The Role And The Mechanism Of Transcription Factor SP1 In The Formation Of Primordial Follicles In Mice

Ovarian follicles are the fundamental functional reproduction unit.The establishment of the primordial follicle(PF)pool is pivotal for the female reproductive lifespan.During folliculogenesis,oocytes and somatic cells undergo dynamic alterations in gene expression,which are regulated by a set of well-coordinated transcription factors.In the current study,we investigated the functional role of SP1 in PF formation in the perinatal mouse ovary.The results showed that SP1 was present in both oocytes and somatic cells in perinatal ovaries.To determine the precise effect of SP1 on the regulation of PF formation,fetal ovaries were injected with Sp1-shRNA lentivirus for knockdown of Spl.The PF formation was arrested and the number of total oocytes in Spl knockdown ovaries increased significantly compared to the control.Furthermore,an in vitro reconstitution result showed that SP1 likely functions in somatic cells rather than oocytes.To investigate the role of SP1in somatic cells,on the one hand,we used Lgr5-EGFP-ires-CreERT2(LGR5-KI)reporter mouse model to find that SP1 deletion significantly inhibited the recruitment of pregranulosa cells from LGR5 positive(LGR5+)cells in the ovarian surface epithelium;on the other hand,SP1 governed the maintenance of pregranulosa cells by using a FOXL2 positive(FOXL2+)cell-specific knockdown model.Consistent with Spl knockdown in the entire ovary,FOXL2+ cell-speicific knockdown of Sp1 disrupted PF formation and oocyte apoptosis.Further studies revealed that Notch2 mediated the function of SP1 in the development of pregranulosa cells and PF formation.ChIP and dual luciferase assay showed that SP1 bond directly to the promoter of the Notch2 gene.Finally,consistent with the critical role of granulosa cells in follicle survival in vitro,a large loss of oocytes in Spl knockdown ovaries was evidenced before puberty after the ovaries were transplanted under the renal capsule,which was similar to the phenotype of premature ovarian insufficiency(POI).A synchronized loss of FOXL2 and Notch2 expression in the area where Spl was silenced,demonstrating that SP1 plays a vital role in determining the survival of follicles by regulating granulosa cell development.Conclusively,our results reveal that SP1 controls establishment of the ovarian reserve by regulating pregranulosa cell development through Notch2 signaling.Our findings provide additional evidence elucidating the importance of ovarian somatic cell development during folliculogenesis,which contributes to a better understanding of the mechanism of folliculogenesis and follicle survival.