Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1997 Jan;71(1):595–600. doi: 10.1128/jvi.71.1.595-600.1997

Tyrosine phosphorylation events during coxsackievirus B3 replication.

M Huber 1, H C Selinka 1, R Kandolf 1
PMCID: PMC191089  PMID: 8985388

Abstract

In order to study cellular and viral determinants of pathogenicity, interactions between coxsackievirus B3 (CVB3) replication and cellular protein tyrosine phosphorylation were investigated. During CVB3 infection of HeLa cells, distinct proteins become phosphorylated on tyrosine residues, as detected by the use of antiphosphotyrosine Western blotting. Two proteins of 48 and 200 kDa showed enhanced tyrosine phosphorylation 4 to 5 h postinfection (p.i.), although virus-induced inhibition of cellular protein synthesis had already occurred 3 to 4 h p.i. Subcellular fractionation experiments revealed distinct localization of tyrosine-phosphorylated proteins of 48 and 200 kDa in the cytosol and membrane fractions of infected cells, respectively. In addition, in Vero cells infected with CVB3, echovirus (EV)11, or EV12, increased tyrosine phosphorylation of a 200-kDa protein was detected 6 h p.i. Herbimycin A, a specific inhibitor of Src-like protein tyrosine kinases, was shown to inhibit virus-induced tyrosine phosphorylations and to reduce the production of progeny virions. In contrast, in cells treated with the inhibitors staurosporine and calphostin C, the synthesis of progeny virions was not affected. Immunoprecipitation experiments suggested that the tyrosine-phosphorylated 200-kDa protein in CVB3-infected cells is of cellular origin. In summary, these investigations have begun to unravel the effect of CVB3 as well as EV11 and EV12 replication on cellular tyrosine phosphorylation and support the importance of tyrosine phosphorylation events for effective virus replication. Such cellular phosphorylation events triggered in the course of enterovirus infection may enhance virus replication.

Full Text

The Full Text of this article is available as a PDF (292.7 KB).

