OR24-3 Persistence of Progenitor Cell Markers Following the Selective Ablation of Musashi in Somatotropes

Abstract

Metabolic and reproductive demands are met and coordinated through the complex control of hormone synthesis and secretion exerted by the anterior pituitary. While pituitary cells are known to possess remarkable plasticity to change their cell fate and alter hormone production in response to ever changing environmental cues, the underlying molecular control of this plasticity has not been established. Our lab has previously introduced the Musashi (MSI) family of RNA binding proteins as important players in control of pituitary hormone levels. Musashi typically governs stem cell fate and promotes self-renewal by repressing translation of target mRNAs needed for differentiation. However, we found that MSI1 is expressed in most differentiated cells of the adult pituitary and can specifically bind to the 3’ UTRs of Prl, Tsh, and Pou1f1 and exert translational control in reporter assays. Confirmation of the requirement for MSI was demonstrated through in vivo analyses where MSI1 and MSI2 were selectively ablated in somatotropes. The mutant animals were subfertile. The mutant males showed reduced serum and pituitary content of LH and FSH, and significant decreases in serum GH and PRL despite 2-fold increases in pituitary protein content of PRL and GH. To further assess the role and downstream pathways regulated by MSI in the pituitary, we collected somatotrope MSI-null pituitaries from males and females for qPCR analysis of common somatotrope target genes. In correlation with our previous findings described above (low serum GH and high pituitary GH content), we found that GHRHR mRNA levels were reduced by 2-fold in male mutants. RNA-seq analysis followed by qPCR validation shows that Prop1 mRNA was significantly increased in male mutants, with no significant change in Pou1f1 mRNA. These data suggest that MSI may be involved in the regulation of progenitor cells giving rise to the somatotrope lineage and that MSI ablation may have caused retention of these progenitors and/or a failure to fully differentiate somatotropes. Our studies of MSI-null somatotrope function support this interpretation because the mutant somatotropes clearly stored GH proteins, but could not secrete them, as demonstrated by the low serum GH values, and the 50% reduction in GHRHR mRNA. RNA-seq evaluation of males revealed a change in the expression of 720 genes between controls and mutants (FDR <0.05). When we examined female MSI-null somatotrope animals, a much smaller cohort of genes showed a change in expression (153 genes, FDR <0.05). Interestingly, 38 genes were altered in both mutant males and females suggesting shared regulation by MSI. Further characterization of the NGS datasets will elucidate additional downstream targets and effector pathways of MSI-dependent control of anterior pituitary cell differentiation and function.