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. 1985 Nov;56(2):502–511. doi: 10.1128/jvi.56.2.502-511.1985

Characterization of the influenza virus M2 integral membrane protein and expression at the infected-cell surface from cloned cDNA.

S L Zebedee, C D Richardson, R A Lamb
PMCID: PMC252606  PMID: 3840537

Abstract

An investigation of properties of the influenza A virus M2 protein indicated that it is synthesized by 2 h postinfection together with other viral polypeptides and is transported to the infected-cell surface with a half-time of approximately 30 to 40 min. The available evidence suggests that M2 is not N-glycosylated even though it contains a potential glycosylation site, and the intracellular pattern of protein distribution includes localization to the Golgi apparatus. Proteolysis of intracellular microsome vesicles followed by immunoprecipitation with antiserum to a synthetic oligopeptide indicated that the M2 protein contains an extensive region of COOH-terminal amino acids exposed on the cytoplasmic side of the infected-cell membrane. A cDNA clone of the M2 mRNA was obtained and expressed in an SV40 recombinant vector. The M2 protein expressed by the vector became associated with the Golgi complex and was found on the surface of vector-infected cells. M2 is antigenically conserved among all strains of influenza virus both in regions exposed on the cell surface and intracellularly.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Allen H., McCauley J., Waterfield M., Gething M. J. Influenza virus RNA segment 7 has the coding capacity for two polypeptides. Virology. 1980 Dec;107(2):548–551. doi: 10.1016/0042-6822(80)90324-4. [DOI] [PubMed] [Google Scholar]
  2. Anderson D. J., Mostov K. E., Blobel G. Mechanisms of integration of de novo-synthesized polypeptides into membranes: signal-recognition particle is required for integration into microsomal membranes of calcium ATPase and of lens MP26 but not of cytochrome b5. Proc Natl Acad Sci U S A. 1983 Dec;80(23):7249–7253. doi: 10.1073/pnas.80.23.7249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dreyfuss G., Choi Y. D., Adam S. A. Characterization of heterogeneous nuclear RNA-protein complexes in vivo with monoclonal antibodies. Mol Cell Biol. 1984 Jun;4(6):1104–1114. doi: 10.1128/mcb.4.6.1104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Erickson A. H., Blobel G. Early events in the biosynthesis of the lysosomal enzyme cathepsin D. J Biol Chem. 1979 Dec 10;254(23):11771–11774. [PubMed] [Google Scholar]
  5. Gahmberg C. G., Jokinen M., Karhi K. K., Andersson L. C. Effect of tunicamycin on the biosynthesis of the major human red cell sialoglycoprotein, glycophorin A, in the leukemia cell line K562. J Biol Chem. 1980 Mar 10;255(5):2169–2175. [PubMed] [Google Scholar]
  6. Gibson R., Leavitt R., Kornfeld S., Schlesinger S. Synthesis and infectivity of vesicular stomatitis virus containing nonglycosylated G protein. Cell. 1978 Apr;13(4):671–679. doi: 10.1016/0092-8674(78)90217-9. [DOI] [PubMed] [Google Scholar]
  7. Guan J. L., Rose J. K. Conversion of a secretory protein into a transmembrane protein results in its transport to the Golgi complex but not to the cell surface. Cell. 1984 Jul;37(3):779–787. doi: 10.1016/0092-8674(84)90413-6. [DOI] [PubMed] [Google Scholar]
  8. Hirt B. Selective extraction of polyoma DNA from infected mouse cell cultures. J Mol Biol. 1967 Jun 14;26(2):365–369. doi: 10.1016/0022-2836(67)90307-5. [DOI] [PubMed] [Google Scholar]
  9. Inglis S. C., Brown C. M. Differences in the control of virus mRNA splicing during permissive or abortive infection with influenza A (fowl plague) virus. J Gen Virol. 1984 Jan;65(Pt 1):153–164. doi: 10.1099/0022-1317-65-1-153. [DOI] [PubMed] [Google Scholar]
  10. Lamb R. A., Choppin P. W. Identification of a second protein (M2) encoded by RNA segment 7 of influenza virus. Virology. 1981 Jul 30;112(2):729–737. doi: 10.1016/0042-6822(81)90317-2. [DOI] [PubMed] [Google Scholar]
  11. Lamb R. A., Choppin P. W. Segment 8 of the influenza virus genome is unique in coding for two polypeptides. Proc Natl Acad Sci U S A. 1979 Oct;76(10):4908–4912. doi: 10.1073/pnas.76.10.4908. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lamb R. A., Choppin P. W. Synthesis of influenza virus proteins in infected cells: translation of viral polypeptides, including three P polypeptides, from RNA produced by primary transcription. Virology. 1976 Oct 15;74(2):504–519. doi: 10.1016/0042-6822(76)90356-1. [DOI] [PubMed] [Google Scholar]
  13. Lamb R. A., Choppin P. W. The gene structure and replication of influenza virus. Annu Rev Biochem. 1983;52:467–506. doi: 10.1146/annurev.bi.52.070183.002343. [DOI] [PubMed] [Google Scholar]
  14. Lamb R. A., Etkind P. R., Choppin P. W. Evidence for a ninth influenza viral polypeptide. Virology. 1978 Nov;91(1):60–78. doi: 10.1016/0042-6822(78)90355-0. [DOI] [PubMed] [Google Scholar]
