YEAST SEC16 GENE ENCODES A MULTIDOMAIN VESICLE COAT PROTEIN THAT INTERACTS WITH SEC23P
YEAST SEC16 GENE ENCODES A MULTIDOMAIN VESICLE COAT PROTEIN THAT INTERACTS WITH SEC23P
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DOI：
10.1083/jcb.131.2.311
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发表时间：
1995-10-01
影响因子：
7.8
通讯作者：
KAISER, CA
中科院分区：
文献类型：
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作者：
ESPENSHADE, P;GIMENO, RE;KAISER, CA
酿酒酵母SEC 16基因中的温度敏感性突变阻断了ER转运囊泡的出芽。SEC 16是通过互补的sec 16 -1突变克隆和编码一个240 kD的蛋白质位于不溶性，颗粒成分的细胞裂解物。Sec 16 p通过高盐从该颗粒部分释放，但不通过非离子洗涤剂或尿素。通过免疫荧光显微镜，一些Sec 16 p定位于ER。膜相关的Sec 16 p被纳入到来自ER的运输囊泡中，这些囊泡在体外囊泡出芽反应中形成，Sec 16 p结合到Sec 23 p，COPII囊泡外壳蛋白，如通过双杂交相互作用测定和细胞提取物中的亲和力研究所示。这些发现表明Sec 16 p与Sec 23 p作为运输囊泡外壳结构的一部分相关联。SEC 16的遗传分析鉴定了三个功能上可区分的结构域，一个结构域由聚集在SEC 16中间的五个温度敏感突变定义。这些突变中的每一个都可以由单独表达的SEC 16的中心结构域补充。SEC 16 p的化学计量对于分泌功能是至关重要的，因为SEC 16 p的过表达引起致死性分泌缺陷。这种致死性功能映射到蛋白质的NH 2-末端，定义了第二个功能结构域。SEC 16 p的COOH-末端结构域的单独功能通过其结合SEC 23 p的能力显示。总之，这些结果表明，Sec 16 p参与多种蛋白质-蛋白质相互作用的ER膜上，并作为一个完整的囊泡的外套的一部分。
Temperature-sensitive mutations in the SEC16 gene of Saccharomyces cerevisiae block budding of transport vesicles from the ER. SEC16 was cloned by complementation of the sec16-1 mutation and encodes a 240-kD protein located in the insoluble, particulate component of cell lysates. Sec16p is released from this particulate fraction by high salt, but not by nonionic detergents or urea, Some Sec16p is localized to the ER by immunofluorescence microscopy. Membrane-associated Sec16p is incorporated into transport vesicles derived from the ER that are formed in an in vitro vesicle budding reaction, Sec16p binds to Sec23p, a COPII vesicle coat protein, as shown by the two-hybrid interaction assay and affinity studies in cell extracts. These findings indicate that Sec16p associates with Sec23p as part of the transport vesicle coat structure.Genetic analysis of SEC16 identifies three functionally distinguishable domains, One domain is defined by the five temperature-sensitive mutations clustered in the middle of SEC16. Each of these mutations can be complemented by the central domain of SEC16 expressed alone, The stoichiometry of Sec16p is critical for secretory function since overexpression of Sec16p causes a lethal secretion defect, This lethal function maps to the NH2-terminus of the protein, defining a second functional domain, A separate function for the COOH-terminal domain of Sec16p is shown by its ability to bind Sec23p. Together, these results suggest that Sec16p engages in multiple protein-protein interactions both on the ER membrane and as part of the coat of a completed vesicle.