CIITA regulates transcription onset viaSer5-phosphorylation of RNA Pol II

Abstract

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.

The S Box of Major Histocompatibility Complex Class II Promoters Is a Key Determinant for Recruitment of the Transcriptional Co-activator CIITA

“…1A) (29). The assay relies on the fact that the known constituents of the MHCII enhanceosome (RFX, CREB, and NF-Y) bind together in a highly cooperative fashion, such that they assemble into a stable higher order complex on DRA promoter fragments (18,24,29,30). This nucleoprotein complex generates a composite interaction surface to which CIITA binds via multiple synergistic protein-protein contacts (29,(32)(33)(34).…”

“…RFX binds in a highly cooperative manner with two other factors, CREB (22) and NF-Y (23), to their respective X, X2, and Y box target sites (24 -28). This generates a higher order MHCII enhanceosome complex that serves as a landing pad for CIITA (29,30). CIITA is recruited to the enhanceosome via multiple protein-protein interactions with RFX, CREB, and NF-Y (29, 31-35).…”

A T Lymphocyte-Specific Transcription Complex Containing RUNX1 Activates MHC Class I Expression

“…CIITA interacts with the constitutively expressed DNA-binding transcription factors RFX5, NF-Y, and CREB to assemble a B lymphocyte-specific enhanceosome that regulates MHC class II gene expression (22)(23)(24)(25)(26)86). Spilianakis et al (23) have recently described the temporal recruitment of transcription factors, cofactors, and basal transcriptional components leading to the expression of the MHC class II DR␣ gene by IFN-␥. CIITA is also a potent activator of MHC class I genes, where it is also thought to assemble a B cell enhanceosome.…”

“…A variety of DNA-binding transcription factors have been identified that interact with these promoter proximal DNA sequence elements. For example, a B lymphocyte-specific enhanceosome consisting of the coactivator CIITA and DNA-bound transcription factors RFX, CREB/ATF, and NF-Y leads to high cell surface class I and II expression in B lymphocytes (22)(23)(24)(25). CIITA, a transcriptional coactivator that induces both MHC class I and class II genes does not bind DNA directly, but rather depends on factors, such as RFX and CREB/ ATF, that bind to the CRE element (16,20,21).…”

ZXDC, a novel zinc finger protein that binds CIITA and activates MHC gene transcription

“…Detection of coimmunoprecipitated DNA was by quantitative real-time PCR. Five microliter of eluted DNA and primers directed against the HLA-DRA promoter (Spilianakis et al, 2003) or the GAPDH promoter (provided in the EZ-ChIP kit) were combined with Reaction Mix (QuantiTect SYBR Green PCR kit; Qiagen Inc.). Data were normalized to the signal detected with non-specific antiserum.…”