1887

Abstract

The human cytomegalovirus (HCMV) UL37 exon 3 (UL37x3) open reading frame (ORF) encodes the carboxyl termini of two immediate-early glycoproteins (gpUL37 and gpUL37). UL37x3 homologous sequences are not required for mouse cytomegalovirus (MCMV) growth ; yet, they are important for MCMV growth and pathogenesis Similarly, UL37x3 sequences are dispensable for HCMV growth in culture, but their requirement for HCMV growth is not known. To determine this requirement, we directly sequenced the complete UL37x3 gene in multiple HCMV primary strains. A total of 63 of the 310 amino acids in the UL37x3 ORF differ non-conservatively in one or more HCMV primary strains. The HCMV UL37x3 genetic diversity is non-random: the -glycosylation (46/186 aa) and basic (9/15 aa) domains have the highest proportion of non-conservative variant amino acids. Nonetheless, most (15/17 signals) of the glycosylation signals are retained in all HCMV primary strains. Moreover, new -glycosylation signals are encoded by 5/20 primary strains. In sharp contrast, the UL37x3 transmembrane (TM) ORF completely lacks diversity in all 20 HCMV sequenced primary strains, and only 1 of 28 cytosolic tail residues differs non-conservatively. To test the functional significance of the conserved carboxyl terminus, gpUL37 mutants lacking the TM and/or cytosolic tail were tested for transactivating activity. The gpUL37 carboxyl-terminal mutants are partially defective in promoter transactivation even though they trafficked similarly to the wild-type protein into the endoplasmic reticulum and to mitochondria. From these results, we conclude that -glycosylated gpUL37, particularly its TM and cytosolic domains, is important for HCMV growth in humans.

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2001-07-01
2019-10-22
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References

  1. Al-Barazi, H. O. & Colberg-Poley, A. M. ( 1996; ). The human cytomegalovirus UL37 immediate-early regulatory protein is an integral membrane N-glycoprotein which traffics through the endoplasmic reticulum and Golgi apparatus. Journal of Virology 70, 7198-7208.
    [Google Scholar]
  2. Baer, R., Bankier, A. T., Biggin, M. D., Deininger, P. L., Farrell, P. J., Gibson, T. J., Hatfull, G., Hudson, G. S., Satchwell, S. C., Seguin, C., Tuffnell, P. S. & Barrell, B. G. ( 1984; ). DNA sequence and expression of the B95-8 Epstein–Barr virus genome. Nature 310, 207-211.[CrossRef]
    [Google Scholar]
  3. Biegalke, B. J. ( 1999; ). Human cytomegalovirus US3 gene expression is regulated by a complex network of positive and negative regulators. Virology 261, 155-164.[CrossRef]
    [Google Scholar]
  4. Borst, E.-M., Hahn, G., Koszinowski, U. H. & Messerle, M. ( 1999; ). Cloning of the human cytomegalovirus (HCMV) genome as an infectious bacterial artificial chromosome in Escherichia coli: a new approach for construction of HCMV mutants. Journal of Virology 73, 8320-8329.
    [Google Scholar]
  5. Britt, W. J. & Alford, C. A. ( 1996; ). Cytomegalovirus. In Fields Virology , pp. 2493-2523. Edited by B. N. Fields, D. M. Knipe & P. M. Howley. Philadelphia:Lippincott–Raven.
  6. Cha, T.-A., Tom, E., Kemble, G. W., Duke, G. M., Mocarski, E. S. & Spaete, R. R. ( 1996; ). Human cytomegalovirus clinical isolates carry at least 19 genes not found in laboratory strains. Journal of Virology 70, 78-83.
    [Google Scholar]
  7. Chee, M. S., Bankier, A. T., Beck, S., Bohni, R., Brown, C. M., Cerny, R., Horsnell, T., Hutchison, C. A.III, Kouzarides, T., Martignetti, J. A., Preddie, E., Satchwell, S. C., Tomlinson, P., Weston, K. M. & Barrell, B. G. ( 1990; ). Analysis of the protein-coding content of the sequence of human cytomegalovirus strain AD169. Current Topics in Microbiology and Immunology 154, 125-169.
    [Google Scholar]
  8. Chou, S. ( 1992; ). Effect of interstrain variation on diagnostic DNA amplification of the cytomegalovirus major immediate-early gene region. Journal of Clinical Microbiology 30, 2307-2310.
