H5N1 virus causes significant perturbations in host proteome very early in influenza virus-infected primary human monocyte-derived macrophages

Abstract

H5N1 influenza viruses, which cause disease in humans, have unusually high pathogenicity. The temporal response of primary human monocyte-derived macrophages infected with highly pathogenic H5N1 and seasonal H1N1 influenza viruses was evaluated using mass spectrometry-based quantitative proteomic profiling. This was done in order to demonstrate significant perturbation of the host proteome upon viral infection, as early as 1 hour after infection. This early host response distinguished H5N1 infection from H1N1 infection, the latter inducing less of a response. The most pronounced effect was observed on the translational machinery, suggesting that H5N1 might gain advantage in replication by using the cell protein synthesis machinery early in the infection.

Figure 1.

Analysis of the differentially abundant proteins using Ingenuity Pathways Analysis (IPA). Protein synthesis, molecular transport, and nucleic acid metabolism networks as defined by IPA. A, The network modulation is shown for H5N1-infected monocyte-derived macrophages (MDMs) at 1 hour p.i. The blue circles identify double-stranded DNA break repair proteins. Blue squares identify proteins associated with virus infection. The red circle defines a group associated with molecular transport and nucleic acid metabolism. B, Protein synthesis machinery proteins differentially expressed at 1 hour p.i. in H5N1-infected MDMs. Blue squares identify proteins that belong to the protein elongation functional group in GO. C, Time-course changes in protein abundance in both H1N1- and H5N1-infected MDMs for the network described in part A.

Figure 2.

A, Gene expression analysis by real-time polymerase chain reaction (PCR) and Western blot. Real-time PCR measurement of mRNA abundance suggests that the dysferlin is expressed at the much lower level to RPS18. RPL14 and RPS5 are expressed at similar levels higher than dysferlin but lower than RPS18. B, Analysis by Western blot for RPS18 in monocyte-derived macrophages infected with influenza viruses C, The corresponding densitometry measurements (mean of 3 independent replicates). Error bars indicate standard error.