Synaptic vesicles (SVs) are neuronal subcellular organelles used for the storage and release of neurotransmitters in communicating with their targets. Efficient SV trafficking from the soma to synapses is critical for neuronal communication. While mechanisms regulating SV trafficking have been studied extensively, our understanding of this biological process is still incomplete. Here we use PLMs (two C. elegans mechanosensory neurons) as a model system to study SV trafficking in vivo. Sam (Synaptic vesicle tag Abnormal in Mechanosensory neurons) mutants were isolated in a visual screen for mutants with SV accumulation defects in PLM chemical synapses. One mutant,
sam-4(
js415) shows defective SV trafficking in PLMs, suggested by retention of SV florescent markers in the proximal regions of neurites. However, other synaptic components appear to traffic normally including mitochondria and active zone proteins. PLMs in
sam-4 mutants are grossly normal in position, polarity, branching, and the morphology of the soma and synaptic varicosities. Molecular cloning showed that
sam-4 encodes a novel protein, which is conserved in both invertebrates and vertebrates, but whose function is unknown.
js415 introduces a termination codon in the middle of the
sam-4 gene coding region, and thus likely represents a null allele. We showed that SAM-4 protein is cytosolic and expressed broadly in neurons. Tissue specific rescue demonstrated that SAM-4 functions cell-autonomously to regulate SV trafficking in PLMs. In neurites, SAM-4-TagRFP puncta colocalize with, but are smaller than, SV puncta.
sam-4 (
js415) null mutants show very mild behavioral defects comparable to mild
unc-104 alleles we have also isolated, which become much severe in the strong
unc-104(
e1265) mutant background. Interestingly, we found that
sam-4(null) and
unc-104(weak) alleles interact synergistically to yield very severe behavioral and SV trafficking defects, suggesting that SAM-4 may play a role in regulating SV trafficking via interaction with motor complexes. We are currently testing how SAM-4 interacts with UNC-104 to regulate these processes.