    [Google Scholar]
  9. Chou, S. & Dennison, K. M. ( 1991; ). Analysis of interstrain variation in cytomegalovirus glycoprotein B sequences encoding neutralization-related epitopes. Journal of Infectious Diseases 163, 1229-1234.[CrossRef]
    [Google Scholar]
  10. Colberg-Poley, A. M. ( 1996; ). Functional roles of immediate early proteins encoded by the human cytomegalovirus UL36–38, UL115–119, TRS1/IRS1 and US3 loci. Intervirology 39, 350-360.
    [Google Scholar]
  11. Colberg-Poley, A. M., Santomenna, L. D., Harlow, P. P., Benfield, P. A. & Tenney, D. J. ( 1992; ). The HCMV US3 and UL36–38 immediate early proteins regulate gene expression. Journal of Virology 66, 95-105.
    [Google Scholar]
  12. Colberg-Poley, A. M., Huang, L., Soltero, V. E., Iskenderian, A., Schumacher, R.-F. & Anders, D. G. ( 1998; ). The acidic domain of pUL37x1 and gpUL37 plays a key role in transactivation of HCMV DNA replication gene promoter constructions. Virology 246, 400-408.[CrossRef]
    [Google Scholar]
  13. Colberg-Poley, A. M., Patel, M. B., Erezo, D. P. P. & Slater, J. E. ( 2000a; ). Human cytomegalovirus UL37 immediate-early regulatory proteins traffic through the secretory apparatus and to mitochondria. Journal of General Virology 81, 1779-1789.
    [Google Scholar]
  14. Colberg-Poley, A. M., Soltero, V. E. & Schumacher, R. F. ( 2000b; ). Analysis of cytomegalovirus gene expression by transfection. In Methods in Molecular Medicine , pp. 67-77. Edited by J. Sinclair. Totowa, New Jersey:Humana Press.
  15. Davison, A. J. & Scott, J. E. ( 1986; ). The complete DNA sequence of varicella-zoster virus. Journal of General Virology 67, 1759-1816.[CrossRef]
    [Google Scholar]
  16. Eustice, D. C., Feldman, P. A., Colberg-Poley, A. M., Buckery, R. M. & Neubauer, R. H. ( 1991; ). A sensitive method for detection of β-galactosidase in transfected mammalian cells. BioTechniques 11, 739-743.
    [Google Scholar]
  17. Goldmacher, V. S., Bartle, L. M., Skaletskaya, A., Dionne, C. A., Kedersha, N. L., Vater, C. A., Han, J.-W., Lutz, R. J., Watanabe, S., McFarland, E. D., Kieff, E. D., Mocarski, E. S. & Chittenden, T. ( 1999; ). A cytomegalovirus-encoded mitochondria-localized inhibitor of apoptosis structurally unrelated to bcl-2. Proceedings of the National Academy of Sciences, USA 96, 12536-12541.[CrossRef]
    [Google Scholar]
  18. Gompels, U. A., Nicholas, J., Lawrence, G., Jones, M., Thomson, B. J., Martin, M. E. D., Efstathiou, S., Craxton, M. & Macaulay, H. A. ( 1995; ). The DNA sequence of human herpesvirus 6: structure, coding content, and genome evolution. Virology 209, 29-51.[CrossRef]
    [Google Scholar]
  19. Hayajneh, W. A., Colberg-Poley, A. M., Skaletskaya, A., Bartle, L. M., Lesperance, M. M., Contopoulos-Ioannidis, D. G., Kedersha, N. L. & Goldmacher, V. S. ( 2001; ). Sequence and antiapoptotic functional domains of the human cytomegalovirus UL37 exon 1 immediate early protein are conserved in multiple primary strains. Virology 279, 233-240.[CrossRef]
    [Google Scholar]
  20. Kouzarides, T., Bankier, A. T., Satchwell, S. C., Preddy, E. & Barrell, B. G. ( 1988; ). An immediate early gene of human cytomegalovirus encodes a potential membrane glycoprotein. Virology 165, 151-164.[CrossRef]
    [Google Scholar]
  21. Lee, M., Xiao, J., Haghjoo, E., Zhan, X., Abenes, G., Tuong, T., Dunn, W. & Liu, F. ( 2000; ). Murine cytomegalovirus containing a mutation at open reading frame M37 is severely attenuated in growth and virulence in vivo. Journal of Virology 74, 11099-11107.[CrossRef]
    [Google Scholar]
  22. Lesperance, M. M., Contopoulos-Ioannidis, D. G., Gutiérrez, M. P. & Colberg-Poley, A. M. ( 1998; ). PCR detection of human cytomegalovirus DNA in clinical specimens using novel UL37 exon 3 and US3 primers. Clinical and Diagnostic Laboratory Immunology 5, 256-258.
