Antigen presentation by class II major histocompatibility antigens (MHC) require not only the HLA-DR surface molecules, but also antigens processing molecules, DMA/DMB encoded within the HLA-D region, as well as the Ii chain protein encoded on chromosome 5. Class II mhc molecule. DMA/DBA and Ii are tightly linked in their function and most relevant to this proposal in their transcriptional control. In addition, the regulation of this group of molecules by IFN-gamma represents a novel pathway distinct from other IFN-gamma inducible systems. The purpose of this proposal is to continue the studies on the coordinate regulation of these genes, with specific emphasis on the IFN-gamma inducible regulation of this group of genes by the shared regulatory elements. W, X and Y, and by the newly described transactivator, Class II transactivator (CIITA). All class II MHC, Ii and DMA/DMB promoters contain shared regulatory elements which mediate majority of the transcriptional activities. Previous studies from our group have shown that IFN-gamma induced a gradual loading of transcription factors onto these elements. Using the in vivo footprint analysis, our laboratory has shown that binding of the NF-Y transcription factors at the Y (also a CCAAT) element is a critical event for subsequent binding at the X elements upon IFN-gamma induction.
The first Aim will determine if loading at the Y element is a key step in the IFN-gamma induction of the Ii promoter, which is much more complicated and contains two separate authentic Y elements. In addition to the importance of NF-Y in IFN-gamma induced promoter loading. Our group has also found that CIITA, a potent positive regulatory identified by complementation cloning of DR-negative cell line, may also be involved in in vivo promoter loading as revealed by studies of G3B, a mutant cell line selectively defective in the IFN-gamma induction of CIITA. A more indepth analysis of this role of CIITA in promoter loading is studied in Aim 2. Published results have also shown that CIITA not only regulates class II MHC genes, but also the Ii land DMB genes. Analysis of other IFN-gamma inducible genes that are regulated by CIITA will be studies in SAim 3. Finally, identification of the basic defect in G3A, which leads to the lack of CIITA induction by IFN-gamma will be identified by complementation cloning.
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