Lipid rafts are cholesterol-enriched microdomains and implicated in many essential physiological ac-tivities such as the neurotransmitter release.Many studies have been carried out on the function of rafts inthe plasma membranes,whereas little is known about the information of such microdomains in subcellularcompartments especially synaptic vesicles(SVs).In the well-studied plasma membranes,several proteinshave been recognized as raft markers,which are used to label or trace rafts.But the raft marker proteinon SVs has not been identified yet.Although some SV proteins,including VAMP and CPE,have beenfound in raft fractions,they cannot be used as markers due to their low abundance in rafts.In this work,we designed several chimera proteins and tested their characteristics for using as SV raft makers.First,we detected whether they located in SVs,and then the chimeras exhibiting the better localization in SVswere further examined for their enrichment in raft using detergent treatment and gradient density floatationanalysis.Our results indicate that one of the chimeric proteins is primarily located in SVs and distributedin raft microdomains,which strongly suggests that it could be served as a raft marker for SVs.
In this paper we report that the C2 domain of synaptotagmin I (syt I) could associate with lipid rafts of plasma membrane. We demonstrate that phosphatidylinositol 4,5-bisphosphate (PIP2) in the target membrane and Ca2+ are the key factors to enhance the raft association of the C2 domain. We also found that the raft association of the C2 domain could be fulfilled by either C2A or C2B alone, suggesting that their raft association might be complementary. Finally, we indicate that destroying lipid rafts or blocking syt I-raft association could significantly reduce the Ca2+-driven release of glutamates. Our data indicate that the raft association of the C2 domain might play an important role in the regulated exocytosis.
