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Annexin II enhances cytomegalovirus binding and fusion to phospholipid membranes
C M Raynor1, J F Wright, D M Waisman
1Canadian Blood Services, R&D Department, University of Ottawa, Ontario, Canada.
Abstract:
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.
Insights
Annexin II bridges human cytomegalovirus (CMV) to anionic phospholipid membranes and enhances virus-membrane fusion. Annexin V inhibits this fusion, suggesting a regulatory role in CMV infection.
Area of Science:
- Virology
- Molecular Biology
- Cell Biology
Background:
- Annexin II, an anionic phospholipid-binding protein, is implicated in cytomegalovirus (CMV) infection.
- Annexin II's known membrane aggregation and fusion properties prompted investigation into its role in CMV.
- Cytomegalovirus (CMV) is a significant human pathogen with complex infection mechanisms.
Purpose of the Study:
- To investigate if annexin II directly exploits its membrane-binding and fusion properties for CMV infection.
- To determine the mechanism by which annexin II interacts with CMV and phospholipid membranes.
- To explore the potential regulatory role of annexin V in annexin II-mediated CMV fusion.
Main Methods:
- Binding assays using purified annexin II and 35S-labeled CMV to anionic phospholipid-coated microtiter wells.
- Fluorescence dequenching assays to measure virus-vesicle fusion rates.
- Experiments utilizing varying concentrations of annexin II, Ca2+, and anionic phospholipids.
- Inhibition studies using unlabeled CMV and annexin V.
Main Results:
- Annexin II, but not annexin V, mediated the binding of CMV to anionic phospholipid membranes in a Ca2+-dependent manner.
- Annexin II significantly enhanced the rate of CMV-anionic phospholipid vesicle fusion, primarily via its heterotetrameric form.
- Annexin V inhibited annexin II-mediated CMV fusion but not binding, indicating distinct mechanisms.
Conclusions:
- Annexin II directly bridges CMV to phospholipid membranes and enhances virus-membrane fusion.
- These findings provide the first direct evidence of annexin II's role in facilitating CMV entry.
- Annexin V may act as a regulator of annexin II's fusogenic function in CMV infection.