Genome-wide analysis of vaccinia virus protein–protein interactions

Abstract

To detect interactions between proteins of vaccinia virus, we carried out a comprehensive two-hybrid analysis to assay every pairwise combination. We constructed an array of yeast transformants that contained each of the 266 predicted viral ORFs as Gal4 activation domain hybrid proteins. The array was individually mated to transformants containing each ORF as a Gal4–DNA-binding domain hybrid, and diploids expressing the two-hybrid reporter gene were identified. Of the ≈70,000 combinations, we found 37 protein–protein interactions, including 28 that were previously unknown. In some cases, e.g., late transcription factors, both proteins were known to have related roles although there was no prior evidence of physical associations. For some other interactions, neither protein had a known role. In the majority of cases, however, one of the interacting proteins was known to be involved in DNA replication, transcription, virion structure, or host evasion, thereby providing a clue to the role of the other uncharacterized protein in a specific process.