A mammalian tripartite enhancer cluster controls hypothalamic <i>Pomc</i> expression, food intake, and body weight.

Abstract

Food intake and energy balance are tightly regulated by a group of hypothalamic arcuate neurons expressing the proopiomelanocortin (<i>POMC)</i> gene. In mammals, arcuate-specific <i>POMC</i> expression is driven by two <i>cis</i>-acting transcriptional enhancers known as nPE1 and nPE2. Because mutant mice lacking these two enhancers still showed hypothalamic <i>Pomc</i> mRNA, we searched for additional elements contributing to arcuate <i>Pomc</i> expression. By combining molecular evolution with reporter gene expression in transgenic zebrafish and mice, here, we identified a mammalian arcuate-specific <i>Pomc</i> enhancer that we named nPE3, carrying several binding sites also present in nPE1 and nPE2 for transcription factors known to activate neuronal <i>Pomc</i> expression, such as ISL1, NKX2.1, and ERα. We found that nPE3 originated in the lineage leading to placental mammals and remained under purifying selection in all mammalian orders, although it was lost in <i>Simiiformes</i> (monkeys, apes, and humans) following a unique segmental deletion event. Interestingly, ablation of nPE3 from the mouse genome led to a drastic reduction (>70%) in hypothalamic <i>Pomc</i> mRNA during development and only moderate (<33%) in adult mice. Comparison between double (nPE1 and nPE2) and triple (nPE1, nPE2, and nPE3) enhancer mutants revealed the relative contribution of nPE3 to hypothalamic <i>Pomc</i> expression and its importance in the control of food intake and adiposity in male and female mice. Altogether, these results demonstrate that nPE3 integrates a tripartite cluster of partially redundant enhancers that originated upon a triple convergent evolutionary process in mammals and that is critical for hypothalamic <i>Pomc</i> expression and body weight homeostasis.