The Neuronal Protein GAP-43 Associates with Microtubules and Lipid Rafts in Platelets and Megakaryocytes and Is Involved in Platelet Genesis

Abstract 2201

GAP-43 (neuromodulin) is a membrane-associated protein involved in neurite outgrowth, pathfinding and branching during development where it regulates neurotransmitter release and endocytosis, and plays a role in cytoskeletal signal transduction at the nerve ending. We have found GAP-43 to be expressed in platelets and megakaryocytes (MK) as evidenced by immunoblot and flow cytometry analysis. Immunofluorescence shows that GAP-43 localizes intracellularly as discrete punctuate structures mostly near the marginal microtubule (MT) coil of platelets and along the MT bundles in MKs and proplatelets. In contrast to neuronal cells where GAP-43 is associated with their detergent insoluble F-actin cytoskeleton, it is mostly soluble in platelets unless the platelets are first treated with taxol, a drug that stabilizes MTs. MT disruption with nocodazole or by chilling decreases the amount of GAP-43 in the detergent-insoluble cytoskeleton and disturbs its localization in proplatelets. GAP-43 colocalizes in platelets and MKs with the glycosylphosphatidylinositol-linked lipid raft marker Thy-1 and Cherry-PH PLC-delta1 (pleckstrin homology domain of phospholipase C-delta1) that stains membrane rafts. Since GAP-43 overexpression in MKs diminishes proplatelet formation, it appears to play a negative role in thrombopoiesis. Together, the data shows that GAP-43 is associated with MT and lipid rafts and suggests a role in signaling reactions between these two components to modulate proplatelet elaboration.