Our laboratory is involved in studies of protein-DNA interactions that regulate gene expression, including general controlled changes in chromatin structure, and specific promoter interactions with transcription factors. As the model system for this study, we use Drosophila spermatogenesis. Transcriptional activation in primary spermatocytes involves a large number of genes and apparently includes both chromatin remodeling and deployment of tissue specific
transcription factors. The mechanisms of transcriptional activation warrant detailed analysis, as this will provide a tool for studies of male fertility and, ultimately, for manipulation of gene expression in spermatogenesis. The information available on the mechanisms of gene regulation in primary spermatocytes of mammals is limited. In contrast, a wealth of relevant background information is available on Drosophila,
thus making it an invaluable model. The remarkable similarity of spermatogenesis between Drosophila and mammals implies that the results will be applicable to the mammalian system.

Testes specific transcription factors: Our analysis of the promoter sequences revealed a conserved testes specific element (TSE) that is present in about 50% of testes specific genes. We also identified a testes specific DNA binding protein, TSE-BF, that is capable of interaction with the TSE sequence. We are now well positioned to dissect the TSE/TSE-BF system in order to understand the role of TSE-BF in spermatogenesis. Our first priority is cloning of the factor, and we are using three different approaches to achieve this goal, including protein purification, and whole-genome transcriptional analysis combined with bioinformatics. Once the cloning is complete, TSE-BF function will be studied both in vivo and in vitro using transgenes and gene knockouts, and in vitro transcription system. Further directions of the project include identification and study of homologous system in mammals, and
characterization of other testes specific transcription factors in Drosophila.

Chromatin remodeling: Our computational studies of gene expression identified a number of clusters of co-expressed genes in the Drosophila genome, including the clusters of testes specific genes. Analysis of chromatin structure revealed that a cluster of non-homologous testes-specific genes corresponds to a regulated chromatin domain. This
suggest the presence of testes specific chromatin regulatory system that is capable of chromatin "opening" within certain domains, resulting in transcriptional activation. Further research is aimed at elucidation of mechanisms responsible for regulated chromatin remodeling. In addition, higher order chromatin structures are studied in interphase nuclei by
fluorescence in situ hybridization, in order to investigate chromosomal folding/anchoring.