Selected References

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

  1. Barrett-Connor E. Is insulin-dependent diabetes mellitus caused by coxsackievirus B infection? A review of the epidemiologic evidence. Rev Infect Dis. 1985 Mar-Apr;7(2):207–215. doi: 10.1093/clinids/7.2.207. [DOI] [PubMed] [Google Scholar]
  2. Billah M. M., Anthes J. C. The regulation and cellular functions of phosphatidylcholine hydrolysis. Biochem J. 1990 Jul 15;269(2):281–291. doi: 10.1042/bj2690281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Borovjagin A. V., Ezrokhi M. V., Rostapshov V. M., Ugarova TYu, Bystrova T. F., Shatsky I. N. RNA--protein interactions within the internal translation initiation region of encephalomyocarditis virus RNA. Nucleic Acids Res. 1991 Sep 25;19(18):4999–5005. doi: 10.1093/nar/19.18.4999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bruns R. F., Miller F. D., Merriman R. L., Howbert J. J., Heath W. F., Kobayashi E., Takahashi I., Tamaoki T., Nakano H. Inhibition of protein kinase C by calphostin C is light-dependent. Biochem Biophys Res Commun. 1991 Apr 15;176(1):288–293. doi: 10.1016/0006-291x(91)90922-t. [DOI] [PubMed] [Google Scholar]
  5. Cheung R. K., Dosch H. M. The tyrosine kinase lck is critically involved in the growth transformation of human B lymphocytes. J Biol Chem. 1991 May 15;266(14):8667–8670. [PubMed] [Google Scholar]
  6. Cohen D. I., Tani Y., Tian H., Boone E., Samelson L. E., Lane H. C. Participation of tyrosine phosphorylation in the cytopathic effect of human immunodeficiency virus-1. Science. 1992 Apr 24;256(5056):542–545. doi: 10.1126/science.1570514. [DOI] [PubMed] [Google Scholar]
  7. Donaldson R. W., Hagedorn C. H., Cohen S. Epidermal growth factor or okadaic acid stimulates phosphorylation of eukaryotic initiation factor 4F. J Biol Chem. 1991 Feb 15;266(5):3162–3166. [PubMed] [Google Scholar]
  8. Filvaroff E., Stern D. F., Dotto G. P. Tyrosine phosphorylation is an early and specific event involved in primary keratinocyte differentiation. Mol Cell Biol. 1990 Mar;10(3):1164–1173. doi: 10.1128/mcb.10.3.1164. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fisher S. N., Kim Y. U., Shin M. L. Tyrosine kinase activation by Newcastle disease virus is required for TNF-alpha gene induction in astrocytes. J Immunol. 1994 Oct 1;153(7):3210–3217. [PubMed] [Google Scholar]
  10. Foulis A. K., Farquharson M. A., Cameron S. O., McGill M., Schönke H., Kandolf R. A search for the presence of the enteroviral capsid protein VP1 in pancreases of patients with type 1 (insulin-dependent) diabetes and pancreases and hearts of infants who died of coxsackieviral myocarditis. Diabetologia. 1990 May;33(5):290–298. doi: 10.1007/BF00403323. [DOI] [PubMed] [Google Scholar]
  11. Fukami Y., Nakamura T., Nakayama A., Kanehisa T. Phosphorylation of tyrosine residues of calmodulin in Rous sarcoma virus-transformed cells. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4190–4193. doi: 10.1073/pnas.83.12.4190. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fukazawa H., Mizuno S., Uehara Y. Effects of herbimycin A and various SH-reagents on p60v-src kinase activity in vitro. Biochem Biophys Res Commun. 1990 Nov 30;173(1):276–282. doi: 10.1016/s0006-291x(05)81053-8. [DOI] [PubMed] [Google Scholar]
  13. Graber M., June C. H., Samelson L. E., Weiss A. The protein tyrosine kinase inhibitor herbimycin A, but not genistein, specifically inhibits signal transduction by the T cell antigen receptor. Int Immunol. 1992 Nov;4(11):1201–1210. doi: 10.1093/intimm/4.11.1201. [DOI] [PubMed] [Google Scholar]
  14. Guinea R., López-Rivas A., Carrasco L. Modification of phospholipase C and phospholipase A2 activities during poliovirus infection. J Biol Chem. 1989 Dec 25;264(36):21923–21927. [PubMed] [Google Scholar]
  15. Holsey C., Cragoe E. J., Jr, Nair C. N. Evidence for poliovirus-induced cytoplasmic alkalinization in HeLa cells. J Cell Physiol. 1990 Mar;142(3):586–591. doi: 10.1002/jcp.1041420319. [DOI] [PubMed] [Google Scholar]
  16. Holsey C., Nair C. N. Poliovirus-induced intracellular alkalinization involves a proton ATPase and protein phosphorylation. J Cell Physiol. 1993 Jun;155(3):606–614. doi: 10.1002/jcp.1041550318. [DOI] [PubMed] [Google Scholar]
  17. Hunter T., Cooper J. A. Protein-tyrosine kinases. Annu Rev Biochem. 1985;54:897–930. doi: 10.1146/annurev.bi.54.070185.004341. [DOI] [PubMed] [Google Scholar]
  18. Irurzun A., Pérez L., Carrasco L. Enhancement of phospholipase activity during poliovirus infection. J Gen Virol. 1993 Jun;74(Pt 6):1063–1071. doi: 10.1099/0022-1317-74-6-1063. [DOI] [PubMed] [Google Scholar]
