Subcellular proteomic analysis of human host cells infected with H3N2 swine influenza virus

Abstract

Cross-species transmissions of swine influenza viruses (SIVs) raise great public health concerns. In this study, subcellular proteomic profiles of human A549 cells inoculated with H3N2 subtype SIV were used to characterize dynamic cellular responses to infection. By 2DE and MS, 27 differentially expressed (13 upregulated, 14 downregulated) cytoplasmic proteins and 20 differentially expressed (13 upregulated, 7 downregulated) nuclear proteins were identified. Gene ontology analysis suggested that these differentially expressed proteins were mainly involved in cell death, stress response, lipid metabolism, cell signaling, and RNA PTMs. Moreover, 25 corresponding genes of the differentially expressed proteins were quantitated by real time RT-PCR to examine the transcriptional profiles between mock- and virus-infected A549 cells. Western blot analysis confirmed that changes in abundance of identified cellular proteins heterogeneous nuclear ribonucleoprotein (hnRNP) U, hnRNP C, ALDH1A1, tryptophanyl-tRNA synthetase, IFI35, and HSPB1 in H3N2 SIV-infected cells were consistent with results of 2DE analysis. By confocal microscopy, nucleus-to-cytoplasm translocation of hnRNP C and colocalization between the viral nonstructural protein 1 and hnRNP C as well as N-myc (and STAT) interactor were observed upon infection. Ingenuity Pathway Analysis revealed that cellular proteins altered during infection were grouped mainly into NFκB and interferon signaling networks. Collectively, these identified subcellular constituents provide an important framework for understanding host/SIV interactions and underlying mechanisms of SIV cross-species infection and pathogenesis.

Aminoacyl-tRNA synthetases, therapeutic targets for infectious diseases

“…An integrated analysis of microarray and proteomics data showed that hepatitis C virus (HCV)-infected hepatocytes up-regulate the WRS, SRS, AlaRS, TRS, and GRS genes and express higher levels of YRS and SRS proteins [24]. In A549 cells inoculated with swine influenza virus, WRS was up-regulated at both the mRNA and protein levels [25]. Increased expression of WRS was also observed in cells infected with Vibrio cholera, human cytomegalovirus, and human hepatitis B virus [22].…”

Circovirus Transport Proceeds via Direct Interaction of the Cytoplasmic Dynein IC1 Subunit with the Viral Capsid Protein

“…Isolation of nuclear components was performed using a nuclear and cytoplasmic protein extraction kit (P0027; Beyotime) as previously stated (54). Briefly, PCV2-infected PK15 cells were treated with 200 l of cytoplasmic protein extraction buffer A containing complete protease inhibitor cocktail for 5 to 10 min on ice and then added to 10 l buffer B.…”

Proteomic Analysis of Differential Expression of Cellular Proteins in Response to Avian H9N2 Virus Infection of A549 Cells

“…The ubiquitin-like modifier, ISG15, has been reported many times in the process of resisting influenza virus (Wu et al, 2013), and can be induced under virus and type I interferon stimulation. ISG15 plays an important role in regulating the antiviral innate immune response, and keeping interferon levels within a certain range, so that it can exert antiviral functions without causing immunologic damage.…”

“…Researchers have applied proteomic approaches to study cellular proteins involved in the process of H5N1, H3N2, and H1N1 virus infection (Baas et al, 2006; Mayer et al, 2007; Coombs et al, 2010; Wu et al, 2013). However, alterations of cellular proteins in human airway epithelial cell lines infected by H9N2 influenza virus have not been reported.…”