Endocrine Abstracts

JOINT2219

Makorin Ring Finger Protein 3 (MKRN3) loss-of-function mutations are the most common genetic cause of central precocious puberty. Its expression levels in the brain drop significantly towards puberty, suggesting effects in repressing the central control of reproduction and its role as a childhood “pubertal brake”. However, both the nature of this repression and the factors responsible for the drop in MKRN3 levels have remained unclear. To investigate these regulatory pathways, we performed bioinformatic analyses of several developmental RNA-seq datasets from rat and mouse brain, and from rat mediobasal hypothalamus (MBH). We identified genes whose expression levels correlate with those of Mkrn3 and change specifically across the pubertal transition, potentially encoding regulators of its expression, downstream effectors, or co-regulated genes. In order to detect its up-stream regulators, we filtered this group of Mkrn3-correlated genes for those linked to both puberty and signaling. Notably, these included Acvr1c, whose expression was strongly negatively correlated with Mkrn3 in all of the datasets and whose knockout was shown previously to delay puberty in mice. Analysis of single-cell RNA-seq data from mice hypothalamic neurons during development indicated that Mkrn3 and Acvr1c are not expressed simultaneously in the same cell. In experimental models, we demonstrated that activation of Acvr1c in GnRH neuronal cells suppresses Mkrn3 mRNA levels, and that this is mediated through Smad2/3 signaling. This repression involves recruitment also of the Kap1 transcription factor, and a marked shift from activating to repressive histone modifications at the gene locus. These findings provide a mechanistic explanation for the reduction in Mkrn3 arising from the developmental increase in expression of this receptor which can be activated by various available ligands. Our study highlights the value of integrating high-throughput gene expression data analysis with experimental studies to uncover novel regulatory networks, and have revealed here a new player in the regulation of pubertal onset.