  19. June C. H., Fletcher M. C., Ledbetter J. A., Schieven G. L., Siegel J. N., Phillips A. F., Samelson L. E. Inhibition of tyrosine phosphorylation prevents T-cell receptor-mediated signal transduction. Proc Natl Acad Sci U S A. 1990 Oct;87(19):7722–7726. doi: 10.1073/pnas.87.19.7722. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kandolf R., Ameis D., Kirschner P., Canu A., Hofschneider P. H. In situ detection of enteroviral genomes in myocardial cells by nucleic acid hybridization: an approach to the diagnosis of viral heart disease. Proc Natl Acad Sci U S A. 1987 Sep;84(17):6272–6276. doi: 10.1073/pnas.84.17.6272. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kandolf R., Canu A., Hofschneider P. H. Coxsackie B3 virus can replicate in cultured human foetal heart cells and is inhibited by interferon. J Mol Cell Cardiol. 1985 Feb;17(2):167–181. doi: 10.1016/s0022-2828(85)80019-5. [DOI] [PubMed] [Google Scholar]
  22. Kandolf R., Hofschneider P. H. Molecular cloning of the genome of a cardiotropic Coxsackie B3 virus: full-length reverse-transcribed recombinant cDNA generates infectious virus in mammalian cells. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4818–4822. doi: 10.1073/pnas.82.14.4818. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kandolf R., Hofschneider P. H. Viral heart disease. Springer Semin Immunopathol. 1989;11(1):1–13. doi: 10.1007/BF00197080. [DOI] [PubMed] [Google Scholar]
  24. Kirchweger R., Ziegler E., Lamphear B. J., Waters D., Liebig H. D., Sommergruber W., Sobrino F., Hohenadl C., Blaas D., Rhoads R. E. Foot-and-mouth disease virus leader proteinase: purification of the Lb form and determination of its cleavage site on eIF-4 gamma. J Virol. 1994 Sep;68(9):5677–5684. doi: 10.1128/jvi.68.9.5677-5684.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kliewer S., Muchardt C., Gaynor R., Dasgupta A. Loss of a phosphorylated form of transcription factor CREB/ATF in poliovirus-infected cells. J Virol. 1990 Sep;64(9):4507–4515. doi: 10.1128/jvi.64.9.4507-4515.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Klingel K., Hohenadl C., Canu A., Albrecht M., Seemann M., Mall G., Kandolf R. Ongoing enterovirus-induced myocarditis is associated with persistent heart muscle infection: quantitative analysis of virus replication, tissue damage, and inflammation. Proc Natl Acad Sci U S A. 1992 Jan 1;89(1):314–318. doi: 10.1073/pnas.89.1.314. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Klump W. M., Bergmann I., Müller B. C., Ameis D., Kandolf R. Complete nucleotide sequence of infectious Coxsackievirus B3 cDNA: two initial 5' uridine residues are regained during plus-strand RNA synthesis. J Virol. 1990 Apr;64(4):1573–1583. doi: 10.1128/jvi.64.4.1573-1583.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kräusslich H. G., Nicklin M. J., Lee C. K., Wimmer E. Polyprotein processing in picornavirus replication. Biochimie. 1988 Jan;70(1):119–130. doi: 10.1016/0300-9084(88)90166-6. [DOI] [PubMed] [Google Scholar]
  29. Kräusslich H. G., Nicklin M. J., Toyoda H., Etchison D., Wimmer E. Poliovirus proteinase 2A induces cleavage of eucaryotic initiation factor 4F polypeptide p220. J Virol. 1987 Sep;61(9):2711–2718. doi: 10.1128/jvi.61.9.2711-2718.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Lacy J., Bartiss A., Coleman D. L. Conversion of a human B cell lymphoma line by Epstein-Barr virus is associated with increased tyrosine phosphorylation of a 50 kilodalton cytosolic protein. Virus Res. 1991 Jun;20(1):85–96. doi: 10.1016/0168-1702(91)90063-2. [DOI] [PubMed] [Google Scholar]
  31. Ledbetter J. A., Linsley P. S. CD28 receptor crosslinking induces tyrosine phosphorylation of PLC gamma 1. Adv Exp Med Biol. 1992;323:23–27. [PubMed] [Google Scholar]
  32. Majumder S., Ray P., Besmer P. Tyrosine protein kinase activity of the HZ4-feline sarcoma virus P80gag-kit-transforming protein. Oncogene Res. 1990;5(4):329–335. [PubMed] [Google Scholar]
  33. Mohammadi M., Dionne C. A., Li W., Li N., Spivak T., Honegger A. M., Jaye M., Schlessinger J. Point mutation in FGF receptor eliminates phosphatidylinositol hydrolysis without affecting mitogenesis. Nature. 1992 Aug 20;358(6388):681–684. doi: 10.1038/358681a0. [DOI] [PubMed] [Google Scholar]
  34. Uehara Y., Fukazawa H., Murakami Y., Mizuno S. Irreversible inhibition of v-src tyrosine kinase activity by herbimycin A and its abrogation by sulfhydryl compounds. Biochem Biophys Res Commun. 1989 Sep 15;163(2):803–809. doi: 10.1016/0006-291x(89)92293-6. [DOI] [PubMed] [Google Scholar]
  35. Vetter M. L., Martin-Zanca D., Parada L. F., Bishop J. M., Kaplan D. R. Nerve growth factor rapidly stimulates tyrosine phosphorylation of phospholipase C-gamma 1 by a kinase activity associated with the product of the trk protooncogene. Proc Natl Acad Sci U S A. 1991 Jul 1;88(13):5650–5654. doi: 10.1073/pnas.88.13.5650. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Werner S., Klump W. M., Schönke H., Hofschneider P. H., Kandolf R. Expression of coxsackievirus B3 capsid proteins in Escherichia coli and generation of virus-specific antisera. DNA. 1988 Jun;7(5):307–316. doi: 10.1089/dna.1.1988.7.307. [DOI] [PubMed] [Google Scholar]
  37. de Verdugo U. R., Selinka H. C., Huber M., Kramer B., Kellermann J., Hofschneider P. H., Kandolf R. Characterization of a 100-kilodalton binding protein for the six serotypes of coxsackie B viruses. J Virol. 1995 Nov;69(11):6751–6757. doi: 10.1128/jvi.69.11.6751-6757.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES