Viral proteomics: The emerging cutting-edge of virus research

Zhou, ShengTao; Liu, Rui; Zhao, Xia; Huang, CanHua; Wei, YuQuan

The recently established quantitative proteomic technology, SILAC, has shown great potential in protein expression profiling. SILAC involves a straightforward metabolic labeling method, where the cells are grown in medium containing amino acids with stable nonabundant isotopes (e.g., 15 N, 13 C and 2 D) and compared with the cells grown in medium containing stable abundant isotopes ( 14 N, 12 C and 1 H). Protein abundance is calculated as the ratios of the peak intensity of the fragment ions from the labeled versus the unlabeled peptides, thus constituting an ideal tool for protein expression analysis upon viral infection [51] . Mannova et al. [52] used 2DE followed by mass spectrometry or SILAC combined with one-dimensional electrophoresis separation and mass spectrometry to quantify the modification of the host lipid raft proteome upon HCV infection. While the 2DE and mass spectrometry strategy identified approximately 100 protein spots, the number of individual proteins characterized by the SILAC approach totaled 1036. Many of these proteins were involved in vesicular and protein trafficking and cell signaling. Using small interfering RNAs modulating small GTPases Cdc42 and RhoA, this study noticed an increase in HCV replication. On the other hand, decreased syntaxin 7 expression led to decreased replication of HCV. This proteomics study indicated that protein subcellular relocalization occurs in HCV-containing cells, which can directly influence HCV replication. Recently, Zhang et al. [53] developed a cell line expressing a SARS-CoV subgenomic replicon and used it to screen inhibitors of SARS-CoV replication. The protein profiles of the SARS-CoV replicon cells and parental BHK21 cells were compared by SILAC coupled with mass spectrometry to identify the host proteins essential for SARS-CoV RNA replication. Among the 1081 host proteins quantified, 74 had significantly altered levels of expression. Of these, BCL2-associated athanogene 3 (BAG3), a multifunctional molecule proved to participate in cell survival, cellular stress response, proliferation, migration, and apoptosis, was subjected to further functional studies. The results indicated that inhibition of BAG3 expression by RNA interference led to significant suppression of SARS-CoV replication, suggesting a critical role of BAG3 in SARS-CoV replication.