A novel Golgi-localisation domain shared by a class of coiled-coil peripheral membrane proteins
A novel Golgi-localisation domain shared by a class of coiled-coil peripheral membrane proteins
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DOI：
10.1016/s0960-9822(99)80168-7
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发表时间：
1999-04-08
期刊：
影响因子：
9.2
通讯作者：
Gleeson, PA
中科院分区：
文献类型：
--
作者：
Kjer-Nielsen, L;Teasdale, RD;Gleeson, PA
外周膜蛋白靶向高尔基体细胞质面的机制知之甚少。以前，我们已经确定了一个跨高尔基体网络（TGN）蛋白p230的羧基末端结构域，负责高尔基体定位[1]，在这里，我们报告了一个类似的高尔基体定位结构域的鉴定（GLD，也被称为“GRIP”结构域-参见Munro和Nichols在本期其他地方的论文）在一个来自低等和高等真核生物的推定外周膜蛋白家族中，大多数家族成员具有与p230相似的结构域结构，具有广泛的卷曲螺旋区域（>80%）和位于羧基末端的非卷曲螺旋结构域中的潜在GLD。以前报道的这个家族的蛋白质包括人golgin-97和酿酒酵母Imh 1 p。通过构建编码这些家族成员的羧基端区域与绿色荧光蛋白（GFP）融合的嵌合cDNA，我们直接证明了p230、golgin-97、新鉴定的人蛋白GCC 1 p和酵母Imh 1 p的GLD在转染的哺乳动物细胞中作为Golgi靶向结构域发挥作用。GLD的定点诱变鉴定了两个保守的芳香族残基，这对该靶向结构域的功能至关重要。内源性p230从高尔基体膜中被置换，在转染细胞中表达高水平的GFP融合到p230或golgin-97的GLD，表明不同的GLD与相似的膜决定簇相互作用。因此，我们已经确定了一个家庭的卷曲螺旋蛋白，共享一个域显示足以定位外周膜蛋白的高尔基体。
The mechanism by which peripheral membrane proteins are targeted to the cytoplasmic face of the Golgi apparatus is poorly understood. Previously, we have identified a carboxy-terminal domain of the trans-Golgi-network (TGN) protein p230 that is responsible for Golgi localisation [1], Here, we report the identification of a similar Golgi-localisation domain (GLD, also termed the 'GRIP' domain - see the paper by Munro and Nichols elsewhere in this issue) in a family of putative peripheral membrane proteins from lower and higher eucaryotes, The majority of family members have a domain structure similar to that of p230, with extensive coiled-coil regions (>80%) and the potential GLD located in a non-coiled-coil domain at the carboxyl terminus. Previously reported proteins in this family include human golgin-97 and Saccharomyces cerevisiae Imh1p. By constructing chimeric cDNAs encoding carboxy-terminal regions of these family members fused to green fluorescent protein (GFP), we have directly demonstrated that the GLD of p230, golgin-97, the newly identified human protein GCC1p and yeast Imh1p functions as a Golgi-targeting domain in transfected mammalian cells. Site-directed mutagenesis of the GLDs identified two conserved aromatic residues that are critical for the function of this targeting domain. Endogenous p230 was displaced from the Golgi membranes in transfected cells expressing high levels of GFP fused to the GLD of either p230 or golgin-97, indicating that different GLDs interact with similar membrane determinants. Thus, we have identified a family of coiled-coil proteins that share a domain shown to be sufficient for the localisation of peripheral membrane proteins to the Golgi apparatus.