Raynor C M, Wright J F, Waisman D M, Pryzdial E L
Canadian Blood Services, R&D Department, University of Ottawa, Ontario, Canada.
Biochemistry. 1999 Apr 20;38(16):5089-95. doi: 10.1021/bi982095b.
A number of studies have suggested that the anionic phospholipid (anPL)-binding protein annexin II may play a role in cytomegalovirus (CMV) infection. Since annexin II has been shown to mediate aggregation and fusion of certain membranes, we investigated whether these properties could be exploited by CMV directly. The experiments showed that purified annexin II, but not the homologous protein annexin V (AnV), can mediate the binding of 35S-CMV (strain AD169) to anPL-coated microtiter wells. This association required Ca2+, could be titrated by varying either annexin II (apparent Kd = 4 x 10(-)8 M) or 35S-CMV, was inhibited by unlabeled CMV, and was observed for the heterotetrameric or monomeric form of annexin II. In experiments utilizing the fluorescence dequenching of octadecyl rhodamine incorporated into the CMV envelope, annexin II was furthermore found to enhance the rate of virus-anPL vesicle fusion. The observed fusion was dependent on the concentration of annexin II, Ca2+, and anPL and was mediated principally by the heterotetramer. Interestingly, AnV was observed to inhibit the effects of annexin II on CMV fusion but not binding to anPL, which indicates that annexin II enhances these processes by distinct mechanisms. The results presented here provide the first direct evidence that annexin II has the capacity to bridge CMV to a phospholipid membrane and to enhance virus-membrane fusion. These observations furthermore suggest that AnV may regulate the fusogenic function of annexin II.
多项研究表明，阴离子磷脂（anPL）结合蛋白膜联蛋白II可能在巨细胞病毒（CMV）感染中发挥作用。由于膜联蛋白II已被证明可介导某些膜的聚集和融合，我们研究了CMV是否能直接利用这些特性。实验表明，纯化的膜联蛋白II而非同源蛋白膜联蛋白V（AnV），可介导35S-CMV（AD169株）与包被有anPL的微量滴定孔结合。这种结合需要Ca2+，可通过改变膜联蛋白II（表观解离常数Kd = 4×10⁻⁸ M）或35S-CMV进行滴定，未标记的CMV可抑制该结合，且在膜联蛋白II的异源四聚体或单体形式中均观察到这种结合。在利用掺入CMV包膜的十八烷基罗丹明荧光猝灭的实验中，还发现膜联蛋白II可提高病毒-anPL囊泡融合的速率。观察到的融合取决于膜联蛋白II、Ca2+和anPL的浓度，且主要由异源四聚体介导。有趣的是，观察到AnV可抑制膜联蛋白II对CMV融合的作用，但不影响其与anPL的结合，这表明膜联蛋白II通过不同机制增强了这些过程。此处呈现的结果提供了首个直接证据，表明膜联蛋白II有能力将CMV与磷脂膜连接起来并增强病毒-膜融合。这些观察结果还表明，AnV可能调节膜联蛋白II的融合功能。
