The Poxviridae family members vaccinia and variola virus enter mammalian cells, replicate outside the nucleus and produce virions that travel to the cell surface along microtubules, fuse with the plasma membrane and egress from infected cells toward apposing cells on actin-filled membranous protrusions. We show that cell-associated enveloped virions (CEV) use Abl- and Src-family tyrosine kinases for actin motility, and that these kinases act in a redundant fashion, perhaps permitting motility in a greater range of cell types. Additionally, release of CEV from the cell requires Abl- but not Src-family tyrosine kinases, and is blocked by STI-571 (Gleevec), an Abl-family kinase inhibitor used to treat chronic myelogenous leukemia in humans. Finally, we show that STI-571 reduces viral dissemination by five orders of magnitude and promotes survival in infected mice, suggesting possible use for this drug in treating smallpox or complications associated with vaccination. This therapeutic approach may prove generally efficacious in treating microbial infections that rely on host tyrosine kinases, and, because the drug targets host but not viral molecules, this strategy is much less likely to engender resistance compared to conventional antimicrobial therapies.
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Esposito, J. & Fenner, F. in Fields Virology 4th edn. Vol. 2 (eds. Knipe, D.M. & Howley, P.M.) Ch. 85, 2885–2921 (Lippincott, Williams and Wilkins, Philadelphia, 2001.)
Moss, B. in Fields Virology 4th edn. Vol. 2 (eds. Knipe, D.M. and Howley, P.M.) Ch. 84, 2849–2883 (Lippincott, Williams and Wilkins, Philadelphia, 2001).
Crotty, S. et al. Cutting edge: long-term B cell memory in humans after smallpox vaccination. J. Immunol. 171, 4969–4973 (2003).
National Research Council, Institute of Medicine. Assessment of Future Scientific Needs for Live Variola Virus 126 (National Academy Press, Washington, DC, 1999).
Harrison, S.C. et al. Discovery of antivirals against smallpox. Proc. Natl. Acad. Sci. USA 101, 11178–11192 (2004).
Amorosa, V.K. & Isaacs, S.N. Separate worlds set to collide: smallpox, vaccinia virus vaccination, and human immunodeficiency virus and acquired immunodeficiency syndrome. Clin. Infect. Dis. 37, 426–432 (2003).
Rotz, L.D., Dotson, D.A., Damon, I.K., Becher, J.A. Advisory Committee on Immunization Practices. Vaccinia (smallpox) vaccine recommendations of the Advisory Committee on Immunization Practices (ACIP), 2001. MMWR Recomm. Rep. 50, 1–25 (2001).
Cono, J., Casey, C.G., Bell, D.M. Centers for Disease Control and Prevention. Smallpox vaccination and adverse reactions guidance for clinicians. MMWR Recomm. Rep. 52, 1–28 (2003).
Smith, J.N. & Ahmer, B.M. Detection of other microbial species by Salmonella: expression of the SdiA regulon. J. Bacteriol. 185, 1357–1366 (2003).
Smith, G.L., Murphy, B.J. & Law, M. Vaccinia virus motility. Annu. Rev. Microbiol. 57, 323–342 (2003).
Carter, G.C. et al. Vaccinia virus cores are transported on microtubules. J. Gen. Virol. 84, 2443–2458 (2003).
Hollinshead, M. et al. Vaccinia virus utilizes microtubules for movement to the cell surface. J. Cell Biol. 154, 389–402 (2001).
Rietdorf, J. et al. Kinesin-dependent movement on microtubules precedes actin-based motility of vaccinia virus. Nat. Cell Biol. 3, 992–1000 (2001).
Ward, B.M. & Moss, B. Vaccinia virus intracellular movement is associated with microtubules and independent of actin tails. J. Virol. 75, 11651–11663 (2001).
Smith, G.L., Vanderplasschen, A. & Law, M. The formation and function of extracellular enveloped vaccinia virus. J. Gen. Virol. 83, 2915–2931 (2002).
Wolffe, E.J., Weisberg, A.S. & Moss, B. Role for the vaccinia virus A36R outer envelope protein in the formation of virus-tipped actin-containing microvilli and cell-to-cell virus spread. Virology 244, 20–26 (1998).
Parkinson, J.E. & Smith, G.L. Vaccinia virus gene A36R encodes a M(r) 43-50 K protein on the surface of extracellular enveloped virus. Virology 204, 376–390 (1994).
Frischknecht, F. et al. Actin-based motility of vaccinia virus mimics receptor tyrosine kinase signalling. Nature 401, 926–929 (1999).
