Abstract
Heterologous expression of the human T-cell lymphotropic virus type 1 (HTLV-1) envelope surface glycoprotein (gp46) in a vaccinia virus/T7 polymerase system resulted in the production of authentic recombinant gp46. Five differentially glycosylated forms of the surface envelope protein were produced by this mammalian system, as demonstrated by tunicamycin inhibition of N-glycosylation and N-glycan removal with endoglycosidase H and glycopeptidase F. These studies revealed that all four potential N-glycosylation sites in gp46 were used for oligosaccharide modification and that the oligosaccharides were mannose-rich and/or hybrid in composition. Conformational integrity of the recombinant HTLV-1 envelope protein was determined by the ability to bind to various HTLV-1-infected human sera and a panel of conformational-dependent human monoclonal antibodies under nondenaturing conditions. Furthermore, this recombinant gp46 was recognized by a series of HTLV-2-infected human sera and sera from a Pan paniscus chimpanzee infected with the distantly related simian T-cell lymphotropic virus STLVpan-p. Maintenance of highly conserved conformational epitopes in the recombinant HTLV-1 envelope protein structure suggests that it may serve as a useful diagnostic reagent and an effective vaccine candidate.
Full Text
The Full Text of this article is available as a PDF (470.4 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Arp J., Ford C. M., Palker T. J., King E. E., Dekaban G. A. Expression and immunogenicity of the entire human T cell leukaemia virus type I envelope protein produced in a baculovirus system. J Gen Virol. 1993 Feb;74(Pt 2):211–222. doi: 10.1099/0022-1317-74-2-211. [DOI] [PubMed] [Google Scholar]
- Bruck C., Rensonnet N., Portetelle D., Cleuter Y., Mammerickx M., Burny A., Mamoun R., Guillemain B., van der Maaten M. J., Ghysdael J. Biologically active epitopes of bovine leukemia virus glycoprotein gp51: their dependence on protein glycosylation and genetic variability. Virology. 1984 Jul 15;136(1):20–31. doi: 10.1016/0042-6822(84)90244-7. [DOI] [PubMed] [Google Scholar]
- Buckner C., Roberts C. R., Foung S. K., Lipka J., Reyes G. R., Hadlock K., Chan L., Gongora-Biachi R. A., Hjelle B., Lal R. B. Immune responsiveness to the immunodominant recombinant envelope epitopes of human T lymphotropic virus types I and II in diverse geographic populations. J Infect Dis. 1992 Nov;166(5):1160–1163. doi: 10.1093/infdis/166.5.1160. [DOI] [PubMed] [Google Scholar]
- Chen Y. M., Lee T. H., Samuel K. P., Okayama A., Tachibana N., Miyoshi I., Papas T. S., Essex M. Antibody reactivity to different regions of human T-cell leukemia virus type 1 gp61 in infected people. J Virol. 1989 Nov;63(11):4952–4957. doi: 10.1128/jvi.63.11.4952-4957.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- DALES S., SIMINOVITCH L. The development of vaccinia virus in Earle's L strain cells as examined by electron microscopy. J Biophys Biochem Cytol. 1961 Aug;10:475–503. doi: 10.1083/jcb.10.4.475. [DOI] [PMC free article] [PubMed] [Google Scholar]
- De B. K., Lairmore M. D., Griffis K., Williams L. J., Villinger F., Quinn T. C., Brown C., Nzilambi, Sugimoto M., Araki S. Comparative analysis of nucleotide sequences of the partial envelope gene (5' domain) among human T lymphotropic virus type I (HTLV-I) isolates. Virology. 1991 May;182(1):413–419. doi: 10.1016/0042-6822(91)90692-5. [DOI] [PubMed] [Google Scholar]
