Protein-coding mutation inAdcy3increases adiposity and alters emotional behaviors sex-dependently in rats

1 Objective Adenylate cyclase 3 (Adcy3) has been linked to both obesity and major depressive disorder (MDD). Our lab identified a protein-coding variant in the 2 nd transmembrane (TM) helix of Adcy3 in rats, and similar obesity variants have been identified in humans. The current study investigates the role of a TM variant in adiposity and behavior. Methods We used CRISPR-SpCas9 to mutate the TM domain of Adcy3 in WKY rats (Adcy3 mut/mut ). We also created a heterozygous knockout rat in the same strain (Adcy3 +/- ). Wild-type (WT), Adcy3 +/- , and Adcy3 mut/mut rats were fed a high-fat diet for 12 weeks. We measured body weight, fat mass, glucose tolerance, food intake, metabolism, emotion-like behaviors, and memory. Results Adcy3 +/- and Adcy3 mut/mut rats weighed more than WT rats due to increased fat mass. There were key sex differences: adiposity was driven by increased food intake in males but by decreased energy expenditure in females. Adcy3 mut/mut males displayed increased passive coping and decreased memory while Adcy3 mut/mut females displayed increased anxiety-like behavior. Conclusions These studies show that the ADCY3 TM domain plays a role in protein function, that Adcy3 may contribute to the relationship between obesity and MDD, and that sex influences the relationships between Adcy3 , metabolism, and behavior. Study Importance Questions What is already known about this subject? Adcy3 has been linked to both obesity and depression in humans, with protein-coding variants in the transmembrane domain leading to increased obesity. Yet the underlying mechanisms are unknown, and it is unknown if the same Adcy3 variants can cause both obesity and depression. Rodent Adcy3 knockout models show increased adiposity and altered emotional behaviors, but no rodent models have assessed the role of protein-coding variants in Adcy3. Very few studies have assessed sex differences. What are the new findings in your manuscript? This is the first study to assess the role of a protein-coding variant in the transmembrane domain of Adcy3 , showing increases in both obesity and emotion-like behaviors in a rat model. We observed striking sex differences, where male and female rats had different factors driving their adiposity and different patterns of altered behavior. How might your results change the direction of research or the focus of clinical practice? Understanding the role of Adcy3 in obesity and MDD may lead to improved treatments for both diseases. Improving our understanding of the sex differences caused by the same Adcy3 mutation emphasizes the importance of studying both sexes and paves the way for personalized medicine approaches.