Localization, dynamics, and protein interactions reveal distinct roles for ER and Golgi SNAREs.
Localization, dynamics, and protein interactions reveal distinct roles for ER and Golgi SNAREs.
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本地化，动力学和蛋白质相互作用揭示了ER和高尔基人的不同作用。
DOI：
10.1083/jcb.141.7.1489
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发表时间：
1998-06-29
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内质网到高尔基体的转运，可能还有高尔基体内的转运，涉及一组相互作用的可溶性N-乙基马来酰亚胺敏感因子附着蛋白受体（SNARE）蛋白，包括syntaxin 5、GOS-28、membrane、rsec 22 b和rbet 1。通过免疫电镜，我们发现rsec 22 b和rbet 1富集在COPII包被的囊泡中，这些囊泡从ER出芽，并可能与附近的囊泡管状簇（VTC）融合。然而，在COPII和COPI包被的膜上都发现了所有的SNARE，这表明类似的SNARE机制指导两种囊泡途径。rsec 22 b和rbet 1不会出现在第一个高尔基池之外，而syntaxin 5和membrin则深入渗透到高尔基堆栈中。温度变化表明，membrane，rsec 22 b，rbet 1，和syntaxin 5一起存在于膜上，快速循环之间的周边和高尔基体中心的位置。另一方面，GOS-28在高尔基体中保持固定的定位。通过免疫沉淀分析，syntaxin 5存在于至少两个主要的亚复合物中：一个含有syntaxin 5（34-kD同种型）和GOS-28，另一个含有syntaxin 5（41-和34-kD同种型），membrane，rsec 22 b和rbet 1。两个亚复合物似乎都涉及每个SNARE与突触融合蛋白5的直接相互作用。我们的研究结果表明，rbet 1，rsec 22 b，membrane，和syntaxin 5（34和41 kD）在两个膜融合界面之间的复合物的核心作用：ER衍生的囊泡与VTC的融合，以及VTC的组装形成顺式高尔基体元件。34 kD的syntaxin 5亚型、membrane和GOS-28可能在高尔基体内转运中起作用。
ER-to-Golgi transport, and perhaps intraGolgi transport involves a set of interacting soluble N-ethylmaleimide–sensitive factor attachment protein receptor (SNARE) proteins including syntaxin 5, GOS-28, membrin, rsec22b, and rbet1. By immunoelectron microscopy we find that rsec22b and rbet1 are enriched in COPII-coated vesicles that bud from the ER and presumably fuse with nearby vesicular tubular clusters (VTCs). However, all of the SNAREs were found on both COPII- and COPI-coated membranes, indicating that similar SNARE machinery directs both vesicle pathways. rsec22b and rbet1 do not appear beyond the first Golgi cisterna, whereas syntaxin 5 and membrin penetrate deeply into the Golgi stacks. Temperature shifts reveal that membrin, rsec22b, rbet1, and syntaxin 5 are present together on membranes that rapidly recycle between peripheral and Golgi-centric locations. GOS-28, on the other hand, maintains a fixed localization in the Golgi. By immunoprecipitation analysis, syntaxin 5 exists in at least two major subcomplexes: one containing syntaxin 5 (34-kD isoform) and GOS-28, and another containing syntaxin 5 (41- and 34-kD isoforms), membrin, rsec22b, and rbet1. Both subcomplexes appear to involve direct interactions of each SNARE with syntaxin 5. Our results indicate a central role for complexes among rbet1, rsec22b, membrin, and syntaxin 5 (34 and 41 kD) at two membrane fusion interfaces: the fusion of ER-derived vesicles with VTCs, and the assembly of VTCs to form cis-Golgi elements. The 34-kD syntaxin 5 isoform, membrin, and GOS-28 may function in intraGolgi transport.