- Edouard E., Legrand E., Astier-Gin T., Dalibart R., Geoffre S., Dalbon P., Guillemain B., Londos-Gagliardi D. Characterization of monoclonal antibodies directed against the gp46 of human T-cell leukemia virus type I. Leukemia. 1994 Apr;8 (Suppl 1):S60–S64. [PubMed] [Google Scholar]
- Elroy-Stein O., Fuerst T. R., Moss B. Cap-independent translation of mRNA conferred by encephalomyocarditis virus 5' sequence improves the performance of the vaccinia virus/bacteriophage T7 hybrid expression system. Proc Natl Acad Sci U S A. 1989 Aug;86(16):6126–6130. doi: 10.1073/pnas.86.16.6126. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fenouillet E., Gluckman J. C. Effect of a glucosidase inhibitor on the bioactivity and immunoreactivity of human immunodeficiency virus type 1 envelope glycoprotein. J Gen Virol. 1991 Aug;72(Pt 8):1919–1926. doi: 10.1099/0022-1317-72-8-1919. [DOI] [PubMed] [Google Scholar]
- Ford C. M., Arp J., Palker T. J., King E. E., Dekaban G. A. Characterization of the antibody response to three different versions of the HTLV-I envelope protein expressed by recombinant vaccinia viruses: induction of neutralizing antibody. Virology. 1992 Nov;191(1):448–453. doi: 10.1016/0042-6822(92)90208-7. [DOI] [PubMed] [Google Scholar]
- Foung S. K., Perkins S. Electric field-induced cell fusion and human monoclonal antibodies. J Immunol Methods. 1989 Jan 6;116(1):117–122. doi: 10.1016/0022-1759(89)90319-0. [DOI] [PubMed] [Google Scholar]
- Fuerst T. R., Earl P. L., Moss B. Use of a hybrid vaccinia virus-T7 RNA polymerase system for expression of target genes. Mol Cell Biol. 1987 Jul;7(7):2538–2544. doi: 10.1128/mcb.7.7.2538. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Geyer H., Holschbach C., Hunsmann G., Schneider J. Carbohydrates of human immunodeficiency virus. Structures of oligosaccharides linked to the envelope glycoprotein 120. J Biol Chem. 1988 Aug 25;263(24):11760–11767. [PubMed] [Google Scholar]
- Giri A., Markham P., Digilio L., Hurteau G., Gallo R. C., Franchini G. Isolation of a novel simian T-cell lymphotropic virus from Pan paniscus that is distantly related to the human T-cell leukemia/lymphotropic virus types I and II. J Virol. 1994 Dec;68(12):8392–8395. doi: 10.1128/jvi.68.12.8392-8395.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goubau P., Liu H. F., De Lange G. G., Vandamme A. M., Desmyter J. HTLV-II seroprevalence in pygmies across Africa since 1970. AIDS Res Hum Retroviruses. 1993 Aug;9(8):709–713. doi: 10.1089/aid.1993.9.709. [DOI] [PubMed] [Google Scholar]
- Hadlock K. G., Goh C. J., Bradshaw P. A., Perkins S., Lo J., Kaplan J. E., Khabbaz R., Foung S. K. Delineation of an immunodominant and human T-cell lymphotropic virus (HTLV)-specific epitope within the HTLV-I transmembrane glycoprotein. Blood. 1995 Aug 15;86(4):1392–1399. [PubMed] [Google Scholar]
- Haigwood N. L., Nara P. L., Brooks E., Van Nest G. A., Ott G., Higgins K. W., Dunlop N., Scandella C. J., Eichberg J. W., Steimer K. S. Native but not denatured recombinant human immunodeficiency virus type 1 gp120 generates broad-spectrum neutralizing antibodies in baboons. J Virol. 1992 Jan;66(1):172–182. doi: 10.1128/jvi.66.1.172-182.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ho D. D., Fung M. S., Cao Y. Z., Li X. L., Sun C., Chang T. W., Sun N. C. Another discontinuous epitope on glycoprotein gp120 that is important in human immunodeficiency virus type 1 neutralization is identified by a monoclonal antibody. Proc Natl Acad Sci U S A. 1991 Oct 15;88(20):8949–8952. doi: 10.1073/pnas.88.20.8949. