes. Furthermore, insulin stimulated adiponectin mRNA expression in adipocytes and augmented transactivation of the adiponectin promoter by ADD1/ SREBP1c. Taken together, these data indicate that ADD1/SREBP1c controls adiponectin gene expression in differentiated adipocytes.

Centromeres of fission yeast are arranged with a central core DNA sequence flanked by repeated sequences. The centromere-associated histone H3 variant Cnp1 (SpCENP-A) binds exclusively to central core DNA, while the heterochromatin proteins and cohesins bind the surrounding outer repeats. CHD (chromo-helicase/ATPase DNA binding) chromatin remodeling factors were recently shown to affect chromatin assembly in vitro. Here, we report that the CHD protein Hrp1 plays a key role at fission yeast centromeres. The hrp1Δ mutant disrupts silencing of the outer repeats and central core regions of the centromere and displays chromosome segregation defects characteristic for dysfunction of both regions. Importantly, Hrp1 is required to maintain high levels of Cnp1 and low levels of histone H3 and H4 acetylation at the central core region. Hrp1 interacts directly with the centromere in early S-phase when centromeres are replicated, suggesting that Hrp1 plays a direct role in chromatin assembly during DNA replication.

Adipocyte determination and differentiation dependent factor 1 (ADD1)/sterol regulatory element binding protein isoform (SREBP1c) is a key transcription factor in fatty acid metabolism and insulin- dependent gene expression. Although its transcriptional and post-translational regulation has been extensively studied, its regulation by interacting proteins is not well understood. To identify cellular proteins that associate with ADD1/SREBP1c, we employed the yeast two-hybrid system with an adipocyte cDNA library. Using the N-terminal domain of ADD1/SREBP1c as bait, we identified Twist2 (also known as Dermo-1), a basic helix-loop-helix (bHLH) protein, as a novel ADD1/SREBP1c interacting protein. Over-expression of Twist2 strongly repressed the transcriptional activity of ADD1/SREBP1c, primarily by reducing its binding to target sequences. Inhibition of histone deacetylase (HDAC) activity with HDAC inhibitors relieved this repression. Our data suggest that physical interaction between Twist2 and ADD1/SREBP1c attenuates transcriptional activation by ADD1/SREBP1c by inhibiting its binding to DNA, and that this inhibition is at least partly dependent on chromatin modification by HDACs.