Synaptotagmin 1 and Synaptic Vesicle Protein 2-like in Arabidopsis

Abstract

The process of synaptic transmission at neuronal synapses is mediated by synaptic vesicle cycle. Synaptotagmins (SYTs) and Synaptic Vesicle Protein 2 (SV2), widely studied in animals since the 1990s, are important synaptic vesicle proteins that regulate synaptic neurotransmission in animal neurons. However, functions of these synaptic vesicle protein homologs in plants remain to be elucidated. In order to gain a better understanding of signal transmission in plants, this study focuses on the subcellular localizations of synaptotagmin 1 and SV2-like in Arabidopsis as well as the functions of these proteins.<br /> Arabidopsis synaptotagmin 1 (SYT1) is localized on the ER-PM contact sites in leaf and root cells. The ER-PM localization of Arabidopsis SYT1 resembles that of the extended synaptotagmins (E-SYTs) in animal cells. In mammals, E-SYTs have been shown to regulate calcium signaling, lipid transfer, and endocytosis. Arabidopsis SYT1 was reported to be essential for maintaining cell integrity by stabilizing the cytoskeleton. Our data provide detailed insight into the subcellular localization of SYT1 and VAP27-1, another ER-PM tethering protein. SYT1 and VAP27-1 were shown to be located on distinct ER-PM contact sites. The VAP27-1-enriched ER-PM contact sites (VECSs) were always associated with the SYT1-enriched ER-PM contact sites (SECSs), but not vice versa. The VAP27-1-enriched contact sites still existed in the leaf epidermal cells of SYT1 null mutant. However, the VAP27-1-enriched contact sites in SYT1 null mutant were less stable than that in the wild type. The polygonal networks of cortical ER disassembled and the mobility of VAP27-1 protein on the ER-PM contact sites increased in leaf cells of SYT1 null mutant. These results suggest that SYT1 is responsible for modulating the stability of ER network and the VAP27-1-enriched contact sites. Furthermore, cells of SYT1 mutant line have smaller BFA-induced compartments in the transition zone of root apices, indicating that the endocytic pathway is modulated by SYT1.<br /> Arabidopsis Synaptic Vesicle Protein 2-Like (SVL) belongs to the major facilitator superfamily and has been shown to be a niacin/trigonelline transporter. However, physiological studies of SVL in Arabidopsis still remain scarce. Our data have shown that Arabidopsis SVL protein is localized on the trans-Golgi network (TGN) and FM-dye stained early endosomes. The subcellular localization pattern of Arabidopsis SVL is similar to mammalian Synaptic Vesicle Protein 2 (SV2) and SV2-related Protein (SVOP) in neurons. The gene expression of SVL is developmental stage-dependent and can be detected in roots, hypocotyls, leaves, and anthers. One SVL mutant (svl-1) was identified and the transgenic Arabidopsis expressing SVL-GFP fusion protein was generated for functional and cytological studies. We have demonstrated that the primary root of SVL mutant grew slightly faster than the wild type during the early seedling stage in the control condition. The subcellular localization of SVL was sensitive to brefeldin A (BFA) treatment. In summary, Arabidopsis SVL participates in the endocytotic pathway in roots. Further studies on other possible functions of SVL in signal transduction and stress responses are still required. These data provide first insights into the sensory functions of SVL in Arabidopsis roots.