Tomato leaf curl disease (ToLCD) is manifested by yellowing of leaf lamina with upward leaf curl, leaf distortion, shrinking of the leaf surface, and stunted plant growth caused by tomato leaf curl virus (ToLCV). In the present study, using computational methods we explored the evolutionary and molecular prospects of viral coat protein derived from an isolate of Vadodara district, Gujarat (ToLCGV-[Vad]), India. We found that the amino acids in coat protein required for systemic infection, viral particle formation, and insect transmission to host cells were conserved amongst Indian strains. Phylogenetic studies on Indian ToLCV coat proteins showed evolutionary compatibility with other viral taxa. Modeling of coat protein revealed a topology similar to characteristic Geminate viral particle consisting of antiparallel β-barrel motif with N-terminus α-helix. The molecular interaction of coat protein with the viral DNA required for encapsidation and nuclear shuttling was investigated through sequence- and structure-based approaches. We further emphasized the role of loops in coat protein structure as molecular recognition interface. 1. Introduction Tomato leaf curl virus (ToLCV) is one of the most devastating causal agents of tomato (Solanum lycopersicum) crop which had emerged causing damage and encroaching new areas in tropical and subtropical continents every year. Plant-infecting geminiviruses belong to the family Geminiviridae in which Begomovirus is one among the genera possessing both mono- and bipartite genomes that infect especially dicotyledonous plant species [1]. The disease is marked by symptoms such as yellowing of leaf lamina with upward leaf curl as well as distortion, reduction in internodes, new leaves size reduction, wrinkle facade, stunted growth, and dissemination of flower from plant before onset of fruiting. ToLCV is primarily transmitted by sweet potato whitefly (Bemisia tabaci) and silver leaf whitefly (also called Biotype B; Bemisia argentifolii). Whiteflies harboring virus can nonspecifically infect a wide spectrum of plant crops and weeds including eggplant, potato, tobacco, pepper, and common bean. Infected plants seem healthy but develop symptoms leading to enormous economic loss [2]. In Indian subcontinent, ToLCV is a major problem for tomato-growing regions as several reports on new strains have been documented including New Delhi, Lucknow, Bangalore, Varanasi, Mirzapur, Vadodara, and so forth and posed a threat to crop productivity [6]. Indian ToLCV isolates are mostly monopartite (DNA-A) in nature with few isolates
References
[1]
B. Gronenbor, “The Tomato yellow leaf curl virus genome,” in Tomato Yellow Leaf Curl Virus Disease: Management, Molecular Biology, Breeding For Resistance, H. Czosnek, Ed., vol. 8, pp. 67–84, Springer, Netherlands, 2007.
[2]
M. J. Melzer, D. Y. Ogata, S. K. Fukuda et al., “First report of Tomato yellow leaf curl virus in Hawaii,” Plant Disease, vol. 94, no. 5, pp. 641–643, 2010.
[3]
A. A. Sanderfoot, D. J. Ingham, and S. G. Lazarowitz, “A viral movement protein as a nuclear shuttle: the geminivirus BR1 movement protein contains domains essential for interaction with BL1 and nuclear localization,” Plant Physiology, vol. 110, no. 1, pp. 23–33, 1996.
[4]
H. Liu, M. I. Boulton, C. L. Thomas, D. A. M. Prior, K. J. Oparka, and J. W. Davies, “Maize streak virus coat protein is karyophyllic and facilitates nuclear transport of viral DNA,” Molecular Plant-Microbe Interactions, vol. 12, no. 10, pp. 894–900, 1999.
[5]
T. Kunik, K. Palanichelvam, H. Czosnek, V. Citovsky, and Y. Gafni, “Nuclear import of the capsid protein of Tomato yellow leaf curl virus (TYLCV) in plant and insect cells,” Plant Journal, vol. 13, no. 3, pp. 393–399, 1998.
[6]
S. Chakraborty, P. K. Pandey, M. K. Banerjee, G. Kalloo, and C. M. Fauquet, “Tomato leaf curl Gujarat virus, a New begomovirus species causing a severe leaf curl disease of tomato in Varanasi, India,” Phytopathology, vol. 93, no. 12, pp. 1485–1495, 2003.
[7]
R. W. Briddon, J. K. Brown, E. Moriones et al., “Recommendations for the classification and nomenclature of the DNA-β satellites of Begomoviruses,” Archives of Virology, vol. 153, no. 4, pp. 763–781, 2008.
[8]
A. Kheyr-Pour, M. Bendahmane, V. Matzeit, G. P. Accotto, S. Crespi, and B. Gronenborn, “Tomato yellow leaf curl virus from Sardinia is a whitefly-transmitted monopartite geminivirus,” Nucleic Acids Research, vol. 19, no. 24, pp. 6763–6769, 1991.
[9]
P. Jyothsna, R. Rawat, and V. G. Malathi, “Predominance of Tomato leaf curl Gujarat virus as a monopartite Begomovirus: association with Tomato yellow leaf curl Thailand betasatellite,” Archives of Virology. In press.