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ho D. D., McKeating J. A., Li X. L., Moudgil T., Daar E. S., Sun N. C., Robinson J. E. Conformational epitope on gp120 important in CD4 binding and human immunodeficiency virus type 1 neutralization identified by a human monoclonal antibody. J Virol. 1991 Jan;65(1):489–493. doi: 10.1128/jvi.65.1.489-493.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ijichi S., Matsuda T., Maruyama I., Izumihara T., Kojima K., Niimura T., Maruyama Y., Sonoda S., Yoshida A., Osame M. Arthritis in a human T lymphotropic virus type I (HTLV-I) carrier. Ann Rheum Dis. 1990 Sep;49(9):718–721. doi: 10.1136/ard.49.9.718. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kaluza G., Rott R., Schwarz R. T. Carbohydrate-induced conformational changes of Semliki forest virus glycoproteins determine antigenicity. Virology. 1980 Apr 30;102(2):286–299. doi: 10.1016/0042-6822(80)90096-3. [DOI] [PubMed] [Google Scholar]
- Kang C. Y., Nara P., Chamat S., Caralli V., Ryskamp T., Haigwood N., Newman R., Köhler H. Evidence for non-V3-specific neutralizing antibodies that interfere with gp120/CD4 binding in human immunodeficiency virus 1-infected humans. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6171–6175. doi: 10.1073/pnas.88.14.6171. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kinoshita T., Tsujimoto A., Shimotohno K. Sequence variations in LTR and env regions of HTLV-I do not discriminate between the virus from patients with HTLV-I-associated myelopathy and adult T-cell leukemia. Int J Cancer. 1991 Feb 20;47(4):491–495. doi: 10.1002/ijc.2910470403. [DOI] [PubMed] [Google Scholar]
- Kiyokawa T., Yoshikura H., Hattori S., Seiki M., Yoshida M. Envelope proteins of human T-cell leukemia virus: expression in Escherichia coli and its application to studies of env gene functions. Proc Natl Acad Sci U S A. 1984 Oct;81(19):6202–6206. doi: 10.1073/pnas.81.19.6202. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Komurian F., Pelloquin F., de Thé G. In vivo genomic variability of human T-cell leukemia virus type I depends more upon geography than upon pathologies. J Virol. 1991 Jul;65(7):3770–3778. doi: 10.1128/jvi.65.7.3770-3778.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kornfeld R., Kornfeld S. Assembly of asparagine-linked oligosaccharides. Annu Rev Biochem. 1985;54:631–664. doi: 10.1146/annurev.bi.54.070185.003215. [DOI] [PubMed] [Google Scholar]
- Kuga T., Hattori S., Yoshida M., Taniguchi T. Expression of human T-cell leukemia virus type I envelope protein in Saccharomyces cerevisiae. Gene. 1986;44(2-3):337–340. doi: 10.1016/0378-1119(86)90199-x. [DOI] [PubMed] [Google Scholar]
- LaGrenade L., Hanchard B., Fletcher V., Cranston B., Blattner W. Infective dermatitis of Jamaican children: a marker for HTLV-I infection. Lancet. 1990 Dec 1;336(8727):1345–1347. doi: 10.1016/0140-6736(90)92896-p. [DOI] [PubMed] [Google Scholar]
