Regulated exocytosis of secretory vesicles is critically dependent on the assembly of SNARE complexes between the plasma membrane SNAREs, synatxin and SNAP-25 and vesicle-associated synaptobrevin, which render vesicles fusion competent. Tomosyn, a syntaxin-binding protein, forms a complex with syntaxin and SNAP-25 in competition with synaptobrevin predicted to limit fusogenic SNARE complex formation. Consistent with these data, the highly conserved C. elegans tomosyn homolog, TOM-1, has been shown to negatively regulate both synaptic and dense-core vesicle release. The regulation and trafficking mechanism(s) for TOM-1 remain to be elucidated. A yeast-2-hybrid screen has revealed a novel TOM-1 binding partner, VPS-39, which could potentially play a role in these unknown processes. VPS-39 is a promising candidate because it is highly conserved among organisms and has previously been implicated in vesicle docking/fusion in other model systems. Homozygous
vps-39 mutants from a balanced strain, grow to adulthood but lay dead embryos. A translational mCherry fusion construct capable of rescuing this embryonic lethality, indicates that VPS-39 is expressed in many tissues including body wall muscles, neurons, and coelomocytes. Consistent with this expression pattern,
vps-39 mutants are aldicarb-resistant and exhibit a reduced evoked postsynaptic response at the neuromuscular junction. These defects are not due to altered synaptogenesis as the neuronal architecture in
vps-39 mutants appears normal, based on pACR2::GFP expression and UNC-18 antibody staining. The presynaptic ultrastructure in
vps-39 mutants also resembles wild type levels for total number of synaptic vesicles and number of docked vesicles, although
vps-39 mutants exhibit a modest increase in cisternae. This phenotype may be indicative of a generalized endocytic defect, as the human homolog (VAMP6p) has been shown to regulate lysosomal clustering and fusion. NLP-21::Venus expression in coelomocytes also appears aberrant, providing further evidence of a possible vesicle recycling defect. Surprisingly, the expression of neuronal TOM-1 appears to be reduced in
vps-39 mutants, although
tom-1 mutants exhibit an opposing phenotype i.e. enhanced synaptic transmission due to an increase in morphologically docked vesicles. Together these data suggest that VPS-39 may have pleiotropic effects. Given the extent of the synaptic transmission defect, we are currently assessing the potential role of VPS-39 in other presynaptic and postsynaptic processes.