Identify transcriptional regulators of Mc4r in the hypothalamus [RNA-Seq]

Human MC4R mutations can increase or decrease obesity risk. Other than intracellular signaling, MC4R function is also influenced by its levels. However, the genetic programs that govern MC4R transcription in the brain remain largely unknown. Moreover, it is unclear whether human genetic variants exist that affect Mc4r expression and obesity risk. Here, we identify the homeodomain transcription factor Otp as one regulator of Mc4r expression in a specific subset of hypothalamic neurons. Selective loss of Otp in these neurons during development or adulthood results in reduced Mc4r expression and excessive body weight gain. Moreover, OTP interacts with upstream regulatory sequences of Mc4r to modulate its transcription. Overall design: To identify transcriptional regulators of Mc4r in the mouse hypothalamus, we crossed Mc4r-Cre mice with Ai14 (R26RLSL-tdTomato) mice to generate double transgenic mice (Mc4r-Cre; Ai14) in which Mc4r-Cre-expressing neurons were labeled by the expression of a Cre-dependent tdTomato reporter. We then purified these neurons by fluorescence-activated cell sorting (FACS) and profiled their transcriptome using bulk RNA sequencing. To elucidate the molecular basis of excessive weight gain, we isolated PVH neurons targeted with either AAV-GFP or AAV-Cre by FACS and compared their transcriptomes using RNAseq.