Differential palmitoylation regulates intracellular patterning of SNAP25
Differential palmitoylation regulates intracellular patterning of SNAP25
复制标题
DOI：
10.1242/jcs.079095
复制
发表时间：
2011-04-15
影响因子：
4
通讯作者：
Chamberlain, Luke H.
中科院分区：
文献类型：
--
作者：
Greaves, Jennifer;Chamberlain, Luke H.
SNAP 25调节质膜和内体系统中的膜融合事件，并且蛋白质的功能库通过ARF 6依赖性循环途径递送到再循环内体（RE）和反式高尔基体网络（TGN）。SNAP 25是一种外周膜蛋白，四个半胱氨酸残基的棕榈酰化介导其与膜的稳定结合。在这里，我们报告棕榈酰化也决定了SNAP 25的精确细胞内分布，并且突变单个棕榈酰化位点增强了RE和TGN处SNAP 25的量。将来自Hras的法尼基化的CAAX基序连接到紧邻富含半胱氨酸的结构域远端截短的SNAP 25突变体上。该构建体显示出与全长SNAP 25相同的细胞内分布，并且减少该构建体中半胱氨酸残基的数量增加了其与RE和TGN的结合，证实了富含半胱氨酸的结构域在指导SNAP 25的细胞内分布中的主导作用。观察到SNAP 25-CAAX和Hras构建体（各自具有两个棕榈酰化位点）的定位的显著差异，表明棕榈酰化序列的细微差异可对细胞内靶向产生重大影响。我们提出SNAP 25的富含半胱氨酸的结构域被设计为促进该SNARE蛋白在质膜和内体的双重功能，并且动态棕榈酰化作为调节SNAP 25的精确细胞内模式的机制。
SNAP25 regulates membrane fusion events at the plasma membrane and in the endosomal system, and a functional pool of the protein is delivered to recycling endosomes (REs) and the trans Golgi network (TGN) through an ARF6-dependent cycling pathway. SNAP25 is a peripheral membrane protein, and palmitoylation of a cluster of four cysteine residues mediates its stable association with the membrane. Here, we report that palmitoylation also determines the precise intracellular distribution of SNAP25, and that mutating single palmitoylation sites enhances the amount of SNAP25 at the RE and TGN. The farnesylated CAAX motif from Hras was ligated onto a SNAP25 mutant truncated immediately distal to the cysteine-rich domain. This construct displayed the same intracellular distribution as full-length SNAP25, and decreasing the number of cysteine residues in this construct increased its association with the RE and TGN, confirming the dominant role of the cysteine-rich domain in directing the intracellular distribution of SNAP25. Marked differences in the localisations of SNAP25-CAAX and Hras constructs, each with two palmitoylation sites, were observed, showing that subtle differences in palmitoylated sequences can have a major impact upon intracellular targeting. We propose that the cysteine-rich domain of SNAP25 is designed to facilitate the dual function of this SNARE protein at the plasma membrane and endosomes, and that dynamic palmitoylation acts as a mechanism to regulate the precise intracellular patterning of SNAP25.