2017 Fiscal Year Research-status Report
Studies on influenza A virus M2-host interactome and its role in virus replication
| Project/Area Number |
16K08014
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| Research Institution | Hokkaido University |
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Principal Investigator |
マンズール ラシッド 北海道大学, 人獣共通感染症リサーチセンター, 特任助教 (90566150)
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| Project Period (FY) |
2016-04-01 – 2019-03-31
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| Keywords | Influenza A virus / M2 protein |
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| Outline of Annual Research Achievements |
Though primarily an ion channel, the M2 protein is not only involved in various steps of viral replication but also affects the host cellular functions in ion channel activity-dependent and -independent manners. While en route to the cell membrane, the M2-ectodomain and cytoplasmic tail are exposed to vesicular/organellar and cytoplasmic environments, where they may interact with various cellular proteins. Therefore, in this study, we plan to investigate the role of M2-interacting host proteins in virus replication.
The M2 protein functions at various steps of virus replication, therefore, the roles of identified host proteins were further evaluated by comparing the infectivity rates of cells and virus release by plaque assay. For this purpose, a plaque assay suitable for large scale screening was developed. Furthermore, to assess the role of host factors in M2-mediated functions, stable cell lines expressing M2 protein were generated using 239 and A549 cell lines and characterized. An assay to assess the M2-ion channel function was also developed and optimized. Moreover, optimal technique for plasma membrane proteome from mock and M2-expressing cells was developed and attempts were made to analyze by 2D-electrophortesis.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
The role of identified host proteins in influenza A virus M2 intracellular transport to plasma membrane is being assessed using M2 expressing stable cell lines. Alongside, further investigations are also being conducted to study the role of shortlisted host proteins in M2-function related stages of virus replication.
Presently, 2D-electrophoresis for studying membrane proteome is being optimized.
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| Strategy for Future Research Activity |
1. To continue to investigate the role of shortlisted host proteins in influenza virus replication with relevance to M2 protein function.
2. To optimize 2D-electrophoresis for studying the host plasma membrane proteome in M2 expressing cells.
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| Causes of Carryover |
The manuscript submission was planned, but got shifted to next fiscal year. The M2 protein is cytotoxic, so development of M2-expressing stable cell lines took time, therefore, the consumables and reagents required could not be purchased during this fiscal year.
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