The class II transactivator (CIITA), the master regulator of the tissue-specific and interferon gammainducible expression of major histocompatibility complex class II genes, synergizes with the histone acetylase coactivator CBP to activate gene transcription. Here we demonstrate that in addition to CBP, PCAF binds to CIITA both in vivo and in vitro and enhances CIITA-dependent transcriptional activation of class II promoters. Accordingly, E1A mutants defective for PCAF or CBP interaction show reduced ability in suppressing CIITA activity. Interestingly, CBP and PCAF acetylate CIITA at lysine residues within a nuclear localization signal. We show that CIITA is shuttling between the nucleus and cytoplasm. The shuttling behavior and activity of the protein are regulated by acetylation: overexpression of PCAF or inhibition of cellular deacetylases by trichostatin A increases the nuclear accumulation of CIITA in a manner determined by the presence of the acetylation target lysines. Furthermore, mutagenesis of the acetylated residues reduces the transactivation ability of CIITA. These results support a novel function for acetylation, i.e., to regulate gene expression by stimulating the nuclear accumulation of an activator.Major histocompatibility complex (MHC) class II genes encode heterodimeric cell surface molecules that are essential for the presentation of foreign antigenic peptides to helper T cells. Human and mouse genes are expressed in antigen-presenting cells as well as in various cell types upon gamma interferon (IFN-␥) stimulation (16, 34, 52). The expression of these genes occurs mainly at the transcriptional level and is regulated by an array of functional cis elements (H/W, X, and Y) that are conserved among all class II genes (16). Transcription of class II genes is orchestrated by the assembly of a higher-order multiprotein complex on the promoter and requires recruitment of the class II transactivator, CIITA (6, 34). Both constitutive and IFN-␥-inducible expression of class II genes are determined by the presence of CIITA in a variety of cell types (8,34,49). Functional analysis of the structure of CIITA revealed the presence of a C-terminal region required for promoter recruitment (44,59) and an N-terminal acidic transactivation domain that can contact the basic transcriptional machinery (13,35).Recently, we and others have demonstrated that the histone acetylase CREB binding protein (CBP) interacts with CIITA and functions as a coactivator for both B-cell-specific and IFN-␥-induced transcription of MHC class II genes (13, 30). Consequently, expression of MHC class II genes was suppressed by the adenovirus E1A protein (30), which is known to strongly bind to and inhibit CBP action (1, 33).The discovery that transcriptional coactivators have histone acetylase activity (4, 41) provided important insights into the process that links chromatin acetylation to transcriptional activation (20,31,50). CBP/p300 and the associated factor PCAF collaborate with many transcription factors as well as with other coa...

The class II transactivator (CIITA) is a key regulatory factor that controls expression of the major histocompatibility complex (MHC) class II genes that are essential components for antigen presentation and thus regulation of the immune response. We show here that the adenovirus E1A protein interferes with the action of CIITA and inhibits both B-cell-specific and gamma interferon (IFN-γ)-induced expression of MHC class II promoters. Transfection studies provide evidence for the functional role of the CREB-binding protein (CBP) in IFN-γ and CIITA-mediated MHC class II promoter activation. We demonstrate that the N-terminally located transcription activation domain of CIITA physically interacts with both the N-terminal and the E1A-binding (C/H3) regions of CBP. These results suggest the involvement of a multisubunit complex, which contains the gene-specific coactivator CIITA and the versatile coactivator CBP, in MHC class II gene regulation, which may be responsible for both high-level expression and modulation by different signaling pathways.

C.Spilianakis, A.Kretsovali and T.Agalioti contributed equally to this workWe describe the temporal order of recruitment of transcription factors, cofactors and basal transcriptional components and the consequent biochemical events that lead to activation of the major histocompatibility class II (MHCII) DRA gene transcription by IFN-g. We found that the gene is`poised' for activation since both the activators and a fraction of the basal transcriptional machinery are pre-assembled at the enhancer and promoter prior to IFN-g treatment. The class II transactivator is synthesized following IFN-g treatment and it is recruited to the enhanceosome leading to the subsequent recruitment of the CBP and GCN5 coactivators. This is followed by histone acetylation and recruitment of the SWI/SNF chromatin remodeling complex. CIITA also recruits the CDK7 and CDK9 kinases and enhances the ability of CDK7 to phosphorylate Pol II at Ser5 leading to initiation of mRNA synthesis. Thus, the gene-speci®c class II transactivator selects the target genes for expression by coordinating a multiple set of biochemical activities ranging from chromatin alterations and pre-initiation complex assembly to promoter clearance.

