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...

Memory of past cellular responses is an essential adaptation to repeating environmental stimuli. We addressed the question of whether gamma interferon (IFN-␥)-inducible transcription generates memory that sensitizes cells to a second stimulus. We have found that the major histocompatibility complex class II gene DRA is relocated to promyelocytic leukemia (PML) nuclear bodies upon induction with IFN-␥, and this topology is maintained long after transcription shut off. Concurrent interaction of PML protein with mixedlineage leukemia generates a prolonged permissive chromatin state on the DRA gene characterized by high promoter histone H3 K4 dimethylation that facilitates rapid expression upon restimulation. We propose that the primary signal-induced transcription generates spatial and epigenetic memory that is maintained through several cell generations and endows the cell with increased responsiveness to future activation signals.

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.

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.

We have isolated a series of human liver cDNA clones encoding glutamate dehydrogenase. The cDNAderived protein sequence specifies a single 558-amino acid long polypeptide including a cleavable signal sequence of 53 amino acids. Blotting analysis of RNA from human, monkey, and rabbit showed that glutamate dehydrogenase mRNA is present in various amounts in all tissues tested. Glutamate dehydrogenase mRNAs are of four sizes and are found in different ratios in different tissues; the predominant ones are ".3.5 and =2.9 kilobases. Blot hybridization of human genomic DNA to nonoverlapping cDNA fragments revealed multiple bands, many of which hybridize with two or more probes in a manner inconsistent with the existence of a single GLUD gene. Moreover, two separate 36-base synthetic oligonucleotides corresponding to the coding region hybridize to multiple genomic fragments, confirming the existence of more than one GLUDrelated gene in human.

Purified mouse fibroblast interferon (IF) directly rendered resting macrophages tumoricidal. The physicochemical properties and species specificity of the stimulatory agent fall within the present definition of IF. Since a number of polyanions induce macrophage IF, the antitumor and antimicrobial activities may result from the ability of newly released IF to modify macrophage activity.

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