[10]
P. Pandey, S. Mukhopadhya, A. R. Naqvi, S. K. Mukherjee, G. S. Shekhawat, and N. R. Choudhury, “Molecular characterization of two distinct monopartite Begomoviruses infecting tomato in india,” Virology Journal, vol. 7, article no. 337, 2010.
[11]
T. Kunik, R. Salomon, D. Zamir et al., “Transgenic tomato plants expressing the Tomato yellow leaf curl virus capsid protein are resistant to the virus,” Nature Biotechnology, vol. 12, no. 5, pp. 500–504, 1994.
[12]
S. K. Raj, R. Singh, S. K. Pandey, and B. P. Singh, “Agrobacterium-mediated tomato transformation and regeneration of transgenic lines expressing Tomato leaf curl virus coat protein gene for resistance against TLCV infection,” Current Science, vol. 88, no. 10, pp. 1674–1679, 2005.
[13]
D. Pratap, S. K. Raj, S. Kumar, S. K. Snehi, K. K. Gautam, and A. K. Sharma, “Coat protein-mediated transgenic resistance in tomato against a IB subgroup Cucumber mosaic virus strain,” Phytoparasitica, vol. 40, no. 4, pp. 375–382, 2012.
[14]
S. Chakraborty, R. Vanitharani, B. Chattopadhyay, and C. M. Fauquet, “Supervirulent pseudorecombination and asymmetric synergism between genomic components of two distinct species of Begomovirus associated with severe tomato leaf curl disease in India,” Journal of General Virology, vol. 89, no. 3, pp. 818–828, 2008.
[15]
N. Kirthi, S. P. Maiya, M. R. N. Murthy, and H. S. Savithri, “Evidence for recombination among the Tomato leaf curl virus strains/species from Bangalore, India,” Archives of Virology, vol. 147, no. 2, pp. 255–272, 2002.
[16]
D. A. Benson, I. Karsch-Mizrachi, D. J. Lipman, J. Ostell, and D. L. Wheeler, “GenBank,” Nucleic Acids Research, vol. 34, pp. D16–D20, 2006.
[17]
A. Marchler-Bauer and S. H. Bryant, “CD-Search: protein domain annotations on the fly,” Nucleic Acids Research, vol. 32, pp. W327–W331, 2004.
[18]
S. F. Altschul, T. L. Madden, A. A. Sch？ffer et al., “Gapped BLAST and PSI-BLAST: a new generation of protein database search programs,” Nucleic Acids Research, vol. 25, no. 17, pp. 3389–3402, 1997.
[19]
A. Bateman, E. Birney, L. Cerruti et al., “The pfam protein families database,” Nucleic Acids Research, vol. 30, no. 1, pp. 276–280, 2002.
[20]
S. Kosugi, M. Hasebe, T. Entani, S. Takayama, M. Tomita, and H. Yanagawa, “Design of peptide inhibitors for the importin α/β nuclear import pathway by activity-based profiling,” Chemistry and Biology, vol. 15, no. 9, pp. 940–949, 2008.
[21]
H. Liu, M. I. Boulton, and J. W. Davies, “Maize streak virus coat protein binds single- and double-stranded DNA in vitro,” Journal of General Virology, vol. 78, no. 6, pp. 1265–1270, 1997.
[22]
P. Rice, L. Longden, and A. Bleasby, “EMBOSS: the European molecular biology open software suite,” Trends in Genetics, vol. 16, no. 6, pp. 276–277, 2000.
[23]
M. A. Larkin, G. Blackshields, N. P. Brown et al., “Clustal W and Clustal X version 2.0,” Bioinformatics, vol. 23, no. 21, pp. 2947–2948, 2007.
[24]
J. Felsenstein, “PHYLIP- phylogeny inference package (version 3. 2),” Cladistics, vol. 5, no. 2, pp. 164–166, 1989.
[25]
M. Johnson, I. Zaretskaya, Y. Raytselis, Y. Merezhuk, S. McGinnis, and T. L. Madden, “NCBI BLAST: a better web interface,” Nucleic acids research, vol. 36, pp. W5–9W, 2008.
[26]
F. C. Bernstein, T. F. Koetzle, and G. J. B. Williams, “The protein data bank: a computer based archival file for macromolecular structures,” Journal of Molecular Biology, vol. 112, no. 3, pp. 535–542, 1977.
[27]
J. J. Ward, L. J. McGuffin, K. Bryson, B. F. Buxton, and D. T. Jones, “The DISOPRED server for the prediction of protein disorder,” Bioinformatics, vol. 20, no. 13, pp. 2138–2139, 2004.
[28]
D. E. Kim, D. Chivian, and D. Baker, “Protein structure prediction and analysis using the Robetta server,” Nucleic Acids Research, vol. 32, pp. W526–W531, 2004.
[29]
L. A. Kelley and M. J. Sternberg, “Protein structure prediction on the web: a case study using the Phyre server,” Nature protocols, vol. 4, no. 3, pp. 363–371, 2009.
[30]
Tripos, A Certara Company, “Tripos benchware 3D explorer for interactive molecular visualizations and structure manipulations,” St. Louis, Miss, USA, http://www.tripos.com/.