Newsome, T.P., Scaplehorn, N. & Way, M. SRC mediates a switch from microtubule- to actin-based motility of vaccinia virus. Science 306, 124–129 (2004).
Frischknecht, F. & Way, M. Surfing pathogens and the lessons learned for actin polymerization. Trends Cell Biol. 11, 30–38 (2001).
Moreau, V. et al. A complex of N-WASP and WIP integrates signalling cascades that lead to actin polymerization. Nat. Cell Biol. 2, 441–448 (2000).
Scaplehorn, N. et al. Grb2 and Nck act cooperatively to promote actin-based motility of vaccinia virus. Curr. Biol. 12, 740–745 (2002).
Nataro, J.P. & Kaper, J.B. Diarrheagenic Escherichia coli . Clin. Microbiol. Rev. 11, 142–201 (1998).
Kenny, B. et al. Enteropathogenic E. coli (EPEC) transfers its receptor for intimate adherence into mammalian cells. Cell 91, 511–520 (1999).
Kalman, D. et al. Enteropathogenic E. coli acts through WASP and Arp2/3 complex to form actin pedestals. Nat. Cell Biol. 1, 389–391 (1999).
Swimm, A. et al. Enteropathogenic Escherichia coli use redundant tyrosine kinases to form actin pedestals. Mol. Biol. Cell 15, 3520–3529 (2004).
Schindler, T. et al. Structural mechanism for STI-571 inhibition of abelson tyrosine kinase. Science 289, 1938–1942 (2000).
Druker, B.J. et al. Chronic myelogenous leukemia. Hematology (Am Soc Hematol Educ Program) 87–112 (2001).
Goldman, J.M. & Druker, B.J. Chronic myeloid leukemia: current treatment options. Blood 98 2039–2042 (2001).
Yuwen, H. et al. Nuclear localization of a double-stranded RNA-binding protein encoded by the vaccinia virus E3L gene. Virology 195, 732–744 (1993).
Ward, B.M. & Moss, B. Visualization of intracellular movement of vaccinia virus virions containing a green fluorescent protein-B5R membrane protein chimera. J. Virol. 75, 4802–4813 (2001).
Pluk, H., Dorey, K. & Superti-Furga, G. Autoinhibition of c-Abl. Cell 108, 247–259 (2002).
Kraker, A.J. et al. Biochemical and cellular effects of c-Src kinase-selective pyrido[2, 3- d]pyrimidine tyrosine kinase inhibitors. Biochem. Pharmacol. 60, 885–898 (2000).
Dorsey, J.F. et al. The pyrido[2,3-d]pyrimidine derivative PD180970 inhibits p210Bcr-Abl tyrosine kinase and induces apoptosis of K562 leukemic cells. Cancer Res. 60, 3127–3131 (2000).
Wisniewski, D. et al. Characterization of potent inhibitors of the Bcr-Abl and the c-kit receptor tyrosine kinases. Cancer Res. 62, 4244–4255 (2002).
Liu, Y. et al. Structural basis for selective inhibition of Src family kinases by PP1. Chem. Biol. 6, 671–678 (1999).
Tatton, L. et al. The Src-selective kinase inhibitor PP1 also inhibits Kit and Bcr-Abl tyrosine kinases. J. Biol. Chem. 278, 4847–4853 (2003).
Ward, B.M., Weisberg, A.S. & Moss, B. Mapping and functional analysis of interaction sites within the cytoplasmic domains of the vaccinia virus A33R and A36R envelope proteins. J. Virol. 77, 4113–4126 (2003).
Law, M. & Smith, G.L. Antibody neutralization of the extracellular enveloped form of vaccinia virus. Virology 280, 132–142 (2001).
Law, M., Hollinshead, R. & Smith, G.L. Antibody-sensitive and antibody-resistant cell-to-cell spread by vaccinia virus: role of the A33R protein in antibody-resistant spread. J. Gen. Virol. 83, 209–222 (2002).
Blasco, R., Sisler, J.R. & Moss, B. Dissociation of progeny vaccinia virus from the cell membrane is regulated by a viral envelope glycoprotein: effect of a point mutation in the lectin homology domain of the A34R gene. J. Virol. 67, 3319–3325 (1993).
Ichihashi, Y. & Oie, M. Neutralizing epitope on penetration protein of vaccinia virus. Virology 220, 491–494 (1996).
Vanderplasschen, A. et al. Extracellular enveloped vaccinia virus is resistant to complement because of incorporation of host complement control proteins into its envelope. Proc. Natl. Acad. Sci. USA 95, 7544–7549 (1998).
