Oxygenated Fatty Acids Enhance Hematopoiesis via the Receptor GPR132

Abstract

After their specification in early development, hematopoietic stem cells (HSCs) maintain the entire blood system throughout adulthood as well as upon transplantation. The processes of HSC specification, renewal, and homing to the niche are regulated by protein, as well as lipid signaling molecules. A screen for chemical enhancers of marrow transplant in the zebrafish identified the endogenous lipid signaling molecule 11,12-epoxyeicosatrienoic acid (11,12-EET). EET has vasodilatory properties, but had no previously described function on HSCs. EET treatment enhanced mouse marrow transplant, and time-lapse imaging showed that this lipid increased HSC specification in zebrafish embryos. These phenotypes were dependent on PI3Kγ signaling as well as AP-1 signaling, which in the zebrafish was specifically required in endothelial cells of the AGM or CHT. EET is known to signal via a G-protein coupled receptor (GPCR), but the identity of this receptor remains unknown, impeding the progress of EET to the clinic and preventing genetic studies of EET signaling. A novel bioinformatic approach identified 10 candidate EET receptors that are expressed in three EET-binding human cell lines, but missing from an EET-non-binding line. Of these, only GPR132 showed EET-responsiveness in vitro. Knockdown of zebrafish gpr132b prevented EET-induced hematopoiesis in the embryo, and marrow from GPR132 KO mice showed a decreased ability to transplant long-term. Others have shown that GPR132 has affinity for a variety of oxygenated fatty acids in vitro. Treatment of zebrafish embryos with these putative GPR132 ligands produced EET-like phenotypes in vivo. Structure-activity-relationship analyses using both in vitro and in vivo assays revealed that a carboxylic acid moiety is required for activity, and oxygenated, unsaturated fatty acids are stronger activators. I have identified GPR132 as an oxygenated fatty acid receptor that mediates both embryonic and adult hematopoiesis. This receptor is a promising target for therapeutic modulation of hematopoiesis and for genetic interrogation of fatty acid signaling. Together these studies reveal the biological and potentially therapeutic importance of lipid signaling molecules and their target GPCRs in regulating stem cell behavior.