  15. Lamb R. A., Lai C. J., Choppin P. W. Sequences of mRNAs derived from genome RNA segment 7 of influenza virus: colinear and interrupted mRNAs code for overlapping proteins. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4170–4174. doi: 10.1073/pnas.78.7.4170. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lamb R. A., Lai C. J. Conservation of the influenza virus membrane protein (M1) amino acid sequence and an open reading frame of RNA segment 7 encoding a second protein (M2) in H1N1 and H3N2 strains. Virology. 1981 Jul 30;112(2):746–751. doi: 10.1016/0042-6822(81)90319-6. [DOI] [PubMed] [Google Scholar]
  17. Lamb R. A., Lai C. J. Expression of unspliced NS1 mRNA, spliced NS2 mRNA, and a spliced chimera mRNA from cloned influenza virus NS DNA in an SV40 vector. Virology. 1984 May;135(1):139–147. doi: 10.1016/0042-6822(84)90124-7. [DOI] [PubMed] [Google Scholar]
  18. Lamb R. A., Lai C. J. Spliced and unspliced messenger RNAs synthesized from cloned influenza virus M DNA in an SV40 vector: expression of the influenza virus membrane protein (M1). Virology. 1982 Dec;123(2):237–256. doi: 10.1016/0042-6822(82)90258-6. [DOI] [PubMed] [Google Scholar]
  19. Lamb R. A., Zebedee S. L., Richardson C. D. Influenza virus M2 protein is an integral membrane protein expressed on the infected-cell surface. Cell. 1985 Mar;40(3):627–633. doi: 10.1016/0092-8674(85)90211-9. [DOI] [PubMed] [Google Scholar]
  20. Matlin K. S., Simons K. Reduced temperature prevents transfer of a membrane glycoprotein to the cell surface but does not prevent terminal glycosylation. Cell. 1983 Aug;34(1):233–243. doi: 10.1016/0092-8674(83)90154-x. [DOI] [PubMed] [Google Scholar]
  21. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  22. McCauley J. W., Mahy B. W., Inglis S. C. Nucleotide sequence of fowl plague virus RNA segment 7. J Gen Virol. 1982 Jan;58(Pt 1):211–215. doi: 10.1099/0022-1317-58-1-211. [DOI] [PubMed] [Google Scholar]
  23. Mulligan R. C., Howard B. H., Berg P. Synthesis of rabbit beta-globin in cultured monkey kidney cells following infection with a SV40 beta-globin recombinant genome. Nature. 1979 Jan 11;277(5692):108–114. doi: 10.1038/277108a0. [DOI] [PubMed] [Google Scholar]
  24. Nakamura K., Compans R. W. Effects of glucosamine, 2-deoxyglucose, and tunicamycin on glycosylation, sulfation, and assembly of influenza viral proteins. Virology. 1978 Feb;84(2):303–319. doi: 10.1016/0042-6822(78)90250-7. [DOI] [PubMed] [Google Scholar]
  25. Okada Y., Frey A. B., Guenthner T. M., Oesch F., Sabatini D. D., Kreibich G. Studies on the biosynthesis of microsomal membrane proteins. Site of synthesis and mode of insertion of cytochrome b5, cytochrome b5 reductase, cytochrome P-450 reductase and epoxide hydrolase. Eur J Biochem. 1982 Feb;122(2):393–402. doi: 10.1111/j.1432-1033.1982.tb05894.x. [DOI] [PubMed] [Google Scholar]
  26. Ortín J., Martínez C., del Río L., Dávila M., López-Galíndez C., Villanueva N., Domingo E. Evolution of the nucleotide sequence of influenza virus RNA segment 7 during drift of the H3N2 subtype. Gene. 1983 Aug;23(2):233–239. doi: 10.1016/0378-1119(83)90055-0. [DOI] [PubMed] [Google Scholar]
  27. Paterson R. G., Harris T. J., Lamb R. A. Analysis and gene assignment of mRNAs of a paramyxovirus, simian virus 5. Virology. 1984 Oct 30;138(2):310–323. doi: 10.1016/0042-6822(84)90354-4. [DOI] [PubMed] [Google Scholar]
  28. Pipas J. M., Peden K. W., Nathans D. Mutational analysis of simian virus 40 T antigen: isolation and characterization of mutants with deletions in the T-antigen gene. Mol Cell Biol. 1983 Feb;3(2):203–213. doi: 10.1128/mcb.3.2.203. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Richardson C. D., Berkovich A., Rozenblatt S., Bellini W. J. Use of antibodies directed against synthetic peptides for identifying cDNA clones, establishing reading frames, and deducing the gene order of measles virus. J Virol. 1985 Apr;54(1):186–193. doi: 10.1128/jvi.54.1.186-193.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sabban E., Marchesi V., Adesnik M., Sabatini D. D. Erythrocyte membrane protein band 3: its biosynthesis and incorporation into membranes. J Cell Biol. 1981 Dec;91(3 Pt 1):637–646. doi: 10.1083/jcb.91.3.637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Virtanen I., Ekblom P., Laurila P. Subcellular compartmentalization of saccharide moieties in cultured normal and malignant cells. J Cell Biol. 1980 May;85(2):429–434. doi: 10.1083/jcb.85.2.429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Walter P., Blobel G. Signal recognition particle contains a 7S RNA essential for protein translocation across the endoplasmic reticulum. Nature. 1982 Oct 21;299(5885):691–698. doi: 10.1038/299691a0. [DOI] [PubMed] [Google Scholar]
  33. Walter P., Blobel G. Translocation of proteins across the endoplasmic reticulum III. Signal recognition protein (SRP) causes signal sequence-dependent and site-specific arrest of chain elongation that is released by microsomal membranes. J Cell Biol. 1981 Nov;91(2 Pt 1):557–561. doi: 10.1083/jcb.91.2.557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Winter G., Fields S. Cloning of influenza cDNA ino M13: the sequence of the RNA segment encoding the A/PR/8/34 matrix protein. Nucleic Acids Res. 1980 May 10;8(9):1965–1974. doi: 10.1093/nar/8.9.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]

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