- Lee T. H., Coligan J. E., Homma T., McLane M. F., Tachibana N., Essex M. Human T-cell leukemia virus-associated membrane antigens: identity of the major antigens recognized after virus infection. Proc Natl Acad Sci U S A. 1984 Jun;81(12):3856–3860. doi: 10.1073/pnas.81.12.3856. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Li Y., Luo L., Rasool N., Kang C. Y. Glycosylation is necessary for the correct folding of human immunodeficiency virus gp120 in CD4 binding. J Virol. 1993 Jan;67(1):584–588. doi: 10.1128/jvi.67.1.584-588.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lipka J. J., Bui K., Reyes G. R., Moeckli R., Wiktor S. Z., Blattner W. A., Murphy E. L., Shaw G. M., Hanson C. V., Sninsky J. J. Determination of a unique and immunodominant epitope of human T cell lymphotropic virus type I. J Infect Dis. 1990 Aug;162(2):353–357. doi: 10.1093/infdis/162.2.353. [DOI] [PubMed] [Google Scholar]
- Mackett M., Smith G. L., Moss B. General method for production and selection of infectious vaccinia virus recombinants expressing foreign genes. J Virol. 1984 Mar;49(3):857–864. doi: 10.1128/jvi.49.3.857-864.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Manca F., Fenoglio D., Valle M. T., Li Pira G., Kunkl A., Ferraris A., Saverino D., Lancia F., Mortara L., Lozzi L. Human CD4+ T cells can discriminate the molecular and structural context of T epitopes of HIV gp120 and HIV p66. J Acquir Immune Defic Syndr Hum Retrovirol. 1995 Jul 1;9(3):227–237. [PubMed] [Google Scholar]
- Manca F., Li Pira G., Fenoglio D., Valle M. T., Kunkl A., Ferraris A., Lancia F., Saverino D., Mortara L., Balderas R. Recognition of human T-leukemia virus (HTLV-1) envelope by human CD4+ T-cell lines from HTLV-1 seronegative individuals: specificity and clonal heterogeneity. Blood. 1995 Mar 15;85(6):1547–1554. [PubMed] [Google Scholar]
- Matsushita S., Robert-Guroff M., Trepel J., Cossman J., Mitsuya H., Broder S. Human monoclonal antibody directed against an envelope glycoprotein of human T-cell leukemia virus type I. Proc Natl Acad Sci U S A. 1986 Apr;83(8):2672–2676. doi: 10.1073/pnas.83.8.2672. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Misumi Y., Misumi Y., Miki K., Takatsuki A., Tamura G., Ikehara Y. Novel blockade by brefeldin A of intracellular transport of secretory proteins in cultured rat hepatocytes. J Biol Chem. 1986 Aug 25;261(24):11398–11403. [PubMed] [Google Scholar]
- Mochizuki M., Watanabe T., Yamaguchi K., Takatsuki K., Yoshimura K., Shirao M., Nakashima S., Mori S., Araki S., Miyata N. HTLV-I uveitis: a distinct clinical entity caused by HTLV-I. Jpn J Cancer Res. 1992 Mar;83(3):236–239. doi: 10.1111/j.1349-7006.1992.tb00092.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Moore J. P., McKeating J. A., Jones I. M., Stephens P. E., Clements G., Thomson S., Weiss R. A. Characterization of recombinant gp120 and gp160 from HIV-1: binding to monoclonal antibodies and soluble CD4. AIDS. 1990 Apr;4(4):307–315. doi: 10.1097/00002030-199004000-00004. [DOI] [PubMed] [Google Scholar]
- Nyunoya H., Ogura T., Kikuchi M., Iwamoto H., Yamashita K., Maekawa M., Takebe Y., Miyamura K., Yamazaki S., Shimotohno K. Expression of HTLV-I envelope protein fused to hydrophobic amino-terminal peptide of baculovirus polyhedrin in insect cells and its application for serological assays. AIDS Res Hum Retroviruses. 1990 Nov;6(11):1311–1321. doi: 10.1089/aid.1990.6.1311. [DOI] [PubMed] [Google Scholar]
- Osame M., Usuku K., Izumo S., Ijichi N., Amitani H., Igata A., Matsumoto M., Tara M. HTLV-I associated myelopathy, a new clinical entity. Lancet. 1986 May 3;1(8488):1031–1032. doi: 10.1016/s0140-6736(86)91298-5. [DOI] [PubMed] [Google Scholar]
