Palmitoylation and membrane interactions of the neuroprotective chaperone cysteine-string protein.
Palmitoylation and membrane interactions of the neuroprotective chaperone cysteine-string protein.
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DOI：
10.1074/jbc.m802140200
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发表时间：
2008-09-05
期刊：
影响因子：
--
通讯作者：
Chamberlain LH
中科院分区：
文献类型：
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作者：
Greaves J;Salaun C;Fukata Y;Fukata M;Chamberlain LH
半胱氨酸串蛋白(CSP)是一种广泛棕榈酰化的DNAJ家族伴侣蛋白，具有重要的神经保护功能。棕榈酰化是CSP细胞内分选和功能所必需的，因此了解CSP的这一基本修饰是如何调节的是重要的。最近的工作在哺乳动物细胞中发现了23个假定的含有保守的DHHC结构域的棕榈酰转移酶，在这里，我们发现CSP的棕榈酰化可以通过高尔基体定位的棕榈酰转移酶DHHC3、DHHC7、DHHC15或DHHC17的共同表达而特异性地增强。事实上，这些DHHC蛋白促进了CSP的稳定膜附着，否则CSP是胞质的。未棕榈酰化的CSP突变体的膜亲和力与随后的棕榈酰化之间呈负相关：膜亲和力增加的突变体定位于内质网，并与高尔基体定位的DHHC蛋白物理分离。灯盏花素A(BFA)可以挽救ER定位突变体的棕榈酰化，它促进了ER和高尔基膜的混合。然而，有趣的是，棕榈酰化突变体在BFA清洗后仍留在内质网，而不会流向更远端的膜室。我们认为CSP具有较弱的膜亲和力，允许蛋白质通过膜‘采样’直接定位其合作伙伴高尔基体定位的DHHC蛋白。增强膜结合的突变阻止采样，并导致CSP在细胞膜上积累，如内质网。因此，CSP棕榈酰化与高尔基膜的偶联可能是后续分选的重要要求。
Cysteine-string protein (CSP) is an extensively palmitoylated DnaJ-family chaperone, which exerts an important neuroprotective function. Palmitoylation is required for the intracellular sorting and function of CSP, and thus it is important to understand how this essential modification of CSP is regulated. Recent work identified 23 putative palmitoyl transferases containing a conserved DHHC domain in mammalian cells, and here we show that palmitoylation of CSP is enhanced specifically by co-expression of the Golgi-localised palmitoyl transferases DHHC3, DHHC7, DHHC15 or DHHC17. Indeed, these DHHC proteins promote stable membrane attachment of CSP, which is otherwise cytosolic. An inverse correlation was identified between membrane affinity of unpalmitoylated CSP mutants and subsequent palmitoylation: mutants with an increased membrane affinity localise to the ER and are physically separated from the Golgi-localised DHHC proteins. Palmitoylation of an ER-localised mutant could be rescued by brefeldin A (BFA) treatment, which promotes the mixing of ER and Golgi membranes. Interestingly though, the palmitoylated mutant remained at the ER following BFA washout, and did not traffic to more distal membrane compartments. We propose that CSP has a weak membrane affinity that allows the protein to locate its partner Golgi-localised DHHC proteins directly by membrane ‘sampling’. Mutations that enhance membrane association prevent sampling and lead to accumulation of CSP on cellular membranes such as the ER. The coupling of CSP palmitoylation to Golgi membranes may thus be an important requirement for subsequent sorting.