Abstract
The coronavirus' (CoV) membrane (M) protein is the driving force during assembly, but this process remains poorly characterized. Previously, we described two motifs in the C-tail of the Middle East respiratory syndrome CoV (MERS-CoV) M protein involved in its endoplasmic reticulum (ER) exit (211DxE213) and trans-Golgi network (TGN) retention (199KxGxYR204). Here, their function in virus assembly was investigated by two different virus-like particle (VLP) assays and by mutating both motifs in an infectious MERS-CoV cDNA clone. It was shown that the 199KxGxYR204 motif was essential for VLP and infectious virus assembly. Moreover, the mislocalization of the M protein induced by mutation of this motif prevented M-E interaction. Hampering the ER export of M by mutating its 211DxE213 motif still allowed the formation of nucleocapsid-empty VLPs, but prevented the formation of fully assembled VLPs and infectious particles. Taken together, these data show that the MERS-CoV assembly process highly depends on the correct intracellular trafficking of its M protein, and hence that not only specific protein-protein interacting motifs but also correct subcellular localization of the M protein in infected cells is essential for virus formation and should be taken into consideration when studying the assembly process.