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Control of the anterior pituitary cell lineage regulator POU1F1 by the stem cell determinant Musashi

Citation

MacNicol, Angus et al. (2021), Control of the anterior pituitary cell lineage regulator POU1F1 by the stem cell determinant Musashi, Dryad, Dataset, https://doi.org/10.5061/dryad.51c59zw5s

Abstract

The cytokine leptin regulates energy homeostasis through ubiquitously expressed leptin receptors.  Leptin has a number of major signaling targets in the brain, including cells of the anterior pituitary (AP).  We have previously reported that mice lacking leptin receptors in AP somatotropes display growth hormone (GH) deficiency, metabolic dysfunction and adult onset obesity.  Amongst other targets, leptin signaling promotes increased levels of the pituitary transcription factor POU1F1 (also referred to as Pit1) which, in turn, regulates the specification of somatotrope, lactotrope and thyrotrope cell lineages within the anterior pituitary.  Leptin’s mechanism of action on somatotropes is sex-dependent, with females demonstrating post-transcriptional control of Pou1f1 mRNA translation.  In this study, we have investigated the mechanism that regulates Pou1f1 mRNA translational control and report that the stem cell marker and mRNA translational control protein, Musashi1, exerts repression of the Pou1f1 mRNA. In FACS-purified female somatotropes, Msi1 mRNA levels and Musashi1 protein levels are increased in the mouse model that lacks leptin signaling (Gh-CRE Lepr-null), coincident with lack of POU1f1 protein, despite normal levels of Pou1f1 mRNA. Single-cell RNA sequencing of pituitary cells from control female animals indicates that both Msi1 and Pou1f1 mRNAs are expressed in Gh-expressing somatrotropes and immunocytochemistry analyses of these cells confirms that Musashi1 protein is present in the somatotrope population. We demonstrate that Musashi interacts directly with the Pou1f1 mRNA 3’ untranslated region and exerts translational repression of a Pou1f1 mRNA translation reporter in a leptin-sensitive manner.  Musashi immunoprecipitation from whole pituitary reveals co-associated Pou1f1 mRNA.  These findings suggest a mechanism in which leptin stimulation is required to reverse Musashi-mediated Pou1f1 mRNA translational control to coordinate AP somatotrope function with metabolic status.

Methods

Publically accessible databases queried for author specific genes/processes as noted in accompanying Table and Figure legends.  

Author generated single cell RNA sequencing data (see Gene Expression Omnibus (GEO) under accession number GSE153045) with differential gene expression analysis between clusters performed as described in accompanying Figure legend and primary manuscript

Individual luciferase reporter assay experiments performed as described in theprimary manuscript that were aggregated to generate teh composite graph Figure 6D in the primary manuscript

EMSA (RNA binding assays) essentially as described in the primary manuscript (see figure 6A and 6B) except full length forms of human Musashi1 and Musashi2 were employed

Funding

National Institutes of Health, Award: R01HD093461

National Institutes of Health, Award: R01DK113776-01

National Institutes of Health, Award: R01HD087057

National Institutes of Health, Award: NIGMS P20 GM103425

National Institutes of Health, Award: NIGMS P30GM11070