12554 Human Sex Determination: SRY Length Regulates Its Cellular Stability And Hence The Robustness Of Testis Differentiation

Abstract Disclosure: M.A. Weiss: None. The abundance of transcription factors (TFs) mediated by the rates of degradation are subjected to be a robust to an appropriate level. This regulation via the proteasome is largely controlled by the stability of individual proteins and then could determine the direction of a gene-regulatory network. Insight is obtained through studies of bistable genetic circuits mediated by initiating transcription factors. A model is provided by SRY, a Y-encoded TF that initiates testicular differentiation. Known functions in human SRY (204 residues) majorly cluster in its high mobility group (HMG) box whereas the functions of the N- and C-terminal non-box segments are not well characterized. Here, we have used cell-based and mouse transgenic studies to measure the transcriptional threshold of SRY regulating the balance between development and dysgenesis. Our findings demonstrate a threshold length in the C-terminal domain of human SRY that determines the protein’s proteosome-enforced half-life. In a pre-Sertoli cell model, truncation of SRY resulted in the reduction of intracellular concentration and twofold attenuation of the male-specific GRN. Expression of the 1-164 fragment of human SRY in CRISPR-Cas9-edited XX mice failed to drive male differentiation whereas the 1-200 of SRY initiated male GRN development. This study provides insight into the robustness of human SRY and illustrates a powerful strategy to link biochemical properties in cultured cells and in vivo developmental outcomes. Our study reveals a checkpoint in a key TF initiating a sex-specific GRN, functioning as an experimental “control knob” in development. Our approach probes molecular determinants of cell fate and so promises to extend structure-function studies of SRY to the flanking and relatively obscure non-box domains. This result implies the balance between robustness and evolvability in metazoan is a game of numbers of initial transcription factor in the networks. Presentation: 6/1/2024