Giuseppe
Emeritus
J Virol. 2009 Feb 4. [Epub ahead of print]
DETECTION AND CHARACTERIZATION OF INFLUENZA A VIRUS PA-PB2 INTERACTION THROUGH A BIMOLECULAR FLOURESENCE COMPLEMENTATION ASSAY.
Hemerka JN, Wang D, Weng Y, Lu W, Kaushik RS, Jin J, Harmon AF, Li F. - Department of Biology and Microbiology, Department of Veterinary Science, Center for Infectious Disease Research and Vaccinology, South Dakota State University, Brookings, SD 57007; Section of Microbial Pathogenesis, Yale University School of Medicine, 295 Congress Ave, New Haven, CT 06536.
The influenza virus polymerase complex, consisting of the PA, PB1, and PB2 subunits, is required for the transcription and replication of the influenza A viral genome.
Previous studies have shown that PB1 serves as a core subunit to incorporate PA and PB2 into the polymerase complex by directly interacting with PA and PB2.
Despite numerous attempts, largely involving biochemical approaches, a specific interaction between PA and PB2 subunits has yet to be detected.
In the current study, we developed and utilized bimolecular fluorescence complementation (BiFC) to study protein-protein interactions in the assembly of the influenza A virus polymerase complex.
Proof-of-concept experiments demonstrated that BiFC can specifically detect PA-PB1 interaction in living cells. Strikingly, BiFC demonstrated an interaction between PA and PB2 that has not been reported previously.
Deletion-based BiFC experiments indicated that the N-terminal 100 amino acid residues of PA are responsible for the PA-PB2 interaction observed in BiFC.
Furthermore, a detailed analysis of subcellular localization patterns and temporal nuclear import of PA-PB2 binary complexes suggested that PA and PB2 subunits interacted in the cytoplasm initially and were subsequently transported as a dimer into the nucleus.
Taken together, results of our studies revealed a previously unknown PA-PB2 interaction and provided a frame work for further investigation of the biological relevance of PA-PB2 interaction in the polymerase activity and viral replication of influenza A virus.
PMID: 19193801 [PubMed - as supplied by publisher]
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DETECTION AND CHARACTERIZATION OF INFLUENZA A VIRUS PA-PB2 INTERACTION THROUGH A BIMOLECULAR FLOURESENCE COMPLEMENTATION ASSAY.
Hemerka JN, Wang D, Weng Y, Lu W, Kaushik RS, Jin J, Harmon AF, Li F. - Department of Biology and Microbiology, Department of Veterinary Science, Center for Infectious Disease Research and Vaccinology, South Dakota State University, Brookings, SD 57007; Section of Microbial Pathogenesis, Yale University School of Medicine, 295 Congress Ave, New Haven, CT 06536.
The influenza virus polymerase complex, consisting of the PA, PB1, and PB2 subunits, is required for the transcription and replication of the influenza A viral genome.
Previous studies have shown that PB1 serves as a core subunit to incorporate PA and PB2 into the polymerase complex by directly interacting with PA and PB2.
Despite numerous attempts, largely involving biochemical approaches, a specific interaction between PA and PB2 subunits has yet to be detected.
In the current study, we developed and utilized bimolecular fluorescence complementation (BiFC) to study protein-protein interactions in the assembly of the influenza A virus polymerase complex.
Proof-of-concept experiments demonstrated that BiFC can specifically detect PA-PB1 interaction in living cells. Strikingly, BiFC demonstrated an interaction between PA and PB2 that has not been reported previously.
Deletion-based BiFC experiments indicated that the N-terminal 100 amino acid residues of PA are responsible for the PA-PB2 interaction observed in BiFC.
Furthermore, a detailed analysis of subcellular localization patterns and temporal nuclear import of PA-PB2 binary complexes suggested that PA and PB2 subunits interacted in the cytoplasm initially and were subsequently transported as a dimer into the nucleus.
Taken together, results of our studies revealed a previously unknown PA-PB2 interaction and provided a frame work for further investigation of the biological relevance of PA-PB2 interaction in the polymerase activity and viral replication of influenza A virus.
PMID: 19193801 [PubMed - as supplied by publisher]
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