UC Berkeley

Mutations affecting the leptin-melanocortin pathway and primary cilia are major causes of obesity, but the neuronal mechanisms remain unclear. In this dissertation, I developed an in vivo Cre-dependent CRISPRi and AAV-delivered sgRNA platform to examine how obesity-associated genes function in melanocortin 4 receptor-expressing neurons of the paraventricular hypothalamus (PVNMC4R neurons). This modular strategy enabled post-developmental, cell-type-specific gene downregulation with spatial and temporal control.In Chapter 2, I established and validated the CRISPRi system by downregulating Mc4r in adult PVNMC4R neurons and developed an in vitro sgRNA pre-screening pipeline to identify effective guides for in vivo use.In Chapter 3, I showed that downregulation of the cilia-specific adenylyl cyclase Adcy3 in adult PVNMC4R neurons is sufficient to drive obesity, demonstrating that ADCY3 is required within these neurons for normal body weight regulation.In Chapter 4, I performed an in vivo screen of obesity-associated primary cilia genes and found that Alms1, Arl13b, Bbs10, Ankrd26, Inpp5e, and Rab23 function within MC4R neurons to regulate body weight, whereas Tub does not. Further exploration of Alms1 revealed roles in both development and adult homeostatic regulation. Mechanistically, we found that Rab23, not Tub, is required for ciliary localization of endogenous MC4R.Together, these findings identify PVNMC4R neurons as a central convergence point for obesity-associated primary cilia genes and demonstrate that disruption of ciliary trafficking, structure, and signaling within this neuronal population is sufficient to impair long-term energy homeostasis.