Highly Conserved STAT3 Motifs Present in the Neuronal Pomc Enhancer nPE1 are Critical for Maximal Hypothalamic Pomc Expression.

Central melanocortins encoded by the Pomc gene control food intake and adiposity. Hypothalamic Pomc expression is driven by three 5'-distal enhancers known as nPE1, nPE2, and nPE3, with nPE1 carrying the strongest transcriptional capacity. Here, we investigated the functional relevance of two highly conserved canonical STAT3 motifs present in nPE1 regarding overall hypothalamic Pomc expression. We generated mutant mice carrying surgical co-allelic deletions of both STAT3 motifs using CRISPR-Cas9. Hypothalamic Pomc mRNA levels were measured by qRT-PCR in homozygous and wild-type adult littermates of both sexes, together with body weight, body length, fat mass, and food intake. A fast and refeeding test was performed with frequent measures of food intake and body weight. Using comparative genomics, we identified two highly conserved STAT3 motifs in nPE1. Homozygous female and male mice carrying simultaneous deletions of both motifs displayed 73.4 ± 0.8% and 76.5 ± 1.3% hypothalamic mRNA levels compared to same sex control littermates. Despite ~ 25% reduction in Pomc mRNA, body weight curves were normal across genotypes and sexes. A fast and refeeding experiment showed similar daily food intake and body weight loss and regain between homozygous and WT littermates. Our results demonstrate that the two STAT3 motifs present in the neuronal Pomc enhancer nPE1 are critical to reach full levels of hypothalamic Pomc expression.