Quantitative Proteomic Analysis of A549 Cells Infected with Human Respiratory Syncytial Virus

Munday, Diane C.; Emmott, Edward; Surtees, Rebecca; Lardeau, Charles-Hugues; Wu, Weining; Duprex, W. Paul; Dove, Brian K.; Barr, John N.; Hiscox, Julian A.

Alteration of Cell Cycle Regulatory Proteins in HRSV-infected Cells-The quantitative proteomics and network pathway analysis (Fig. 3) identified several proteins with roles in cell cycle regulation whose abundance differed between mock-infected and HRSV-infected cells. These included Cdc2 (also known as cyclin-dependent kinase 1 (cdk1)); histone deacetylase 2 (HDAC2); proliferation-associated 2G4, 38 kDa (PA2G4); and SIN3 homolog A transcription regulator (yeast) (SIN3A) (ϳ6-, 4-, 3-, and 3-fold less abundant, respectively, in HRSV-infected cells). Network pathway analysis (Fig. 3 ) also predicted that cyclin A might be altered in HRSV-infected cells. Cdc2 has been shown to bind to cyclins such as A, E, and B types and can regulate cell cycle progression (59, 60) . In addition, in HRSV-infected A549 cells, cell cycle arrest has been observed; however, the abundance of cell cycle regulatory complexes was not elucidated (18) . Western blot analysis of nuclear and cytoplasmic fractions from mock-and HRSVinfected cells indicated that Cdc2 was less abundant in the nuclear fraction in HRSV-infected cells (confirming the quantitative proteomic analysis) and that cyclins D2, A, E, and B1 were also less abundant (Fig. 11) . In such analysis, it is essential to ensure that equal protein loading is used to allow for relative protein abundance observations. Lamin B and tubulin were therefore selected as internal controls to confirm protein content.