Structure-Function Analysis of Mouse Interferon Alpha Species: MuIFN-α10, a Subspecies with Low Antiviral Activity Free

Abstract

Summary

A mouse interferon alpha gene (MuIFN-α10) was isolated from a BALB/c cosmid genomic library. The gene was located on a 1.8 kb dIII fragment and a 5.1 kb RI fragment. The coding region and parts of the 5′ and 3′ non-coding regions were sequenced. The results showed that the MuIFN-α10 gene encoded a protein of 167 amino acids. Like most other MuIFN-α species it contained a putative -glycosylation site at amino acid positions 78 to 80. It also possessed cysteine residues at positions 1, 29, 86, 99 and 129. In the signal peptide, in addition to cysteine 21, which is present in all MuIFN-α species sequenced so far, a cysteine was found at position 22. At the amino acid level MuIFN-α10 showed strong homology to MuIFN-α1 (only 15 out of 167 amino acids were different). The MuIFN-α10 gene was transiently expressed in monkey COS cells under the direction of the simian virus 40 early promoter. The protein product secreted by COS cells was equally active on mouse (L929) and hamster (CHO) cells. However, as compared to MuIFN-α1 and MuIFN-α4 the specific activity on mouse cells of the protein was 10- to 100-fold lower. To find out which region of its structure was responsible for this low activity, hybrids of the genes encoding MuIFN-α10 and MuIFN-α1 were constructed using the two common I sites which correspond to positions between amino acids 67 and 68 and 123 and 124, respectively. The data showed that hybrid constructs which were MuIFN-α1-like from amino acid 68 or MuIFN-α10-like from position 124 to the C terminus possessed high antiviral activity. Other hybrid constructs were hardly active at all. This implied that the amino acid 68 to 123 region was mainly responsible for the low antiviral activity of MuIFN-α10. In this part of the molecule MuIFN-α1 and MuIFN-α10 differed in only five amino acids. A serine at position 110 and a valine at 85 were unique to MuIFN-α10 as compared to all known MuIFN-α and human IFN-α subspecies.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-69-1-67
1988-01-01
2024-03-29
Loading full text...

Full text loading...

/deliver/fulltext/jgv/69/1/JV0690010067.html?itemId=/content/journal/jgv/10.1099/0022-1317-69-1-67&mimeType=html&fmt=ahah

References

  1. DANDOY F., KELLEY K. A., DEMAEYER-GUIGNARD J., DEMAEYER E., PITHA P. M. 1984; Linkage analysis of the murine interferon-a locus on chromosome 4. Journal of Experimental Medicine 160:294–302
    [Google Scholar]
  2. DANDOY F., DEMAEYER E., BONHOMME F., GUENET J. L., DEMAEYER-GUIGNARD J. 1985; Segregation of restriction fragment length polymorphism in an interspecies cross of laboratory and wild mice indicates a tight linkage of the murine IFN-β gene to the murine IFN-α genes. Journal of Virology 56:216–220
    [Google Scholar]
  3. DAUGHERTY B., MARTIN-ZANCA D., KELDER B., COLLIER K., SEAMANS T. C, HOTTA K., PESTKA S. 1984; Isolation and bacterial expression of a murine alpha leucocyte interferon gene. Journal of Interferon Research 4:635–643
    [Google Scholar]
  4. KELLEY K. A., PITHA P. M. 1985a; Characterization of a mouse interferon a gene locus. II. Differential expression of α-interferon genes. Nucleic Acids Research 13:825–839
    [Google Scholar]
  5. KELLEY K. A., PITHA P. M. 1985b; Characterization of a mouse interferon gene locus. I. Isolation of a cluster of four interferon genes. Nucleic Acids Research 13:805–823
    [Google Scholar]
  6. KELLEY K. A., KOZAK C. A., DANDOY F., SOR F., SKUP D., WINDASS J. D., DEMAEYER-GUIGNARD J., PITHA P. M., DEMAEYER E. 1983; Mapping of murine interferon-a genes to chromosome 4. Gene 26:181–188
    [Google Scholar]
  7. KELLEY K. A., RAY N. B. K., KELLUM M., PITHA P. M. 1986; Synthesis of fusion and mature murine alpha interferons in Escherichia coli. Gene 45:317–325
    [Google Scholar]
  8. LEROSCOUET D., VODJDANI G., LEMAIGZE-DUBREUIL Y., TOVEY M. G., LATTA M., DOLY J. 1985; Structure of a murine alpha interferon pseudogene with a repetitive R-type sequence in the 3′-flanking region. Molecular and Cellular Biology 5:1343–1348
    [Google Scholar]
  9. LOVETT M., COX D. R., YEE D., BOLL W., WEISSMANN C, EPSTEIN C. J., EPSTEIN L. B. 1984; The chromosomal location of mouse interferon a genes. EMBO Journal 3:1643–1646
    [Google Scholar]
  10. MANIATIS T., FRITSCH E. F., SAMBROOK J. 1982 Molecular Cloning: A Laboratory Manual New York: Cold Spring Harbor Laboratory;
    [Google Scholar]
  11. MESSING J., CREA R., SEEBURG P. H. 1981; A system for shotgun DNA sequencing. Nucleic Acids Research 9:309–321
    [Google Scholar]
  12. SANGER F., NICKLEN S., COULSON R. 1977; DNA sequencing with chain-terminating inhibitors. Proceedings of the National Academy of Sciences, U.S.A 74:5463–5467
    [Google Scholar]
  13. SEIF I., DEMAEYER-GUIGNARD J. 1986; Structure and expression of a new murine interferon-alpha gene: MuIFN-α9. Gene 43:111–121
    [Google Scholar]
  14. SHAW G. D., BOLL W., TAIRA H., MANTEL N., LENGYEL P., WEISSMANN C. 1983; Structure and expression of cloned murine IFN-α genes. Nucleic Acids Research 11:555–573
    [Google Scholar]
  15. TRAPMAN J., VAN HEUVEL M., ZWARTHOFF E. C, BOSVELD I. J., KLAASSEN P. 1986; Properties of modified mouse interferon alpha species. Journal of Cellular Biochemistry supplement 10c:246
    [Google Scholar]
  16. VAN DER KORPUT A. G. M., HILKENS J., KROEZEN V., ZWARTHOFF E. C., TRAPMAN J. 1985; Mouse interferon alpha and beta genes are linked at the centromere proximal region of chromosome 4. Journal of General Virology 66:493–502
    [Google Scholar]
  17. VAN HEUVEL M., BOSVELD I. J., MOOREN A. T. A., TRAPMAN J., ZWARTHOFF E. C. 1986; Properties of natural and hybrid murine alpha interferons. Journal of General Virology 67:2215–2222
    [Google Scholar]
  18. VAN HEUVEL M., BOSVELD I. J., KLAASSEN P., ZWARTHOFF E. C., TRAPMAN J. 1987; Structure-function analysis of murine interferon-alpha: antiviral properties of novel hybrid interferons. Journal of Interferon Research in press
    [Google Scholar]
  19. WEISSMANN C., WEBER H. 1986 The interferon genes. Progress in Nucleic Acid Research and Molecular Biology 33251–300 Edited by Cohn W. E., Moldave K. New York: Academic Press;
    [Google Scholar]
  20. ZWARTHOFF E. C, MOOREN A. T. A., TRAPMAN J. 1985; Organization, structure and expression of murine interferon alpha genes. Nucleic Acids Research 13:791–804
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/0022-1317-69-1-67
Loading
/content/journal/jgv/10.1099/0022-1317-69-1-67
Loading

Data & Media loading...

Most cited Most Cited RSS feed