- Palker T. J., Scearce R. M., Miller S. E., Popovic M., Bolognesi D. P., Gallo R. C., Haynes B. F. Monoclonal antibodies against human T cell leukemia-lymphoma virus (HTLV) p24 internal core protein. Use as diagnostic probes and cellular localization of HTLV. J Exp Med. 1984 Apr 1;159(4):1117–1131. doi: 10.1084/jem.159.4.1117. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Palker T. J., Tanner M. E., Scearce R. M., Streilein R. D., Clark M. E., Haynes B. F. Mapping of immunogenic regions of human T cell leukemia virus type I (HTLV-I) gp46 and gp21 envelope glycoproteins with env-encoded synthetic peptides and a monoclonal antibody to gp46. J Immunol. 1989 Feb 1;142(3):971–978. [PubMed] [Google Scholar]
- Perkins S., Foung S. K. Stabilizing antibody secretion of human Epstein Barr virus-activated B-lymphocytes with hybridoma formation by electrofusion. Methods Mol Biol. 1995;48:295–307. doi: 10.1385/0-89603-304-X:295. [DOI] [PubMed] [Google Scholar]
- Perkins S., Zimmermann U., Foung S. K. Parameters to enhance human hybridoma formation with hypoosmolar electrofusion. Hum Antibodies Hybridomas. 1991 Jul;2(3):155–159. [PubMed] [Google Scholar]
- Pierotti M., DeLeo A. B., Pinter A., O'Donnell P. V., Hämmerling U., Fleissner E. The GIX antigen of murine leukemia virus: an analysis with monoclonal antibodies. Virology. 1981 Jul 30;112(2):450–460. doi: 10.1016/0042-6822(81)90292-0. [DOI] [PubMed] [Google Scholar]
- Pique C., Pham D., Tursz T., Dokhélar M. C. Human T-cell leukemia virus type I envelope protein maturation process: requirements for syncytium formation. J Virol. 1992 Feb;66(2):906–913. doi: 10.1128/jvi.66.2.906-913.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pique C., Tursz T., Dokhelar M. C. Mutations introduced along the HTLV-I envelope gene result in a non-functional protein: a basis for envelope conservation? EMBO J. 1990 Dec;9(13):4243–4248. doi: 10.1002/j.1460-2075.1990.tb07872.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Plummer T. H., Jr, Elder J. H., Alexander S., Phelan A. W., Tarentino A. L. Demonstration of peptide:N-glycosidase F activity in endo-beta-N-acetylglucosaminidase F preparations. J Biol Chem. 1984 Sep 10;259(17):10700–10704. [PubMed] [Google Scholar]
- Rademacher T. W., Parekh R. B., Dwek R. A. Glycobiology. Annu Rev Biochem. 1988;57:785–838. doi: 10.1146/annurev.bi.57.070188.004033. [DOI] [PubMed] [Google Scholar]
- Rademacher T. W. Therapeutic challenges: does Glycobiology have a role? Trends Biotechnol. 1992 Jul;10(7):227–230. doi: 10.1016/0167-7799(92)90228-n. [DOI] [PubMed] [Google Scholar]
- Ratner L., Josephs S. F., Starcich B., Hahn B., Shaw G. M., Gallo R. C., Wong-Staal F. Nucleotide sequence analysis of a variant human T-cell leukemia virus (HTLV-Ib) provirus with a deletion in pX-I. J Virol. 1985 Jun;54(3):781–790. doi: 10.1128/jvi.54.3.781-790.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Robert-Guroff M., Nakao Y., Notake K., Ito Y., Sliski A., Gallo R. C. Natural antibodies to human retrovirus HTLV in a cluster of Japanese patients with adult T cell leukemia. Science. 1982 Feb 19;215(4535):975–978. doi: 10.1126/science.6760397. [DOI] [PubMed] [Google Scholar]
- Samuel K. P., Lautenberger J. A., Jorcyk C. L., Josephs S., Wong-Staal F., Papas T. S. Diagnostic potential for human malignancies of bacterially produced HTLV-I envelope protein. Science. 1984 Nov 30;226(4678):1094–1097. doi: 10.1126/science.6208612. [DOI] [PubMed] [Google Scholar]
