The generation of high affinity antibodies is essential to protect us from infections with pathogenic organisms and damage from toxic substances. This affinity maturation of the antibody response requires the hypermutation of heavy and light chain variable (V) region genes. The detailed molecular and biochemical mechanisms responsible the regulation and targeting of this mutational process remain largely unknown, though transcription appears to be involved. We have shown that the mutation of V regions of stably transfected heavy chain genes in the NSO mouse plasma cell line shares many of the characteristics of V region mutation in vivo. This has allowed us to identify an AP1 like motif in the coding exons of gamma constant region genes that is required for V region hypermutation in NSO. Preliminary evidence suggests that this sequence recruits the GCN5 histone acetyltransferase to the heavy chain gene and that histone acetylation activates the mutational process in already highly transcribed genes. We now propose to confirm the role of histone acetylation in the regulation of V region hypermutation in human Burkitt lymphoma cell lines that originate from germinal center cells that carry out somatic mutation in vivo. We will use the chromatin histone immunoprecipitation (CHIP) assay to determine whether the histones associated with the mutating heavy chain genes are acetylated. We will use overexpression and down regulation of GCN5 expression to examine whether this particular histone acetylase is responsible for histone acetylation in vitro and in vivo in transgenic mice. We will also identify the DNA binding protein that recruits GCN5 to the mutating heavy chains and study its role and expression both in vitro and in vivo.
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