RT Journal Article SR Electronic T1 Identification of Genes Involved in Synaptogenesis Using a Fluorescent Active Zone Marker in Caenorhabditis elegans JF The Journal of Neuroscience JO J. Neurosci. FD Society for Neuroscience SP 3833 OP 3841 DO 10.1523/JNEUROSCI.4978-04.2005 VO 25 IS 15 A1 Edward Yeh A1 Taizo Kawano A1 Robby M. Weimer A1 Jean-Louis Bessereau A1 Mei Zhen YR 2005 UL http://www.jneurosci.org/content/25/15/3833.abstract AB Active zones are presynaptic regions where synaptic vesicles fuse with plasma membrane to release neurotransmitters. Active zones are highly organized structurally and are functionally conserved among different species. Synapse defective-2 (SYD-2) family proteins regulate active zone morphology in Caenorhabditis elegans and Drosophila. Here, we demonstrate by immunoelectron microscopy that at C. elegans synapses, SYD-2 localizes strictly at active zones and can be used as an active zone marker when fused to green fluorescent protein (GFP). By driving expression of SYD-2::GFP fusion protein in GABAergic neurons, we are able to visualize discrete fluorescent puncta corresponding to active zones in living C. elegans. During development, the number of GABAergic synapses made by specific motoneurons increases only slightly from larvae to adult stages. In contrast, the number of SYD-2::GFP puncta doubles, suggesting that individual synapses accommodate the increasing size of their synaptic targets mainly by incorporating more active zone materials. Furthermore, we used this marker to perform a genetic screen to identify genes involved in the development of active zones. We recovered 16 mutants with altered SYD-2::GFP expression, including alleles of five genes that have been implicated previously in synapse formation or nervous-system development. Mapping of 11 additional mutants suggests that they may represent novel genes involved in active zone formation. Figure 6. Quantification of SNB-1::GFP and SYD-2::GFP puncta during development. Using Punc-25-DsRed to label GABAergic neurons, SYD-2::GFP (hpIs3) puncta were counted in the dorsal cord region between the commissures of motoneurons VD10 and VD12. A-C, Examples of DsRed-labeled commissures meeting the dorsal cord (arrows). Scale bar, 10 μm. D, Schematic representation of this region (courtesy of E. Jorgensen). E, The results of SNB-1::GFP (juIs1) and SYD-2::GFP (hpIs3) puncta quantification within this region (as shown in A-D) are illustrated by dark filled bars. Puncta were also quantified by counting the number of puncta within a 50 μm distance at different stages during C. elegans development. These results are shown as shaded bars. Numbers for juIs1 puncta are shown in yellow, and numbers for hpIs3 puncta are shown in blue.