Distinct roles for the AAA ATPases NSF and p97 in the secretory pathway
Distinct roles for the AAA ATPases NSF and p97 in the secretory pathway
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DOI：
10.1091/mbc.e03-02-0097
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发表时间：
2004-02-01
影响因子：
3.3
通讯作者：
Hanson, PI
中科院分区：
文献类型：
--
作者：
Dalal, S;Rosser, MFN;Hanson, PI
NSF和p97是涉及膜运输和细胞器生物发生的相关AAA蛋白。p97还参与导致泛素依赖性蛋白水解的途径，包括ER相关降解（ERAD）。在这项研究中，我们使用了显性干扰ATP水解缺陷突变体（NSF（E329 Q）和p97（E578 Q）），比较这些AAA蛋白在哺乳动物细胞分泌途径中的功能。表达NSF（E329 Q）促进高尔基体堆叠分解成分散的囊泡结构。它还迅速抑制糖胺聚糖硫酸化，反映了高尔基体内转运的破坏。相反，表达p97（E578 Q）不影响高尔基体的结构或功能;糖胺聚糖通常是硫酸化和分泌的，VSV-G ts 045蛋白也是如此。相反，p97（E578 Q）的表达导致泛素化蛋白在ER膜上积累，并减缓ERAD底物囊性纤维化跨膜电导调节剂的降解。此外，p97（E578 Q）的表达最终导致ER肿胀。p97（E578 Q）对细胞器组装的影响的更具体的评估表明，高尔基体在用布雷菲德菌素A处理后和有丝分裂期间正常分散和重新组装。这些发现表明，在细胞中的NSF和p97的ATP水解依赖性活动是不相等的，并建议，只有NSF是直接参与调节膜融合。
NSF and p97 are related AAA proteins implicated in membrane trafficking and organelle biogenesis. p97 is also involved in pathways that lead to ubiquitin-dependent proteolysis, including ER-associated degradation (ERAD). In this study, we have used dominant interfering ATP-hydrolysis deficient mutants (NSF(E329Q) and p97(E578Q)) to compare the function of these AAA proteins in the secretory pathway of mammalian cells. Expressing NSF(E329Q) promotes disassembly of Golgi stacks into dispersed vesicular structures. It also rapidly inhibits glycosaminoglycan sulfation, reflecting disruption of intra-Golgi transport. In contrast, expressing p97(E578Q) does not affect Golgi structure or function; glycosaminoglycans are normally sulfated and secreted, as is the VSV-G ts045 protein. Instead, expression of p97(E578Q) causes ubiquitinated proteins to accumulate on ER membranes and slows degradation of the ERAD substrate cystic-fibrosis transmembrane-conductance regulator. In addition, expression of p97(E578Q) eventually causes the ER to swell. More specific assessment of effects of p97(E578Q) on organelle assembly shows that the Golgi apparatus disperses and reassembles normally after treatment with brefeldin A and during mitosis. These findings demonstrate that ATP-hydrolysis-dependent activities of NSF and p97 in the cell are not equivalent and suggest that only NSF is directly involved in regulating membrane fusion.