Regulation Of IPLA2 Activity And Localization By Rab5

Ca²⁺-independent phospholipase A₂?iPLA2?is a subclass of enzyme that catalyzes the release of fatty acids from phospholipids.Previous finding in our lab showed that iPLA2 activity is required for microglia chemotaxis and that inhibition of iPLA2 causes a trafficking defect of c-Src,remaining in the endosomal recycling compartment.This suggests the requirement of iPLA2 activity for vesicular recycling.Rab proteins,a group of small GTPases,regulate intracellular trafficking and maintain organellar identity by controlling budding,transport,tethering,docking,and fusion of recycling vesicles.Thus,one might imagine a possibility of the regulation of iPLA2 by Rab proteins.In this study,we investigated the interaction between iPLA2 and Rab proteins using GST pull down assays and found Rab5 is specifically binding to iPLA2.Using recombinant His-tag ankyrin repeats?His ANK?protein,we confirmed that Rab5 is binding to ankyrin repeats of iPLA2.We also analyzed the effect of ATP binding to the nucleotide-binding domain of iPLA2 on the interaction with Rab5 and demonstrated that ATP binding to iPLA2 inhibits the interaction with Rab5.Microscopic observations showed that ADP stimulation of microglia induces co-localization of iPLA2 and Rab5in vesicular compartments aound nucleus in starved microglia cells.Furthermore,expression of constitutively active Rab5(Rab5^CA)facilitates the association of iPLA2with vesicles.These results suggest that Rab5 interacts with iPLA2 and regulates the activity or localization for iPLA2.