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Tobacco mosaic virus. M.M. Hernandez. Tobacco mosaic virus TMV. Genus Tobamovirus 15 members naked, rigid rod, + unsegmented ss RNA No family assignation by ICTV. 2 nm. 18 nm. NC protein. STRUCTURAL FEATURES OF TMV. CAPSID. 30 nm . Helical symmetry. RNA genome.
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Tobacco mosaic virus M.M. Hernandez
Tobacco mosaic virusTMV • Genus Tobamovirus 15 members naked, rigid rod, + unsegmented ss RNA • No family assignation by ICTV
2 nm 18 nm NC protein STRUCTURAL FEATURES OF TMV CAPSID 30 nm Helical symmetry RNA genome Images:P. Sforza, 2001 www.ppws.vt.edu/~sforza/tmv/tmv.html
TMV genome organization 6,395 nt Capsid CP Replicase RdRp tRNAhis 183K 17.6 K 30 K UAG (leaky) Movement MP MT-Hel 5’ cap 126 K
Host:Nicotiana tabaccum(Solanaceous ) & other 9 plant families susceptible too
Chlorosis Necrosis Stunting Mosaic disease: significant losses in yield & $ value
TMV Life cycle 5’ cap Host Rb a) Virus entry trough abrasions on plant tissue. Inside cell associates with ER b) spontaneous release of few capsid (CP) subunits 5' end of genome is uncovered c) Host ribosome attaches to viral RNA, moves down displacing more CP units …/… Modification of a diagram from: L. Stannard, Department of Medical Microbiology, U. of Cape Town
TMV Life cycle (cont.) 5’ d) Ribosome meets start codon, translates first two proteins (126K ,183 K) while uncoating continues “co-traslational disassembly” e) 126 K ( MET-Hel) & 183 K ( RdRp) use viral RNA as template to make full length complementary neg. strand RNA …/… Modification of a diagram from: L. Stannard, Department of Medical Microbiology, U. of Cape Town
TMV Life cycle (cont.) RdRp + Strand (genome) Neg. strand f) Neg. RNA strand used by viral replicase (RdPp/MET-Hel ) as template for +RNA g) Also, neg. RNA strand has internal promoters used by replicase to make mRNA for 30K protein (MP) and 17.5 K (CP) promoters Transcription by RdRp MP mRNA CP mRNA
TMV Life cycle (cont.) h) MP combines with viral +RNA to move it into new plant cells through plasmodesmata http:www.sunysb.edu/biochem/BIOCHEM/facultypages/citovsky/TMV-TransportModel-III2.jpg
TMV Life cycle (cont.) i) Accumulation of +RNA & CP proteins stimulates assembly of progeny virions Viral RNA interacts with discs CP units self assemble into discs Electrostatic interaction stabilizes structure into helix M. Olvera de la Cruz
TMV Life cycle (cont.) j) massive TMR replication occur in the X-bodies (viroplasmas) Szecsi et al. 1999 Hamacher et al. 2003
TMV Life cycle (cont.) k) Virus will remain in plant tissue and sap until delivered to a new plant host usually by mechanical means
TMV diagnosis • Symptoms • E.M. & Serology • RT-PCR & RFLP Letschert et al. 2002 HTTP;//BAKERLAB.BERKELEY.EDU/?IMG=1
Current Research on TMV • Improving diagnostic techniques • Characterization of MP and viral movement through plant • Characterization of the Helicase & RdRp
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Khan J. A. and J. Dijkstra editors, 2002. Plant Viruses as Molecular Pathogens Food Product Press 7 An Imprint of the Haworth Press Inc. New York Lazarowitz S. G., 2001. Plant Viruses In: Fundamental Virology Fourth Edition. Knipe D. and P.M. Howley Editors. Lippincott Williams & Wilkins, Philadelphia. pp:377-429 Lesemann D.-E., S. Winters. 1998. Detection and specific identification of tobamoviruses in horticultural crops by immunoelectron microscopical methods (IEM) and ELISA. Mitteilungender Biologischen Bundesanstalt fur Land-und Forstwirtschaft 357 Letschert B., A Gunter, L. Dietrich-Eckhardt, P. Willingmann, C. Heinze., 2002. Detection and differentiation of serologically cross-reacting tobamoviruses of economical importance by RT-PCR and RT-PCR-RFLP. Journal of Virological Methods 106:1-10 Lucas G. B. , C.L. Campbell, and L.T. Lucas, 1985. Introduction to Plant Diseases Identification and Management. The AVI Publishing Company, Inc. Westport Connecticut. pp 254-261 Martin J.H. , W.H. Leonard and D. L. Stamp 1976 Principles of Field Crop Production Third Edition MacMills Publishing Co., INC. London Matthews R.E.F., 1992. Fundamentals of Plant Virology Academic Press Inc San Diego Matthews R.E.F., 1999 Plant Virology Third Edition. Academic Press Inc San Diego. pp:207-225 Osman T. A. M., and K.W. Buck. 2003 Identification of a Region of the Tobacco Mosaic Virus 126- and 183-Kilodalton Replication Proteins Which bind Specifically to the Viral 3'-Terminal tRNA-like Structure. Journal of Virology 77(16 ) 8669-8675 Padmanadban M.S., Goregaoker S.P., S. Golem, H. Shiferaw, and J. N. Culver. 2005. Interaction of the Tobacco Mosaic Virus Replicase Protein with the Aux/IAA Protein PAP1/IAA26 Is associated with Disease Development. Journal of Virology. 79 (4) 2549 - 2558 Sacristan S., J. M. Malpica, A. Fraile, F. Garcia-Arenal. 2003. Estimation of Population Bottlenecks during Systemic Movement of Tobacco Mosaic Virus in Tobacco Plants. Journal of Virology 77(18) 9906-9911 Seemanpillai M., R. Elamawi, C. Ritzenthhaler, M. Heinlein. 2006. Challenging the Role of Microtubules in Tobacco Mosaic Virus Movement by Drug Treatment is Disputable. Journal of Virology 80(13): 6712-6715 Torok V.A. and J.W. Randles. 2001. Tobacco mosaic virus RNA as an internal control for duplex RT-PCR assay of pea germaplsm. Australian Plant Pathology 30: 227-230 Zaitlin M., 2001. Tobamovirus in: The Spriger Index of Viruses. Tidona C. A. and G. Darai Editors. Springer-Verlag, Berlin
2. Add ssTMV+ RNA (assembly infective virus): Electrostatic attraction stabilizes viral assembly. 1. Solution of TMV Capsid Proteins Electrostatic repulsion inhibits + protein +protein aggregation. Salt (M) ZRNA= -1/mer 3. Sphere-like (icosahedral) RNA virus (Tsuruta et al) Acidity 4. Capsid + Hole (density ≈ RNA crystal) self-assembles from capsid protein + viral RNA solution