Coupling high-throughput genetics with phylogenetic information reveals an epistatic interaction on the XXXX M segment.

@entity11317 @entity72818 @entity72817
BACKGROUND: Epistasis is one of the central themes in viral evolution due to its importance in drug resistance, immune escape, and interspecies transmission. However, there is a lack of experimental approach to systematically probe for epistatic residues. RESULTS: By utilizing the information from natural occurring sequences and high-throughput genetics, this study established a novel strategy to identify epistatic residues. The rationale is that a substitution that is deleterious in one strain may be prevalent in nature due to the presence of a naturally occurring compensatory substitution. Here, high-throughput genetics was applied to @entity11317 M segment to systematically identify deleterious substitutions. Comparison with natural sequence variation showed that a deleterious substitution M1 @entity72817 was prevalent in circulating strains. A coevolution analysis was then performed and indicated that M1 residues 121, 207, 209, and 214 naturally coevolved as a group. Subsequently, we experimentally validated that M1 @entity72818 was a compensatory substitution for M1 @entity72817 . CONCLUSIONS: This work provided a proof-of-concept to identify epistatic residues by coupling high-throughput genetics with phylogenetic information. In particular, we were able to identify an epistatic interaction between M1 substitutions @entity72818 and @entity72817 . This analytic strategy can potentially be adapted to study any protein of interest, provided that the information on natural sequence variants is available.