Histone deacetylase inhibition in stem cell differentiation

A gene profiling study of mouse embryonic stem cells treated with the histone deacetylase inhibitor trichostatin A shows that inhibition of histone deacetylases accelerates the early events of differentiation, by regulating the expression of pluripotency-and differentiation-associated in an opposite manner.

Histone deacetylase inhibitors (HDACi) are small molecules that have important and pleiotropic effects on cell homeostasis. Under distinct developmental conditions, they can promote either self-renewal or differentiation of embryonic stem cells. In addition, they can promote directed differentiation of embryonic and tissue-specific stem cells along the neuronal, cardiomyocytic, and hepatic lineages. They have been used to facilitate embryo development following somatic cell nuclear transfer and induced pluripotent stem cell derivation by ectopic expression of pluripotency factors. In the latter method, these molecules not only increase effectiveness, but can also render the induction independent of the oncogenes c-Myc and Klf4. Here we review the molecular pathways that are involved in the functions of HDAC inhibitors on stem cell differentiation and reprogramming of somatic cells into pluripotency. Deciphering the mechanisms of HDAC inhibitor actions is very important to enable their exploitation for efficient and simple tissue regeneration therapies.

HATs off! The development of the first cell‐permeable small‐molecule inhibitor of the human histone acetyltransferase (HAT) Gcn5 opens up new possibilities for understanding the histone code. Based on kinetic data and the proposed mechanism of the acetylation by Gcn5, the relatively simple butyrolactone structure of the inhibitor was identified.

Sall1 is a multi-zinc finger transcription factor that regulates kidney organogenesis. It is considered to be a transcriptional repressor, preferentially localized on heterochromatin. Mutations or deletions of the human SALL1 gene are associated with the Townes-Brocks syndrome. Despite its high expression, no function was yet assigned for Sall1 in embryonic stem (ES) cells. In the present study, we show that Sall1 is expressed in a differentiation-dependent manner and physically interacts with Nanog and Sox2, two components of the core pluripotency network. Genome-wide mapping of Sall1-binding loci has identified 591 genes, 80% of which are also targeted by Nanog. A large proportion of these genes are related to self-renewal and differentiation. Sall1 positively regulates and synergizes with Nanog for gene transcriptional regulation. In addition, our data show that Sall1 suppresses the ectodermal and mesodermal differentiation. Specifically, the induction of the gastrulation markers T brachyury, Goosecoid, and Dkk1 and the neuroectodermal markers Otx2 and Hand1 was inhibited by Sall1 overexpression during embryoid body differentiation. These data demonstrate a novel role for Sall1 as a member of the transcriptional network that regulates stem cell pluripotency.

Embryonic stem (ES) cells have high self-renewal capacity and the potential to differentiate into a large variety of cell types. To investigate gene networks operating in pluripotent ES cells and their derivatives, the “Functional Genomics in Embryonic Stem Cells” consortium (FunGenES) has analyzed the transcriptome of mouse ES cells in eleven diverse settings representing sixty-seven experimental conditions. To better illustrate gene expression profiles in mouse ES cells, we have organized the results in an interactive database with a number of features and tools. Specifically, we have generated clusters of transcripts that behave the same way under the entire spectrum of the sixty-seven experimental conditions; we have assembled genes in groups according to their time of expression during successive days of ES cell differentiation; we have included expression profiles of specific gene classes such as transcription regulatory factors and Expressed Sequence Tags; transcripts have been arranged in “Expression Waves” and juxtaposed to genes with opposite or complementary expression patterns; we have designed search engines to display the expression profile of any transcript during ES cell differentiation; gene expression data have been organized in animated graphs of KEGG signaling and metabolic pathways; and finally, we have incorporated advanced functional annotations for individual genes or gene clusters of interest and links to microarray and genomic resources. The FunGenES database provides a comprehensive resource for studies into the biology of ES cells.