Role of Rab27 in synaptic transmission at the squid giant synapse

Abstract

Small GTPase Rab is a member of a large family of Ras-related proteins, highly conserved in eukaryotic cells, and thought to regulate specific type(s) and/or specific step(s) in intracellular membrane trafficking. Given our interest in synaptic transmission, we addressed the possibility that Rab27 (a close isoform of Rab3) could be involved in cytosolic synaptic vesicle mobilization. Indeed, preterminal injection of a specific antibody against squid Rab27 (anti-sqRab27 antibody) combined with confocal microscopy demonstrated that Rab27 is present on squid synaptic vesicles. Electrophysiological study of injected synapses showed that the anti-sqRab27 antibody inhibited synaptic release in a stimulation-dependent manner without affecting presynaptic action potentials or inward Ca 2؉ current. This result was confirmed in in vitro synaptosomes by using total internal reflection fluorescence microscopy. Thus, synaptosomal Ca 2؉ -stimulated release of FM1-43 dye was greatly impaired by intraterminal anti-sqRab27 antibody. Ultrastructural analysis of the injected giant preterminal further showed a reduced number of docked synaptic vesicles and an increase in nondocked vesicular profiles distant from the active zone. These results, taken together, indicate that Rab27 is primarily involved in the maturation of recycled vesicles and/or their transport to the presynaptic active zone in the squid giant synapse.Griscelli syndrome ͉ rabphilin ͉ small GTPase ͉ total internal reflection fluorescence microscopy ͉ vesicle exocytosis

Quantitative Analysis of Synaptic Vesicle Rabs Uncovers Distinct Yet Overlapping Roles for Rab3a and Rab27b in Ca2+-Triggered Exocytosis

“…These findings complement recent observations in invertebrate systems, where recycling defects were noted following genetic ablation or antibody-induced depletion of Rab27 proteins (Mahoney et al, 2006;Yu et al, 2008). Interestingly, we found that both GTP-and GDP-preferring Rab27b mutants profoundly impair stimulus-dependent recycling of SVs.…”

“…sion in invertebrates (Mahoney et al, 2006;Yu et al, 2008); but their precise subcellular localization has been unclear. In the following experiments, we have compared the subcellular localization of Rab27b with that of Rab3a using complementary approaches to determine whether they reside on identical or different synaptic vesicle pools.…”

“…Although it is still uncertain whether endosomes and their associated Rab repositories are involved as mandatory intermediates in every SV recycling event, the endosomal Rab5 GTPase has been shown to be involved in SV recycling in flies and mice (Shimizu et al, 2003;Wucherpfennig et al, 2003;Star et al, 2005). Furthermore, both Rab11b (Khvotchev et al, 2003) and Rab27 (Mahoney et al, 2006;Yu et al, 2008) have been recently suggested to play a role in the synapse, but their SV localizations and function(s) have yet to be assigned.…”

Distinct yet overlapping roles of Rab GTPases on synaptic vesicles

“…This synergy was not unexpected given the recent attention focused on Rab27 in SV docking and release in invertebrate systems. 28,29 Interestingly, we found that along with its inhibitory effect on uptake of a dye marker for endocytosis, dominantnegative Rab27b overexpression correlated with a depletion of endogenous Rab3a. As Rab3/Rab27b proteins share evolutionary conserved GTP-exchange factors i.e., Rab3GEF 28,30 together with the knowledge that GDP-mutants sequester cognate RabGEFs, 31 we reasoned that the GDPpreferring Rab27b mutants compete out Rab3a activation, thus accounting for the more pronounced SV recycling defect.…”

EPI64 Protein Functions as a Physiological GTPase-activating Protein for Rab27 Protein and Regulates Amylase Release in Rat Parotid Acinar Cells

“…Because treatment of parotid acinar cells with EPI64 antisense LNA significantly reduced the amount of amylase released, cytosolic EPI64 increase after IPR stimulation is also likely to regulate the biogenesis and/or recycling of secretory granules (e.g. transport of secretory granules to the apical region), in which Rab27 may be involved (42,43). A proper GTP-GDP cycle of Rab27 is presumably required for these processes.…”