Abstract
Regulation of Neurotrophin 3 Expression by SRY During Male Sex Determination
Biology of reproduction, Vol.78(Suppl_1), pp.189-189
05/01/2008
Abstract
Neurotrophin 3 (Nt3) plays a significant role in male gonadal development. It is required for mesonephric cell migration into the testis for testis cord formation. The expression profiles for Nt3 are consistent with those expected for a gene directly regulated by SRY, and potential SRY binding HMG box elements were identified in the Nt3 promoter. Therefore, Nt3 regulation by SRY was investigated. Nt3 expression profiles in rat were verified spatially and temporally by immunohistochemical localization and Nt3 was found to be in the developing testis cords in Sertoli cells of rat gonads. Nt3 promoter constructs with luciferase reporters were made using mouse and rat Nt3 promoters and HMG boxes mutated by site specific mutagenesis. Promoter activity was monitored using transfection assays in primary P20 rat Sertoli cells and in E13 rat testis cell lines. SRY element site specific mutant promoters had reduced luciferase reporter activity compared to wild type promoters. Since SRY and SOX9 are important for sex determination and SOX9 binds to DNA sequences highly similar to those bound by SRY, the potential roles of both SRY and SOX9 in regulation of the Nt3 promoter were investigated. The potential for SRY and SOX9 to affect Nt3 promoter activity was examined in co-transfection assays using SRY and SOX9 expression vectors. Results indicate that both SRY and SOX9 can stimulate the Nt3 promoter in transfection assays. Further analysis is required to reveal the key players in vivo: Knock down of Sry and Sox9 expression is being used to determine if there is a change in Nt3 expression when Sry or Sox9 are silenced at the initiation of gonadal differentiation. Gel shifts and high sensitivity ChIP assays are being utilized to determine the relative binding affinities of SRY and SOX9 to HMG boxes in the Nt3 promoter and investigate binding to the Nt3 promoter in vivo. These studies have helped clarify the regulation of Nt3 in the gonad during sex determination and reveal the potential for Nt3 to be regulated directly by SRY.
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Details
- Title
- Regulation of Neurotrophin 3 Expression by SRY During Male Sex Determination
- Creators
- Tracy M. Clement - Washington State UniversityRamji Bhandari - Washington State UniversityMichael Skinner - Washington State University
- Publication Details
- Biology of reproduction, Vol.78(Suppl_1), pp.189-189
- Academic Unit
- Biological Sciences, School of
- Identifiers
- 99901080818301842
- Language
- English
- Resource Type
- Abstract