Role of DAX1 in Testis Determination and Function

Over the last 8 years, we identified and characterized patients with naturally occurring mutations inthe orphan nuclear receptors, DAXl and SF1, and demonstrated that DAXl acts in part by inhibiting SF1-mediated transcription. Using targeted mutagenesis, we developed a murine Daxl knockout (KO) model andidentified roles for Daxl in gonadal determination, testis development, and adult testis function. In this grant,we propose to extend these studies, which have provided unexpected insight into mechanisms of gonadaldevelopment. We propose 3 inter-related aims that focus on Daxl structure and function as a transcriptionalrepresser, identify genetic pathways regulated by Daxl, and develop animal models to unravel the interplayof Daxl with other genes involved in testis development and function. Specifically, in Aim 1 we will identifymolecular partners that mediate DAXl transcriptional repression. Naturally occurring DAX1 mutationswill be identified and characterized to elucidate key structural domains required for DAX1 function in vivo.Candidate proteins identified in a yeast two-hybrid screen of an embryonic gonadal library will be furthercharacterized. The goal of Aim 2 is to identify the genetic pathways regulated by Daxl using microarrayanalyses of genes expressed in wild type versus Daxl-deficient embryonic gonads at 12 dpc, a timepointwhen Daxl expression diverges in males and females. Aim 3 is designed to explore the interaction ofDaxl with other genetic pathways in vivo. Using mice that lack Daxl, we will explore the gonadalphenotypes of mice with selective rescue of Daxl in various cell types. In addition, Daxl-deficient mice willbe crossed to other strains with alterations in genetic pathways proposed to intersect with Daxl (e.g., Sfl,Sry, Ptc). Success in these aims should provide a critical link between DAX1 and transcriptional repressionpathways that link a variety of other transcription factors involved in gonadal development.