Regulation of murine follicle-stimulating hormone ß subunit transcription by newly identified enhancers [scATAC-seq; Mouse Pituitaries]

Activin-class ligands of the TGFß family induce follicle-stimulating hormone (FSH) production by pituitary gonadotrope cells in mice via the actions of the transcription factors SMAD3, SMAD4, and FOXL2, which bind to cis-elements in the FSHß subunit (Fshb) promoter. An enhancer region for murine Fshb transcription was identified in vitro. However, deletion of the region using CRISPR-Cas9 did not affect FSH synthesis or secretion in mice. Using single-nucleus ATAC-seq of whole pituitaries, we identified three additional chromatin regions upstream of Fshb open exclusively in murine gonadotropes. We found these regions, as well as the Fshb gene, to be fully or partially closed in gonadotropes of FSH-deficient mice with genetically or pharmacologically inactivated activin type II receptors. The data indicate that there may be as many as four activin-sensitive enhancers upstream of murine Fshb. This data is part of a SuperSeries that includes bulk ATAC-seq and ChIP-seq data of gonadotrope-derived LßT2 cells, that substantiate the above conclusions. Overall design: Two sgRNAs were designed to target a previously identified enhancer region for murine Fshb transcription. Enhancer-knockout mice were generated using CRISPR-Cas9, after electroporation of the two sgRNAs and the Cas9 protein into zygotes. Pituitary glands were dissected from 8- to 10-week-old wild-type and enhancer-knockout mice, and prepared for scATAC-sequencing. Pituitary glands were similarly collected and processed from wild-type mice treated with either IgG or Bimagrumab, an activin type II receptor antagonist.