Alteration of Protein Levels during Influenza Virus H1N1 Infection in Host Cells: A Proteomic Survey of Host and Virus Reveals Differential Dynamics

Abstract

We studied the dynamics of the proteome of influenza virus A/PR/8/34 (H1N1) infected Madin-Darby canine kidney cells up to 12 hours post infection by mass spectrometry based quantitative proteomics using the approach of stable isotope labeling by amino acids in cell culture (SILAC). We identified 1311 cell proteins and, apart from the proton channel M2, all major virus proteins. Based on their abundance two groups of virus proteins could be distinguished being in line with the function of the proteins in genesis and formation of new virions. Further, the data indicate a correlation between the amount of proteins synthesized and their previously determined copy number inside the viral particle. We employed bioinformatic approaches such as functional clustering, gene ontology, and pathway (KEGG) enrichment tests to uncover co-regulated cellular protein sets, assigned the individual subsets to their biological function, and determined their interrelation within the progression of viral infection. For the first time we are able to describe dynamic changes of the cellular and, of note, the viral proteome in a time dependent manner simultaneously. Through cluster analysis, time dependent patterns of protein abundances revealed highly dynamic up- and/or down-regulation processes. Taken together our study provides strong evidence that virus infection has a major impact on the cell status at the protein level.

Proteomic analysis reveals down-regulation of surfactant protein B in murine type II pneumocytes infected with influenza A virus

“…NS1 has multiple roles during IAV infection, including inhibition of cellular gene expression and the type I interferon response, and is also expressed at high levels in infected cells (Fernandez-Sesma et al, 2006; Garcia-Sastre et al, 1998). Proteins that form the viral RNA-dependent RNA polymerase complex, PB1, PB2, and PA, the nuclear export protein, NEP, and M2 protein are expressed in relatively low abundance in infected cells (Kummer et al, 2014) and were not detected by our analysis. Overall, we detected a similar profile of viral proteins as others have reported in proteomics analyses of influenza virus- infected cells and animals (Brown et al, 2010; Lietzen et al, 2011; Liu et al, 2012).…”

“…Peptides mapping to NP and HA were also detected in two or fewer of the mock samples. Kummer et al demonstrated that these five proteins are the most abundant viral proteins in PR8-infected MDCK cells (Kummer et al, 2014). HA, NA, NP, and M1 are present in infectious virions and are produced in great abundance during viral assembly (Hutchinson et al, 2014; Kummer et al, 2014).…”

“…Kummer et al demonstrated that these five proteins are the most abundant viral proteins in PR8-infected MDCK cells (Kummer et al, 2014). HA, NA, NP, and M1 are present in infectious virions and are produced in great abundance during viral assembly (Hutchinson et al, 2014; Kummer et al, 2014). Therefore, detecting these proteins late during the replication cycle (24 h post-infection) is not surprising and suggests that influenza virus is actively replicating in these cells.…”

“…Proteomics studies of influenza virus infections in macaques (Brown et al, 2010), mice (Kumar et al, 2014), continuous cell lines (Coombs et al, 2010; Kummer et al, 2014; Vester et al, 2009), and primary human cells (Kroeker et al, 2012; Lietzen et al, 2011; Liu et al, 2012) have provided insight into how IAV strains alter the host proteome at cellular and organismal levels. However, host responses to IAV infection are also, in part, specific to the cell type.…”

Positive Selection in CD8+T-Cell Epitopes of Influenza Virus Nucleoprotein Revealed by a Comparative Analysis of Human and Swine Viral Lineages

“…Our goal is to determine whether epitopes targeted by human CD8 ϩ T cells are under selection in influenza viruses that circulate in human hosts. The two most highly expressed influenza virus proteins are NP and M1 (49), and epitopes in these proteins are major targets of CD8 ϩ T cells (3,4,8,9). The NP and M1 proteins in contemporary human H3N2 influenza viruses have circulated in humans since at least 1918 (50,51).…”