08492 ATM (Ataxia-Telangiectasia Mutated) Has a Role in Mitochondrial Pathway of Apoptosis Andrew Behesnilian Mentor: Leman Yel

Ataxia-Telangiectasia (A-T) is a genetic disorder characterized by immunodeficiency, ataxia and telangiectasia. ATM (Ataxia-Telangiectasia Mutated) is the defective gene in A-T, which shows an altered programmed cell death (apoptosis). The function of ATM in apoptosis is not clarified. Since DNA damage activates the mitochondrial pathway of apoptosis, and since A-T cells have defects in DNA damage repair, we hypothesize that ATM has a role in the mitochondrial pathway of apoptosis. We examined cell viability and apoptosis induced by etoposide, an agent that causes DNA damage, in fibroblast cell lines from two patients with A-T and one control subject. Cells treated with etoposide 50uM were studied for cell viability using an MTT assay. At 24 hr of etoposide treatment, A-T cells showed higher cell viability (99% and 84%) compared to the control (81% and 79%), which was due to decreased apoptosis demonstrated by Hoechst staining. In A-T cells, mitochondrial membrane potential () assessed by JC-1 staining was less than in control cells. Cytochrome c and apoptosis-inducing factor (AIF) release was determined by immunohistochemistry using fluorescence imaging. At 24 hr of etoposide induction, 65% of A-T cells released cytochrome c from mitochondria to the cytosol compared to the 90% of the control cells. AIF was released in 85% of A-T cells and 95% of control cells. These results show that etoposide induces apoptosis through the mitochondrial pathway in A-T and suggest that ATM renders cells with apoptotic susceptibility through the mitochondrial pathway.