    [Google Scholar]
  23. Lurain, N. S., Kapell, K. S., Huang, D. D., Short, J. A., Paintsil, J., Winkfield, E., Benedict, C. A., Ware, C. F. & Bremer, J. W. ( 1999; ). Human cytomegalovirus UL144 open reading frame: sequence hypervariability in low-passage clinical strains. Journal of Virology 73, 10040-10050.
    [Google Scholar]
  24. McGeoch, D. J. ( 1989; ). The genomes of the human herpesviruses: contents, relationships, and evolution. Annual Review of Microbiology 43, 235-265.[CrossRef]
    [Google Scholar]
  25. Meyer-König, U., Haberland, M., von Laer, D., Haller, O. & Hufert, F. T. ( 1998; ). Intragenic variability of human cytomegalovirus glycoprotein B in clinical strains. Journal of Infectious Diseases 177, 1162-1169.[CrossRef]
    [Google Scholar]
  26. Nicholas, J. ( 1996; ). Determination and analysis of the complete nucleotide sequence of human herpesvirus 7. Journal of Virology 70, 5975-5989.
    [Google Scholar]
  27. Nicholas, J. & Martin, M. E. D. ( 1994; ). Nucleotide sequence analysis of a 38·5-kilobase-pair region of the genome of human herpesvirus 6 encoding human cytomegalovirus immediate-early gene homologs and transactivating functions. Journal of Virology 68, 597-610.
    [Google Scholar]
  28. Pari, G. S., Field, A. K. & Smith, J. A. ( 1995; ). Potent antiviral activity of an antisense oligonucleotide complementary to the intron–exon boundary of human cytomegalovirus genes UL36 and UL37. Antimicrobial Agents and Chemotherapy 39, 1157-1161.[CrossRef]
    [Google Scholar]
  29. Rawlinson, W. D., Farrell, H. E. & Barrell, B. G. ( 1996; ). Analysis of the complete DNA sequence of murine cytomegalovirus. Journal of Virology 70, 8833-8849.
    [Google Scholar]
  30. Russo, J. J., Bohenzky, R. A., Chien, M.-C., Chen, J., Yan, M., Maddalena, S., Parry, J. P., Peruzzi, D., Edelman, I. S., Chang, Y. & Moore, P. S. ( 1996; ). Nucleotide sequence of the Kaposi sarcoma-associated herpesvirus (HHV8). Proceedings of the National Academy of Sciences, USA 93, 14862-14867.[CrossRef]
    [Google Scholar]
  31. Smith, J. A. & Pari, G. S. ( 1995; ). Expression of human cytomegalovirus UL36 and UL37 genes is required for viral DNA replication. Journal of Virology 69, 1925-1931.
    [Google Scholar]
  32. Tenney, D. J., Santomenna, L. D., Goudie, K. B. & Colberg-Poley, A. M. ( 1993; ). The human cytomegalovirus US3 immediate early protein lacking the putative transmembrane domain regulates gene expression. Nucleic Acids Research 21, 2931-2937.[CrossRef]
    [Google Scholar]
  33. Vink, C., Beuken, E. & Bruggeman, C. A. ( 2000; ). Complete DNA sequence of the rat cytomegalovirus genome. Journal of Virology 74, 7656-7665.[CrossRef]
    [Google Scholar]
  34. Wilkinson, G. W. G., Akrigg, A. & Greenaway, P. J. ( 1984; ). Transcription of the immediate early genes of human cytomegalovirus AD169. Virus Research 1, 101-116.[CrossRef]
    [Google Scholar]
  35. Zhang, H., Al-Barazi, H. O. & Colberg-Poley, A. M. ( 1996; ). The acidic domain of the human cytomegalovirus UL37 immediate early protein is dispensable for its transactivating activity and localization but is not for its synergism. Virology 223, 292-302.[CrossRef]
    [Google Scholar]
  36. Zweygberg Wirgart, B., Brytting, M., Linde, A., Wahren, B. & Grillner, L. ( 1998; ). Sequence variation within three important cytomegalovirus gene regions in isolates from four different patient populations. Journal of Clinical Microbiology 36, 3662-3669.
    [Google Scholar]
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