The Membrane Protein of Severe Acute Respiratory Syndrome Coronavirus Functions as a Novel Cytosolic Pathogen-Associated Molecular Pattern To Promote Beta Interferon Induction via a Toll-Like-Receptor-Related TRAF3-Independent Mechanism

Wang, Yi; Liu, Li

One critical question remaining to be answered is whether or not M-mediated IFN-␤ induction could occur in real virus infection. It has been shown that codelivering the M, N, and S genes of SARS-CoV into HEK293 cells readily produced SARS-CoV pseudovirus with the corona-like halo (35) . Earlier results demonstrate that valine-toalanine mutation at residue 68 [abbreviated as M(V68A)] inhibited IFN-␤ induction (34) . We employed a SARS-CoV pseudovirus system to mimic the real SARS-CoV infection. Cell lysate supernatants isolated from either M, N, and S cotransfection or M(V68A), N, and S cotransfection were first incubated with 293-ACE2 stable cells (Fig. 8A) for 4 h. Then, the cell culture medium was replaced with fresh medium and further incubated at 37°C for another 24 h before harvesting. The SARS-CoV pseudovirus VLP(S-M-N) markedly upregulated IFN-␤ expression at both RNA and protein levels (Fig. 8B) , whereas the point mutation at the valine 68 residue of M protein completely diminished SARS-CoV pseudovirus-mediated IFN-␤ induction, strongly indicating that the M protein residing on SARS-CoV virion could specifically induce IFN-␤ production upon infecting the targeted cells. Taken together, our data indicate for the first time that SARS-CoV M protein may function as a novel cytosolic PAMP to activate IFN-␤ induction through an intracellular TLR-related signaling pathway in a TRAF3-independent manner.