CARCINOGENS AND CHROMATIN STRUCTURE AND FUNCTION

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DESCRIPTION: This is a once-amended proposal to continue Dr. Smulson's studies into Three aims are proposed to pursue this objective. The first is structured to attempt to clarify the interaction of PADPRP with DNA breaks in chromatin. Initial studies will evaluate the cycling of PADPRP on and off ends of DNA during repair, using bacterially-expressed PADPRP mutant proteins. Also, using stably transfected synchronized 3T3-L1 cells, evidence will be sought to establish the direct role of PADPRP with DNA replication/repair by defining the physical association with DNA polymerase alpha, as well as a multi-enzyme DNA replicative complex. The cleavage of PADPRP during apoptosis in this system also will be evaluated. Aim II is designed to determine whether the poly(ADP-ribose) signal generated subsequent to DNA strand breaks plays a significant role in the accumulation of p53 protein and the arrest of cells at G1 until repair can proceed. In aim III the mechanistic role of PADPRP in V(D)J recombination, which shares several enzymatic features with strand break rejoining during repair, will be explored. The effects of the PADPRP expression constructs on this process will be examined in mouse 3T3 cells expressing the RAG 1 and RAG 2 genes and in the context of the whole animal by use of newly-established "knockout" mice.