~ Department of Genetics, University of Cambridge, Downing Street, Cambridge, CB2 3EH, U.K. email: bdmc@mole.bio.cam.ac.uk , § Department of Genetics, University of Glasgow, Church St., Glasgow, U.K., # Department of Molecular and Cell Physiology, Beckman Center, Stanford University, California., % Department of Zoology, Arizona State University, Tempe, Arizona,
The exocytosis of synaptic vesicles is a specialised and tightly regulated process essential to neuronal function. However many of the key proteins involved in this process have homologs which are expressed outside of the nervous system and which may participate in constitutive vesicle exocytosis in these tissues. Synaptobrevins have been demonstrated to play an essential role in vesicle trafficking at the synapse. Mammals have several synaptobrevin isoforms which are expressed predominately in the nervous system in addition to non-neuronally expressed forms such as Cellubrevin. Drosophila too has a neurally expressed synaptobrevin protein (N-SYB) and a ubiquitously expressed synaptobrevin (SYB). We have disrupted the function of N-SYB protein by transgenic expression of tetanus toxin light chain (TeTxLC) and demonstrated that this completely eliminates evoked synaptic transmission. SYB protein on the other hand is resistant to the toxin, and expression of TeTxLC outside the nervous system appears to have no adverse effects. Therefore, in order to examine the function of synaptobrevins outside of the nervous system we have recovered mutants in the syb gene. A number of transposable element and point mutations in the syb gene have been isolated. Preliminary characterisation of syb mutants demonstrates that they are lethal, homozygotes dying during the first larval instar. Loss of syb function appears to be attenuated by the large maternal contribution which is sufficient to allow completion of embryogenesis. However during late embryogenesis defects are observed in syb mutants. We will present data showing a range of these defects. In addition, vesicle trafficking is being examined in several tissues during the larval lethal stage. Of particular interest is the suggestion from cell culture studies that synaptobrevins may be involved in baso-lateral but not apical secretion in epithelial cells. This is being examined in syb mutants by utilising antibody markers specific for each pathway. As mentioned, disruption of N-SYB by TeTxLC eliminates all evoked synaptic transmission, however spontaneous or 'mini' vesicle fusion's continue at 50% of the wild type rate at NMJs where the toxin is expressed. One possible explanation for this is that SYB protein could be functioning in the exocytosis of these synaptic vesicles. The combination of syb mutations and transgenic tetanus toxin enables the dissection of the roles of SYB and N-SYB in 'mini' vesicle fusion at the synapse and is currently in progress. The inter-relationship between constitutive and neuronal secretion in Drosophila also offers the possibility to isolate genes involved in synaptic transmission by screening for mutants with defects in constitutive exocytosis. Identification of components common to both constitutive and regulated exocytosis will enable a comparison of vesicle trafficking between neurons and other secretory cells.