A computational-experimental approach identifies mutations that enhance surface expression of an
Jesse D Bloom1, Jagannath S Nayak, David Baltimore
1Division of Biology, California Institute of Technology, Pasadena, California, United States of America.
Abstract:
The His274→Tyr (H274Y) oseltamivir (Tamiflu) resistance mutation causes a substantial decrease in the total levels of surface-expressed neuraminidase protein and activity in early isolates of human seasonal H1N1 influenza, and in the swine-origin pandemic H1N1. In seasonal H1N1, H274Y only became widespread after the occurrence of secondary mutations that counteracted this decrease. H274Y is currently rare in pandemic H1N1, and it remains unclear whether secondary mutations exist that might similarly counteract the decreased neuraminidase surface expression associated with this resistance mutation in pandemic H1N1. Here we investigate the possibility of predicting such secondary mutations. We first test the ability of several computational approaches to retrospectively identify the secondary mutations that enhanced levels of surface-expressed neuraminidase protein and activity in seasonal H1N1 shortly before the emergence of oseltamivir resistance. We then use the most successful computational approach to predict a set of candidate secondary mutations to the pandemic H1N1 neuraminidase. We experimentally screen these mutations, and find that several of them do indeed partially counteract the decrease in neuraminidase surface expression caused by H274Y. Two of the secondary mutations together restore surface-expressed neuraminidase activity to wildtype levels, and also eliminate the very slight decrease in viral growth in tissue-culture caused by H274Y. Our work therefore demonstrates a combined computational-experimental approach for identifying mutations that enhance neuraminidase surface expression, and describes several specific mutations with the potential to be of relevance to the spread of oseltamivir resistance in pandemic H1N1.
Insights
The His274→Tyr mutation in influenza H1N1 reduces neuraminidase activity. Computational and experimental methods identified secondary mutations that restore neuraminidase expression and activity, potentially impacting oseltamivir resistance.
Area of Science:
- Virology
- Computational Biology
- Drug Resistance
Background:
- The His274→Tyr (H274Y) mutation confers oseltamivir resistance to influenza H1N1.
- This mutation significantly reduces surface neuraminidase (NA) protein levels and activity.
- Secondary mutations were required for H274Y to become widespread in seasonal H1N1, but their role in pandemic H1N1 is unclear.
Purpose of the Study:
- To investigate the potential for secondary mutations to counteract the NA reduction caused by H274Y in pandemic H1N1.
- To evaluate computational approaches for predicting such compensatory mutations.
- To experimentally validate predicted mutations.
Main Methods:
- Retrospective analysis of seasonal H1N1 to assess computational prediction accuracy.
- Application of the most successful computational method to predict secondary mutations in pandemic H1N1 NA.
- Experimental screening of predicted mutations for their effect on NA surface expression and activity.
Main Results:
- Several predicted secondary mutations partially restored NA surface expression reduced by H274Y.
- Two identified mutations restored NA activity to wild-type levels.
- These mutations also compensated for the slight decrease in viral growth associated with H274Y.
Conclusions:
- A combined computational-experimental approach can identify mutations that enhance NA surface expression.
- Specific secondary mutations can counteract the effects of the H274Y resistance mutation in pandemic H1N1.
- These findings are relevant to understanding and potentially managing oseltamivir resistance spread.
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