The role for apoptosis in pathological processes, particularly in nervous tissues, has been recently established. Although little is known about the molecular mechanisms triggering this process in CNS, the mechanisms leading to execution of cell death are better understood. Caspase-3 activation has been proved to lead to the morphological and biochemical features of apoptosis. The investigator's recent studies established an important role of this caspase in neuronal apoptosis in vivo and in vitro and suggested that blockade of caspase-2 leads to reduction of cell death and associated neurological dysfunction. These results, confirmed now by other investigators, have shown that caspase-3 can be regulated in neuronal cells at the level of transcription. On the basis of these observations, they propose to elucidate mechanisms responsible for the regulation of caspase-3 gene activity. The general working hypothesis is that repression of caspase-3 transcription must have functional consequences for control of neuronal apoptosis. Accordingly, the new information proposed in this application may provide the basis for where the essential role for apoptosis has been established. In order to begin testing this general hypothesis, they will identify a gene control region for the rat caspase-3 gene and analyze general transcription factors required for its activity. The proposed studies are intended to address the hypothesis that the activity of caspase-3 is regulated, in part, at the level of transcription by gene regulatory factor(s). There are four closely related specific aims, all focused on defining the mechanisms of the caspase-3 gene regulation: (1) to determine the structure of the 5' flanking region of the rat caspase-3 gene; (2) to identify a transcription start site for this gene; (3) to identify the promoter and promoter regulatory elements; and (4) to examine the role of specific regulatory elements in the regulation of the caspase-3 gene promoter activity.