- Sattentau Q. J., Moore J. P. Human immunodeficiency virus type 1 neutralization is determined by epitope exposure on the gp120 oligomer. J Exp Med. 1995 Jul 1;182(1):185–196. doi: 10.1084/jem.182.1.185. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schwarz R. T., Datema R. Inhibition of the dolichol pathway of protein glycosylation. Methods Enzymol. 1982;83:432–443. doi: 10.1016/0076-6879(82)83041-3. [DOI] [PubMed] [Google Scholar]
- Seiki M., Hattori S., Hirayama Y., Yoshida M. Human adult T-cell leukemia virus: complete nucleotide sequence of the provirus genome integrated in leukemia cell DNA. Proc Natl Acad Sci U S A. 1983 Jun;80(12):3618–3622. doi: 10.1073/pnas.80.12.3618. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sodora D. L., Cohen G. H., Muggeridge M. I., Eisenberg R. J. Absence of asparagine-linked oligosaccharides from glycoprotein D of herpes simplex virus type 1 results in a structurally altered but biologically active protein. J Virol. 1991 Aug;65(8):4424–4431. doi: 10.1128/jvi.65.8.4424-4431.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stamatatos L., Cheng-Mayer C. Structural modulations of the envelope gp120 glycoprotein of human immunodeficiency virus type 1 upon oligomerization and differential V3 loop epitope exposure of isolates displaying distinct tropism upon virion-soluble receptor binding. J Virol. 1995 Oct;69(10):6191–6198. doi: 10.1128/jvi.69.10.6191-6198.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sugawara K., Kitame F., Nishimura H., Nakamura K. Operational and topological analyses of antigenic sites on influenza C virus glycoprotein and their dependence on glycosylation. J Gen Virol. 1988 Mar;69(Pt 3):537–547. doi: 10.1099/0022-1317-69-3-537. [DOI] [PubMed] [Google Scholar]
- Trimble R. B., Maley F. Optimizing hydrolysis of N-linked high-mannose oligosaccharides by endo-beta-N-acetylglucosaminidase H. Anal Biochem. 1984 Sep;141(2):515–522. doi: 10.1016/0003-2697(84)90080-0. [DOI] [PubMed] [Google Scholar]
- Varki A. Biological roles of oligosaccharides: all of the theories are correct. Glycobiology. 1993 Apr;3(2):97–130. doi: 10.1093/glycob/3.2.97. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Vidal S., Mottet G., Kolakofsky D., Roux L. Addition of high-mannose sugars must precede disulfide bond formation for proper folding of Sendai virus glycoproteins. J Virol. 1989 Feb;63(2):892–900. doi: 10.1128/jvi.63.2.892-900.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Vile R. G., Schulz T. F., Danos O. F., Collins M. K., Weiss R. A. A murine cell line producing HTLV-I pseudotype virions carrying a selectable marker gene. Virology. 1991 Jan;180(1):420–424. doi: 10.1016/0042-6822(91)90050-l. [DOI] [PubMed] [Google Scholar]
- Yoshida M., Seiki M., Yamaguchi K., Takatsuki K. Monoclonal integration of human T-cell leukemia provirus in all primary tumors of adult T-cell leukemia suggests causative role of human T-cell leukemia virus in the disease. Proc Natl Acad Sci U S A. 1984 Apr;81(8):2534–2537. doi: 10.1073/pnas.81.8.2534. [DOI] [PMC free article] [PubMed] [Google Scholar]
- de Thé G., Bomford R. An HTLV-I vaccine: why, how, for whom? AIDS Res Hum Retroviruses. 1993 May;9(5):381–386. doi: 10.1089/aid.1993.9.381. [DOI] [PubMed] [Google Scholar]
- van Drunen Littel-van den Hurk S., Hughes G., Babiuk L. A. The role of carbohydrate in the antigenic and immunogenic structure of bovine herpesvirus type 1 glycoproteins gI and gIV. J Gen Virol. 1990 Sep;71(Pt 9):2053–2063. doi: 10.1099/0022-1317-71-9-2053. [DOI] [PubMed] [Google Scholar]