Identification of FSH-regulated and estrous stage-specific transcriptional networks in mouse ovaries.

Follicle-stimulating hormone (FSH) acts by binding to FSHRs expressed on ovarian granulosa cells and produces estradiol. FSH is essential for female fertility because mice lacking FSH (Fshb KO) are anestrous and infertile. Although several in vitro cell culture and ex vivo approaches combined with pharmacological hormone treatment were used to identify FSH-regulated genes, how FSH orchestrates ovarian gene networks in vivo has not been investigated. Whether FSH-regulated genes display estrous stage-specific expression changes has also not been studied. Here, we functionally rescued Fshb null mice with a gonadotrope-targeted HFSHB transgene and performed RNA-Seq analysis on ovarian RNAs obtained from FSH-intact (WT), FSH-deficient (Fshb KO), and FSH-rescue (HFSHB+ rescue) mice. By comparing WT vs. Fshb KO and Fshb KO vs. HFSHB+ rescue ovarian gene expression datasets, we identified FSH-responsive genes in vivo. Cross interrogation of these datasets further allowed us to identify several transcription factors (TFs) and RNA-binding proteins specific to FSH-regulated genes. In an independent set of experiments, we performed RNA-Seq analysis on ovarian RNAs from mice in diestrous (DE), proestrous (PE), and estrous (E) and identified estrous stage-specific ovarian gene expression patterns. Interestingly, many of the FSH-regulated TFs themselves were estrous-stage specifically expressed. We found that ESR2 and GATA6, two known FSH-responsive TFs, and their target genes are reciprocally regulated with distinct patterns of expression in estrous stages. Together, our in vivo models and RNA-Seq analyses identify FSH-regulated ovarian genes in specific estrous stages that are under transcriptional and posttranscriptional control.