Many types of DNA damage in eukaryotic cells activate the chromosomal enzyme poly(adenosine diphosphate ribose) polymerase. The increase in the enzyme's activity can cause rapid depletion of cellular levels of its substrate NAD+ and abrupt increases in its product poly(ADP-ribose). The polymer is usually covalently linked to chromosomal proteins, however the function of this posttranslational modificaion has not been defined. We have proposed that the function of the poly(ADP-ribose) synthesized in response to DNA damage is to alter chromatin structure in such a way that regions of DNA damage become more readily accessible to the enzymes of DNA repair. The overall objectives of this proposal re to test this hypothesis for the role of poly(ADP-ribose) synthesis in DNA repair and to define the factors that regulate the activity of poly(ADP-ribose) polymerase and its response to DNA damage. Parallel experiments will be conducted in vitro in an SV 40 minichromosome model and in vivo in cells derived from normal human donors and patients with genetic disorders of DNA repair including xeroderma pigmentosu, Fanconi's anemia, Bloom's syndrome and ataxia telangiectasia. The SV 40 minichromosome, reconstituted with specifc proteins and enzymes, will provide a well defined in vitro model to study the effect of specific factors affecting the interaction of poly(ADP-ribose) synthesis with DNA damage and repair. The studies in cells derived from normal donors and from patients with disorders of DNA repair will provide systems to investigate different aspects of the regulation of poly(ADP-ribose) synthesis. These cells will also be used to further define the possibility that defects in the poly(ADP-ribose) synthesis system may be etiologically related to some of the abnormalities that affect patients with these diseases.
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