Small GTPases of the Rab family are involved in vesicular transport between different intracellular compartments. Two dominant alleles of
unc-108/rab-2 have been isolated in C. elegans exhibiting slow movement. The uncoordinated phenotype suggest defects in the synaptic transmission at the neuromuscular junction (NMJ). In C. elegans, fast synaptic transmission at the NMJ is mediated by acetylcholine released from synaptic vesicles (SV). Using pharmacological assays we showed that the movement defect observed in
rab-2 mutants is caused by decreased release of acetylcholine from pre-synaptic sites, while the postsynaptic side is not affected. A high pressure freeze electron microscopic study of synapses in
rab-2 mutants confirmed that the morphology as well as the number and distribution of synaptic vesicles was similar to wild type. The decreased release of acetylcholine in
rab-2 mutant animals could be rescued by the addition of phorbol esters. This suggests that the readily releasable SV pool in
rab-2 mutants is similar to wild type, however, the neuronal diacyl glycerol (DAG) levels are reduced leading to less SV exocytosis at steady state. It has been shown that SV exocytosis can be regulated by neuro-peptides, which are stored and secreted from dense core vesicles (DCV) in motor neurons at the NMJ. An electron microscopic (EM) analysis revealed that the number and distribution of DCVs in
rab-2 mutants is not affected. In contrast, by expressing fluorescently labeled neuro-peptides, which are packaged into DCVs, we show that DCVs in
rab-2 mutants contain and release less neuro-peptides. In addition, DCVs in
rab-2 mutants are bigger and more variable in size as judged by EM. This implicates RAB-2 in the biogenesis of DCVs. In agreement with that, Rab-2 is localized at Golgi but not at synapse in C. elegans neurons. Recently, the diabetes auto-antigen ICA69 has been reported to be an effector of Rab2. ICA69 has been shown to be involved in insulin secretion from secretory granules of beta cells in humans. Furthermore, ICA69 is also highly expressed in the brain, however, its function still remains unknown. We show that the C. elegans ICA69 ortholog, RIC-19, which is predominantly expressed in neurons is also involved in proper loading DCVs with neuro-peptides. RIC-19 also localizes to the Golgi and
ric-19 mutants exhibit DCV defects similar to
rab-2 mutants. Therefore, we propose that RIC-19 together with Rab-2 is involved in DCV biogenesis and loading at the Golgi in C. elegans motor-neurons.