Molecular mechanisms of sex determination and testis differentiation

Decreasing sperm counts in human populations over the last century is a trend of increasing concern. Several recent studies have shown that embryonic environment and exposures can lead to adult onset and trans-generational disease including reduced sperm count and motility in rodent models. To better understand how atypical sexual development can lead to reduced fertility, it is critical to determine the mechanisms of the SRY directed program of male sexual differentiation, its disruption, and the fetal basis of adult onset disease. In this thesis, the mechanisms involved in mammalian testis development are investigated. Transcriptional regulation during sexual differentiation was investigated using a genomic microarray approach to determine how transcriptional and morphological events are orchestrated. Cellular pathways and processes affected by genes regulated in testis differentiation included Wnt and Notch signaling, cellular differentiation, proliferation, focal contact, RNA localization, and development. Transcriptional alterations of testis differentiation by the fungicide vinclozolin were also investigated using a microarray approach to identify candidate mechanisms of action leading to adult onset and trans-generational transmission of disease. The genes and processes affected suggest apoptosis and vascular formation may be affected by vinclozolin treatment and transcriptional and epigenetic mechanisms may be important for mediation of affects seen with vinclozolin treatment. These studies have identified many new candidate genes with known functions that suggest roles which may help to fill in gaps in the understanding of testis development and its disruption leading to adult onset trans-generational disease. To connect candidate genes and processes in the sex determination cascade to one another, their regulation and function will also need to be studied. In this thesis, Nt3 transcriptional activation was investigated as a direct target of SRY action. Results show SRY and SOX9 have the ability to activate the Nt3 promoter in a site specific manner providing preliminary evidence that it may be one of the missing links directly downstream of Sry for induction of male sex determination. The sum of work in this thesis has contributed to the understanding of functional and disrupted sex determination and testis development. These studies have significantly contributed to information on gene regulation during sex determination and provide multiple new candidate genes.