The proposed research is designed to determine the role of chromatin function and structure in the radioresponse of mammalian cells. Regarding chromatin function, we hope to determine: 1) whether radiation induces any changes in the rate of addition or replacement (turnover) of chromsomal proteins, and 2) whether any radiation-induced changes in the turnover of chromosomal proteins play a role in radiation-induced division delay. Regarding chromatin structure, the objectives are: 1) to determine whether the in situ arrangement of chromosomal proteins on the DNA molecule or changes therein affect the induction of DNA damage and its repair, and 2) to determine what role thermally induced structural changes in the protein-DNA interaction in chromatin play in heat-induced cell killing and radiosensization. Chromatin isolated from cultured mammalian cells will be used to assay both radiation and thermal effects. The isolated chromatin will be fractionated by gel electrophoresis so that alterations of individual protein components may be examined. The particular class of DNA damage we will study 5,6-dihydroxydihydrothymine (T gamma)-type products. T gamma-type products are formed in DNA by OH radicals and are removed by an excision repair process. We will examine the formation and repair of T gamma-type products as a function of the cell's position in the cell cycle as a function of the DNA-protein organization of chromatin. Also flow cytometry will be used to monitor nuclear protein changes following gamma-irradiation.
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