3118 - GPRASP Family Members Function as Novel Negative Regulators of Murine Hematopoietic Stem Cell Transplantation via Controlling CXCR4 Degradation

Abstract

Hematopoietic stem cell transplantation (HSCT) is the only curative therapy for many patients with leukemia and hematologic disease. Sadly, immunological complications and infections contribute to high morbidity and mortality post-transplant. Recent work suggests that transplanted HSC are subject to distinct molecular regulation compared with HSC at steady-state. Thus, we seek to identify novel molecular regulators of HSCT. Here we report that multiple GPRASP (G-protein coupled Receptor (GPCR) associated Sorting Proteins) family members function as novel barriers to HSCT in mice. Treatment of hematopoietic stem-progenitor cells (HSPCs) with Gprasp1 or Gprasp2-shRNAs results in up to a 4-fold increase in hematopoietic reconstitution by 4 weeks post-transplant and maintained for more than 16 weeks post-transplant. GPRASPs regulate GPCR degradation by modulating endosomal/lysosomal sorting. The GPCR, CXCR4, a master regulator of HSC engraftment, cell cycle and survival, contains a C-terminal GPRASP-binding motif, implicating it as a target. Gprasp1- and Gprasp2-shRNAs increased survival and quiescence in HSPCs ex vivo and 20 days post-transplant. In contrast, Gprasp loss in Cxcr4-/- HSPCs had no effect on their survival or cell cycle status. Further, Gprasp1 and 2 losses increased CXCR4 levels in HSPCs and perturbed CXCR4 cellular localization. In pilot studies, GPRASP2 appears to physically interact with CXCR4. These data suggest that loss of GPRASP1 or GPRASP2 in HSPCs stabilizes CXCR4, resulting in increased engraftment, survival and quiescence acutely post-transplant. In sum, we have discovered that multiple members of the GPRASP family function as barriers to optimal HSCT by effecting HSC survival, cell cycle status and engraftment via regulation of CXCR4 stability. This protein family has never before been implicated in hematopoiesis or HSC biology. Further, our data illuminate a novel means to modulate CXCR4 levels and open up new avenues to developing therapies that improve HSCT.