RF21 | PSAT71 Unravelling the Role of CDK7-mediated SF1 Serine 203 Phosphorylation in Adrenal Homeostasis

Adrenal homeostasis relies on the centripetal differentiation and migration of peripheral adrenocortical progenitors to replenish the steroidogenic cells in the cortex. This homeostatic process is essential for steroidogenic functions and its dysregulation has been implicated in many adrenal diseases. Steroidogenic Factor 1(SF1) is a nuclear receptor which functions as a transcriptional factor to regulate global steroidogenic gene expression and adrenal development. However, the regulation of SF1 in the processes of differentiation (steroidogenesis) versus self-renewal of progenitors necessary for adult adrenal homeostasis remains largely unknown. Transcriptional activity of SF1 is regulated by post-translational modifications including phosphorylation (S203) and SUMOylation (K119 and K194). SUMOylation inhibits CDK7-mediated SF1 phosphorylation and SUMO-defective SF1 preferentially activates "SUMO-sensitive" target genes including Shh, rather than many well-characterized "SUMO-insensitive" target genes mediating differentiation. Based on these findings, together with data that CDK7 uniquely links cell cycle to transcriptional programs engaged in proliferating cells and the marked stem cell failure reported in multiple tissues of CDK7-KO mice, we hypothesized that CDK7-mediated SF1 phosphorylation in Shh-expressing adrenocortical progenitors regulates a unique transcriptional program to maintain their undifferentiated proliferative state during adrenal homeostasis. We engineered SF1-S203A/D mouse models which contain biallelic non-phosphorylatable/phosphomimetic germline mutations of SF1 at Serine 203. We crossed S203D mice with a) Shh-LacZ reporter strain to visualize Shh-expressing progenitors locked in an SF1 phosphomimic state and b) Shh-CreERT2;mTmG strain to GFP label Shh progenitors for lineage tracing. We observed increased beta-galactosidase staining in the zG of SF1-S203D reporter mice compared to controls, indicating an expanded Shh progenitor pool when SF1 is phosphorylated at S203. After 14 days of lineage tracing, GFP was co-stained with Cyp11B2, a marker for zG steroidogenic lineage, to quantify the percentage of Shh-expressing progenitors that differentiated into Cyp11B2-expressing zG cells. SF1-S203D mice exhibited a markedly lower differentiation rate of Shh-expressing progenitors compared to wild type, suggesting that SF1 phosphorylation preserved their undifferentiated state. Moreover, SF1-S203A mutation leads to a decreased adrenal/body weight ratio as well as downregulation of Shh. For insights into the role of CDK7 in SF1 phosphorylation during adrenocortical homeostasis, we generated conditional CDK7-KO mice. Adrenocortical-restricted CDK7-KO mice had decreased adrenal size, proliferation rate and Shh expression. Lineage tracing revealed increased differentiation of Shh-expressing progenitors when CDK7 was deleted specifically and temporally in Shh-expressing progenitors. Finally, we compared the differentiation rate in WT, CDK7-KO and CDK7-KO on an S203D background. The rapid differentiation induced by CDK7 loss was partially rescued by SF1-S203D, confirming the contribution of SF1 phosphorylation in mediating the functions of CDK7 in adrenocortical progenitors. Based on these findings, we propose a novel model in which CDK7-mediated SF1 phosphorylation regulates a unique transcriptional program that contributes to maintaining the undifferentiated proliferative state of adrenocortical progenitors. Presentation: Saturday, June 11, 2022 1:00 p.m. - 3:00 p.m., Sunday, June 12, 2022 12:30 p.m. - 12:35 p.m.