Detection and characterization of proteins encoded by the second ORF of the M2 gene of pneumoviruses

• Published in: Journal of general virology Vol. 80; no. 8; pp. 2011 - 2016
• Main Authors: Ahmadian, G, Chambers, P, Easton, A. J
• Format: Journal Article
• Language: English
• Published: England Soc General Microbiol 01-08-1999
• Subjects: Amino Acid Sequence; Animals; Avian pneumovirus; Base Sequence; Bovine respiratory syncytial virus; Cell Line; Chlorocebus aethiops; DNA, Viral; Human respiratory syncytial virus; Humans; Mice; Molecular Sequence Data; Open Reading Frames; Pneumonia virus of mice; Pneumovirus; Pneumovirus - genetics; Pneumovirus - immunology; Viral Matrix Proteins - genetics; Viral Matrix Proteins - immunology
• ISSN: 0022-1317; 1465-2099
• DOI: 10.1099/0022-1317-80-8-2011

Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK 1
Author for correspondence: Andrew Easton.Fax [phone]. e-mail ae{at}dna.bio.warwick.ac.uk
The nucleotide sequence of the M2 gene of pneumonia virus of mice (PVM) was determined. The sequence showed that the gene encoded a protein of 176 amino acids with a predicted molecular mass of 20165 Da from a major ORF, which is smaller than the equivalent proteins encoded by human, bovine and ovine respiratory syncytial (RS) viruses. The PVM M2 protein is conserved, having 41% similarity to the equivalent human RS virus protein. In common with the M2 genes of the RS viruses and avian pneumovirus (APV), the PVM mRNA also contained a second ORF (ORF2) that partially overlaps the first ORF and which is capable of encoding a 98 residue polypeptide. No significant sequence identity could be detected between the putative M2 ORF2 proteins of PVM, APV and the RS viruses. The expression of the M2 ORF2 proteins of the pneumoviruses was investigated by using monospecific antisera raised against GST fusion proteins. Western blot analysis demonstrated the presence of polypeptides encoded by M2 ORF2 of PVM and RS virus corresponding with those predicted by in vitro translation studies, but this was not the case for APV. The PVM polypeptide was present as three distinct products in vivo . The PVM and RS virus polypeptides were also detected in cells by immunofluorescence, which showed that both were present in the cytoplasm with a degree of localization in inclusion bodies. No APV M2 ORF2 protein could be detected in vivo . The RS virus M2 ORF2 polypeptide was shown to accumulate during infection and the potential implications of this are discussed.