Abstract
The activation of transcription factor Pou1f1 at embryonic day 13 gives rise to the pituitary populations of somatotropes, lactotropes, and thyrotropes and these populations maintain expression of Pou1f1 throughout life. The Musashi family of RNA regulatory proteins is known to regulate stem cell fate by repressing translation of target mRNAs needed for differentiation. Previously our lab has shown that female Lepr-null somatotropes have reduced POU1F1 protein levels but do not have changes in Pou1f1 mRNA expression. Stimulation with leptin increased the POU1F1 protein levels 3-fold, but did not change Pou1f1 mRNA suggesting a post-transcriptional mechanism for leptin’s regulation of Pou1f1. An in silico analysis indicated the presence a number of potential regulatory elements (MBEs) within the Pou1f1 mRNA 3’ UTR, including 8 consensus Musashi binding elements. Interestingly, we found musashi mRNA and protein levels were increased in Lepr-null somatotropes. This suggested that leptin regulates the expression of musashi in somatotrope populations and may be a candidate translational regulator of the Pou1f1 mRNA. We verified that MSI binds directly to the Pou1f1 mRNA 3’ UTR MBEs by EMSA in vitro and exerts translational repression (using reporter mRNA assays in transfected cell populations). Single cell RNA sequencing of pituitary cells from control male and female mice indicates that MSI and Pou1f1 mRNAs are co-expressed in somatotropes, lactotropes as well as thyrotropes. Immunocytochemical analyses confirmed that Musashi protein is present in mixed and purified somatotrope populations. Furthermore, immunoprecipitation with Musashi1 antibody showed a 5-fold enrichment of Pou1f1 mRNA in control female mouse pituitaries. These findings point to a critical in vivo role for Musashi-mediated mRNA translational regulation within the Pou1f1 lineage and specifically in the control of somatotrope maturation and response to metabolic cues.