A single tyrosine in the severe acute respiratory syndrome coronavirus membrane protein cytoplasmic tail is important for efficient interaction with spike protein.
Severe acute respiratory syndrome coronavirus (SARS-CoV) encodes 3 major envelope proteins: spike (S), membrane (M), and envelope (E). Previous work identified a dibasic endoplasmic reticulum retrieval signal in the cytoplasmic tail of SARS-CoV S that promotes efficient interaction with SARS-CoV M. The dibasic signal was shown to be important for concentrating S near the virus assembly site rather than for direct interaction with M. Here, we investigated the sequence requirements of the SARS-CoV M protein that are necessary for interaction with SARS-CoV S. The SARS-CoV M tail was shown to be necessary for S localization in the Golgi region when the proteins were exogenously coexpressed in cells. This was specific, since SARS-CoV M did not retain an unrelated glycoprotein in the Golgi. Importantly, we found that an essential tyrosine residue in the SARS-CoV M cytoplasmic tail, Y(195), was important for S-M interaction. When Y(195) was mutated to alanine, M(Y195A) no longer retained S intracellularly at the Golgi. Unlike wild-type M, M(Y195A) did not reduce the amount of SARS-CoV S carbohydrate processing or surface levels when the two proteins were coexpressed. Mutating Y(195) also disrupted SARS-CoV S-M interaction in vitro. These results suggest that Y(195) is necessary for efficient SARS-CoV S-M interaction and, thus, has a significant involvement in assembly of infectious virus.
McBride
Corrin E
CE
Department of Cell Biology, The Johns Hopkins University School of Medicine, 725 N. Wolfe St., Baltimore, MD 21205, USA.
Machamer
Carolyn E
CE
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D002460:Cell Line; D000086862:Coronavirus M Proteins; D004279:DNA, Viral / Q000235:genetics; D006056:Golgi Apparatus / Q000378:metabolism / Q000821:virology; D006367:HeLa Cells; D006801:Humans; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids / Q000235:genetics; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D064370:Spike Glycoprotein, Coronavirus; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D014763:Viral Matrix Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D019065:Virus Assembly / Q000235:genetics / Q000502:physiology
D000086862:Coronavirus M Proteins; D004279:DNA, Viral; C000706237:M protein, SARS-CoV; D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; D014763:Viral Matrix Proteins; D014443:Tyrosine