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. 2011 Aug 15;6(8):e23598. doi: 10.1371/journal.pone.0023598

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Wanner et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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A, Schematic representation of SYG-2 transgene expression in the HSN under the unc-86 promoter. B, SYG-1::GFP expressed in the HSN localizes to the vulva area in wild type worms. Under the same promoter, SYG-2::GFP also localizes to the vulva area. Fluorescence and DIC image. C, syg-1(ky652) mutant animals display SNB-1::YFP vesicle clusters in the SSR. Ectopic expression of syg-1 in the HSN of syg-1(ky652) mutants rescues the mutant phenotype. syg-1(ky652) mutants with ectopic expression of syg-2 in the HSN, however, display synaptic vesicle clusters in the SSR. D, syg-1(ky652) mutants with ectopic expression of syg-2ex_syg-1in and syg-2ig1-5_syg-1in in the HSN also display synaptic vesicle clusters in the SSR. Fluorescence image and schematic drawing. Arrowhead, HSN cell body. Arrow, SNB-1 cluster. Asteriks, vulva. Scale bar represents 10 µm. Lateral view, anterior is to the left, ventral down. E, Quantification of SNB-1 punctae in the SSR. Ectopic expression of syg-2, syg-2ex_syg-1in, and syg-2ig1-5_syg-1in does not reduce the syg-1(ky652) mutant phenotype contrary to expression of syg-1. Mann-Whitney rank sum test. ***, p<0,001. n.s., not significant. n>30. Error bars, SEM. F, SYG-2 interacts with SYG-2 and chimerical SYG-2_SYG-1 proteins. V5.SYG-2 and Flag-tagged SYG-2, SYG-2ex_SYG-1in, SYG-2ig1-5_SYG-1in, or RGS3 were expressed in transiently transfected HEK 293T cells. After immunoprescipitation (IP) with anti-Flag antibody, the immobilized SYG-2 was detected with anti-V5 antibody in the precipitates containing SYG-2, SYG-2ex_SYG-1in, SYG-2ig1-5_SYG-1in, but not RGS3 (upper panel). The middle panel shows expression of V5.SYG-2 in the lysates, the lower panel expression of the Flag-tagged proteins. kD, kilodalton.