The CM2 protein of influenza C virus is an oligomeric integral membrane glycoprotein structurally analogous to influenza A virus M2 and influenza B virus NB proteins.
We have undertaken a characterization of the CM2 protein of influenza C virus. The CM2 coding region of RNA segment 6 (nucleotides 731-1147) was cloned from two strains of influenza C virus and expressed using the vaccinia virus-bacteriophage T7 RNA polymerase (vac-T7) system. An antiserum raised to a C-terminal peptide in the CM2 open reading frame recognized the CM2 protein in influenza C virus-infected cells and after vac-T7 expression of the CM2 open reading frame. CM2 is posttranslationally modified by addition of high-mannose carbohydrate chains (Mr approximately 18 kDa) and by further addition of polylactosaminoglycans (Mr approximately 21-35 kDa). The available data indicate that CM2 has a cleavable signal peptide at the N-terminus of the protein. Site-directed mutagenesis eliminated the single potential N-linked carbohydrate attachment site on CM2 and indicated that the protein has an NoutCin orientation in membranes. Nonreducing SDS-PAGE indicated that the protein was expressed as disulfide-linked dimers and tetramers. Cell surface biotinylation and indirect immunofluorescence showed the protein to be expressed at the cell surface. Elimination of the N-linked carbohydrate attachment site and addition of a C-terminal HA epitope tag did not adversely affect surface expression of CM2. The NoutCin membrane orientation of CM2, the size of the ectodomain and cytoplasmic tail of CM2, and its ability to form disulfide-linked oligomers are reminiscent of the structural properties of influenza A virus M2 and influenza B virus NB proteins.
Pekosz
A
A
Howard Hughes Medical Institute, Northwestern University, Evanston, Illinois 60208-3500, USA.
Lamb
R A
RA
D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis; D019281:Dimerization; D009980:Influenza A virus / Q000378:metabolism; D009981:Influenza B virus / Q000378:metabolism; D009982:Gammainfluenzavirus / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D014763:Viral Matrix Proteins / Q000737:chemistry* / Q000235:genetics; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics; D000086082:Viroporin Proteins
D018076:DNA, Complementary; D014763:Viral Matrix Proteins; D014764:Viral Proteins; C090877:CM2 protein, Influenza C virus; C031540:M-protein, influenza virus; C488937:M2 protein, Influenza A virus; D000086082:Viroporin Proteins; C051094:NB glycoprotein, Influenza B virus