Boulter, E.A. & Appleyard, G. Differences between extracellular and intracellular forms of poxvirus and their implications. Prog. Med. Virol. 16, 86–108 (1973).
Appleyard, G., Hapel, A.J. & Boulter, E.A. An antigenic difference between intracellular and extracellular rabbitpox virus. J. Gen. Virol. 13, 9–17 (1971).
Wolff, N.C. & Ilaria, R.L. Jr. Establishment of a murine model for therapy-treated chronic myelogenous leukemia using the tyrosine kinase inhibitor STI571. Blood 98, 2808–2816 (2001).
Chahroudi, A., Chavan, R., Kozyr, N., Silvestri, G. & Feinberg, M.B. Vaccinia virus tropism for primary hematolymphoid cells is determined by restricted expression of a unique virus receptor. J. Virol. in the press (2005).
Ichaso, N. & Dilworth, S.M. Cell transformation by the middle T-antigen of polyoma virus. Oncogene 20, 7908–7916 (2001).
Gruenheid, S. et al. Enteropathogenic E. coli Tir binds Nck to initiate actin pedestal formation in host cells. Nat. Cell Biol. 3, 856–859 (2001).
Bishop, J.M. Molecular themes in oncogenesis. Cell 64, 235–248 (1991).
Baker, R.O., Bray, M. & Huggins, J.W. Potential antiviral therapeutics for smallpox, monkeypox and other orthopoxvirus infections. Antiviral Res. 57, 13–23 (2003).
Idemyor, V. Bacterial resistance to antimicrobial agents--the time for concern. Ann. Pharmacother. 27, 1285 (1993).
Li, X.Z. & Nikaido, H. Efflux-mediated drug resistance in bacteria. Drugs 64, 159–204 (2004).
Gould, I.M. Antibiotic policies and control of resistance. Curr. Opin. Infect. Dis. 15, 395–400 (2002).
Koleske, A.J. et al. Essential roles for the Abl and Arg tyrosine kinases in neurulation. Neuron 21, 1259–1272 (1998).
Nagar, B. et al. Crystal structures of the kinase domain of c-Abl in complex with the small molecule inhibitors PD173955 and imatinib (STI-571). Cancer Res. 62, 4236–4243 (2002).
Kalman, D. et al. Ras family GTPases control growth of astrocyte processes. Mol. Biol. Cell 10, 1665–1683 (1999).
Swedlow, J.R., Sedat, J.W. & Agard, D.A. Deconvolution in Optical Microscopy, in Deconvolution of Images and Spectra (ed. Jansson, P.A.) 284–307 (Academic Press, San Diego, 1997).
Ramirez, J.C. et al. Tissue distribution of the Ankara strain of vaccinia virus (MVA) after mucosal or systemic administration. Arch. Virol. 148, 827–839 (2003).
The authors thank D. Kalman, D. Steinhauer, O. Weiner, J. Taunton and K. Saxe for discussions; A. Family, S. Staprans, N. Kozyr and R. Griffith for assistance and advice; B. Meyer and J. Wang for Abl cDNAs; T. Koleske for Abl1−/−Abl2−/− cells, Abl1 and Abl2 cells, and c-Arg-YFP cDNA; B. Moss and J. Yudell for α-TW2.3 mAb; R. Blasco for GFP-VV; C. Lowell for Src−/− cells, Src−/−Fyn−/− cells, and Src−/−Yes1−/− cells; G. Smith for α-IMV mAb; and L. Burleigh, D. Steinhauer, and C. Moran for commenting on the manuscript. The work was supported by US National Institutes of Health grant PO1 AI 46007 (to M.B.F.), and by grants from the University Research Council, Emory University, the Southeastern Regional Center for Excellence in Bioterrorism (SERCEB) Pilot Project Feasibility Award, grant R01-AI056067-01 from the N.I.A.I.D., and an award from the Emtech Biotechnology Foundation (all to D.K.).
The authors declare no competing financial interests.
Protein sequences surrounding phosphorylation sites in VV A36R and enteropathogenic E. coli Tir. (PDF 53 kb)
Identification of VV-infected cells. (PDF 352 kb)
Effects of PD-166326 and STI-571. (PDF 339 kb)
Characterization of kinases sufficient for VV actin motility. (PDF 166 kb)
Abl-family kinases do not affect viral entry or replication. (PDF 356 kb)
Distinguishing the effects of Abl and Arg on EEV and IMV. (PDF 62 kb)
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