Research objectives are directed toward understanding the recognition and processing of radiation induced lesions with a particular emphasis on the formation of chromosomal aberrations. Nuclear DNA changes significantly impact human health with clear relationships to cell death, mutation and oncogenic change. There is uncertainly however as to the potentia contributions of stresses induced in non-nuclear targets to long term deleterious effects. It is clear that molecular interplay between membrane, cytoplasm and nucleus contributes to damage recognition, signaling and respons pathways. The p53 molecule is one pathway component implicated in a broad rang of cellular control responses and in a mutated or abrogated form is involved i many human cancers. An understanding of the role of sub-cellular components in radiation responses can efficiently be obtained by use of a microbeam. A directed beam of high LET charges particles can initiate significant levels of damage in sub-cellular microbolumes and single cell microscopic examination ca potentially localize gene product/s to such sub-cellular regions. In this proposal we will use cell site specific microbeam irradiation to ask the questions: Can non-nuclear irradiation initiate chromosomal clastogenic responses or cell cycle delay and p53 expression and localization. What is the kinetics of a p53 response? Can irradiated cells initiate responses in unirradiated neighbors? (uniquely achievable with a microbeam). Cells of human origin, normal and mutant (ATM), and cells expressing wild type and mutant p53 constructs will be used in these studies. Based on chromosomal aberration formation kinetics, it is hypothesized that p53 is not involved in initial DNA damage sensing and repair but is a surveillance component that helps to ensure genomic integrity. These microbeam based studies will help to provide a spatio-temporal framework for the recognition and resolution of radiation induced damage and the molecular components.
