Enzymatic activities that post-translationally modify the histones are central to the regulation of gene expression. The best studied histone modification at present is acetylation of lysine residues. Increased histone acetylation accompanies gene activation, whereas decreased acetylation is associated with gene repression. Acetylation levels are governed by opposing histone acetyltransferase (HAT) and histone deacetylase (HDAC) activities. Although a number of transcriptional coactivators and corepressors that house these activities have been described in the last few years, the role of these enzymes in transcriptional programs of cellular differentiation is currently understudied. Few mutations in mouse genes for these enzymes have been created or identified, although mutations in specific HATs and HDACs are associated with human diseases including colon cancer and leukemias. Gcn5 was the first nuclear HAT activity to be identified, and it serves as a useful paradigm for HAT structure and function. Previously, we demonstrated that loss of Gcn5 in mouse causes embryonic lethality just after gastrulation, with a loss of particular mesodermal lineages. These lineages are specified normally but failafter 7.5 days of gestation due to increased apoptosis. These experiments demonstrate the importance of Gcn5 to normal development, but the death of Gcn5 null embryos precludes analysis of Gcn5 functions at later embryonic time points or mechanistic studies to determine the molecular basis of the increased apoptosis. Experiments here will make use of newly developed genetic tools to address these questions.
Our specific aims are to 1) Determine the importance of Gcn5 at specific developmental stages using a conditional (floxed) allele 2) Determine the importance of Gcn5 HAT activity to development using alleles that carry point mutations in the catalytic center 3) Determine whether developmental defects and apoptosis are cell autonomous in mosaic animals 4) Determine the molecular effects of Gcn5 loss on cell growth, transcription, and DNA repair in Gcn5 null embryonic stem cells. These studies will provide important and novel information about the functions of this HAT during mammalian development and will provide a springboard for long term genetic studies to define the functions of this and other histone modifying activities in normal and diseased states.
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