Apoptosis has been defined as a form of programmed cell death in all eukaryotic organisms and is related to a
variety of human diseases, such as viral infections, AIDS or cancer. In recent years, apoptotic cell death has also
been described for the yeast Saccharomyces cerevisiae. In this organism Cdc48p, a member of the AAA-family of
proteins (ATPases associated with a variety of cellular activities), plays an important role in the process of
apoptotic transformation. Cdc48p is stimulated upon hydrogen peroxide by transcription factor Yap1p, a protein
involved in the oxidative stress response. Since it has been shown that oxidative stress can lead to apoptosis it is
crucial to investigate the interplay of the two proteins, Yap1p and Cdc48p. The aim of my proposed project is to
elucidate the role of yeast transcription factor Yap1p in both regulation of Cdc48p and the apoptotic network. In
order to assess this question I will use strains carrying CDC48 and/or YAP1 mutant alleles. Mutations in CDC48
and YAP1 should provide more information about Yap1-mediated Cdc48p expression. In addition, mutant strains
will be examined for hallmarks of apoptosis including phosphatidylserine presentation on the cell surface,
chromatin condensation and DNA fragmentation and should help to gain further insight into the apoptotic
transformation. Combined biochemical and microscopic approaches including Northern blotting and DNA
microarray analyses, as well as fluorescence and electron microscopy will be utilized. Taking yeast as a genetically
tractable model organism, the proposed experiments might help to gain further understanding of oxidative stress
and apoptotic transformation of metazoan cells. In addition, information obtained from my work may have
practical implications for the diagnostic and treatment of disease states characterized by aberrantly regulated
apoptosis.