This project focuses on understanding how dietary fluxes from w-3 fatty acids activate the FFAR4 receptor, expressed in primary cilia, in both preadipocytes to trigger adipogenesis and in pancreatic b cells to enhance insulin secretion. The proposal extends from a recent discovery in the Jackson lab of a dramatic requirement for primary cilia in de novo adipogenesis and for the accumulation of fat mass in mice (Cell 2019). Using a knockout of TULP3, a critical regulator of ciliary GPCR trafficking, the group has established a requirement for ciliary GPCR signaling and screened 36 candidates to discover that the w-3 fatty acid (FA) receptor FFAR4/GPR120 is critical. Using FFAR4 and ciliary markers, the group finds that ciliated FFAR4+ preadipocytes comprise a perivascular stem cell population in all fat pads. Further, DHA-FFAR4 signaling synergizes with insulin signaling to direct adipogenic fate change and that FFAR4 agonists rapidly activate localized cAMP in the cilium. To better define the steps in adipogenesis downstream of FFAR4, two genome-wide CRISPR screens for genes important in adipogenesis were conducted, one using a traditional adipogenesis formulation and the other activated by FFAR4 agonists. These screens recapitulated most known requirements for adipogenesis and provided a wealth of new genes important for the adipogenic program. Among these there is a strong requirement for new factors controlling translation initiation and cap-independent translation. Finally, looking for the importance of FFAR4 in other metabolic systems, the group discovered that all pancreatic islet cell types are ciliated, but that both b and a cells express FFAR4 in cilia. Detailed pharmacology and genetics show that FFAR4 agonists can drive glucose stimulated insulin secretion (GSIS) in isolated b cells or islets at levels comparable to known secretagogues like GLP-1. FFAR4 agonists also stimulate glucagon secretion in a cells. Both effects require ciliary trafficking.
The first Aim will focus on validating and explaining new adipogenesis factors found in the screen. This will focus on identifying specific adenylate cyclases, phosphodiesterases, cAMP effectors, and translational control genes.
Aim 2 will focus on FFAR4 signaling in pancreatic b cells to understand the mechanisms of how FFAR4 and cAMP drive insulin secretion and how another w-3 fatty acid receptor, FFAR1, drives calcium influx to cooperate with FFAR4. Finally, in Aim 3, the focus is to use mouse models systematically knocking out both FFAR4 and FFAR1 to examine the cooperation of these pathways in obesity, insulin secretion and diabetes.
This aim will also examine human patient adipose tissue to determine if FFAR4+ ciliary in preadipocytes are lost in aging, diabetic or obese patients, also comparing sex differences. Overall, Aim 3 will test whether FFAR4 or ciliary loss is an important driver of diabetic disease progression, setting up critical tests of FFAR4 and other GPCRs for normalizing adipogenesis and improving insulin secretion in aging, diabetic patients.
The project focuses w-3 fatty acids activating FFAR4 and other GPCRs to trigger adipogenesis and insulin secretion in pancreatic b cells. Our functional studies, proteomics, and CRISPR screens comprise a multiomics approach to new human disease genes and targets for diabetes.