[31]
G. N. Ramachandran, C. Ramakrishnan, and V. Sasisekharan, “Stereochemistry of polypeptide chain configurations,” Journal of Molecular Biology, vol. 7, pp. 95–99, 1963.
[32]
S. C. Lovell, I. W. Davis, W. B. Arendall et al., “Structure validation by Cα geometry: φ,ψ and Cβ deviation,” Proteins, vol. 50, no. 3, pp. 437–450, 2003.
[33]
E. Pascal, A. A. Sanderfoot, B. M. Ward, R. Medville, R. Turgeon, and S. G. Lazarowitz, “The geminivirus BR1 movement protein binds single-stranded DNA and localizes the cell nucleus,” Plant Cell, vol. 6, no. 7, pp. 995–1006, 1994.
[34]
M. R. Rojas, A. O. Noueiry, W. J. Lucas, and R. L. Gilbertson, “Bean dwarf mosaic Geminivirus movement proteins recognize DNA in a form- and size-specific manner,” Cell, vol. 95, no. 1, pp. 105–113, 1998.
[35]
L. Wang and S. J. Brown, “BindN: a web-based tool for efficient prediction of DNA and RNA binding sites in amino acid sequences,” Nucleic Acids Research, vol. 34, pp. W243–W248, 2006.
[36]
Y. Tsuchiya, K. Kinoshita, and H. Nakamura, “PreDs: a server for predicting dsDNA-binding site on protein molecular surfaces,” Bioinformatics, vol. 21, no. 8, pp. 1721–1723, 2005.
[37]
M. van Dijk and A. M. Bonvin, “3D-DART: a DNA structure modelling server,” Nucleic Acids Research, vol. 37, no. 2, pp. W235–W239, 2009.
[38]
S. J. de Vries, M. van Dijk, and A. M. Bonvin, “The HADDOCK web server for data-driven biomolecular docking,” Nature protocols, vol. 5, no. 5, pp. 883–897, 2010.
[39]
S. Jo, M. Vargyas, J. Vasko-Szedlar, B. Roux, and W. Im, “PBEQ-Solver for online visualization of electrostatic potential of biomolecules,” Nucleic acids research, vol. 36, pp. W270–W275, 2008.
[40]
T. J. Dolinsky, J. E. Nielsen, J. A. McCammon, and N. A. Baker, “PDB2PQR: an automated pipeline for the setup of Poisson-Boltzmann electrostatics calculations,” Nucleic Acids Research, vol. 32, pp. W665–W667, 2004.
[41]
M. A. Lill and M. L. Danielson, “Computer-aided drug design platform using PyMOL,” Journal of Computer-Aided Molecular Design, vol. 25, no. 1, pp. 13–19, 2011.
[42]
A. O. Noueiry, W. J. Lucas, and R. L. Gilbertson, “Two proteins of a plant DNA virus coordinate nuclear and plasmodesmal transport,” Cell, vol. 76, no. 5, pp. 925–932, 1994.
[43]
K. Palanichelvam, T. Kunik, V. Citovsky, and Y. Gafni, “The capsid protein of Tomato yellow leaf curl virus binds cooperatively to single-stranded DNA,” Journal of General Virology, vol. 79, no. 11, pp. 2829–2833, 1998.
[44]
S. Qin, B. M. Ward, and S. G. Lazarowitz, “The bipartite geminivirus coat protein aids BR1 function in viral movement by affecting the accumulation of viral single-stranded DNA,” Journal of Virology, vol. 72, no. 11, pp. 9247–9256, 1998.
[45]
M. Hussain, S. Mansoor, S. Iram, A. N. Fatima, and Y. Zafar, “The nuclear shuttle protein of Tomato leaf curl New Delhi virus is a pathogenicity determinant,” Journal of Virology, vol. 79, no. 7, pp. 4434–4439, 2005.
[46]
M. R. Rojas, H. Jiang, R. Salati et al., “Functional analysis of proteins involved in movement of the monopartite Begomovirus, Tomato yellow leaf curl virus,” Virology, vol. 291, no. 1, pp. 110–125, 2001.
[47]
E. Noris, A. M. Vaira, P. Caciagli, V. Masenga, B. Gronenborn, and G. P. Accotto, “Amino acids in the capsid protein of Tomato yellow leaf curl virus that are crucial for systemic infection, particle formation, and insect transmission,” Journal of Virology, vol. 72, no. 12, pp. 10050–10057, 1998.
[48]
T. A. Jones and L. Liljas, “Structure of satellite tobacco necrosis virus after crystallographic refinement at 2.5？？ resolution,” Journal of Molecular Biology, vol. 177, no. 4, pp. 735–767, 1984.
[49]
W. Zhang, N. H. Olson, T. S. Baker et al., “Structure of the maize streak virus geminate particle,” Virology, vol. 279, no. 2, pp. 471–477, 2001.
[50]
M. S. Pinner, V. Medina, K. A. Plaskitt, and P. G. Markham, “Viral inclusions in monocotyledons infected by Maize streak and related viruses,” Plant Pathology, vol. 42, no. 1, pp. 75–87, 1993.
[51]
A. Zrachya, P. P. Kumar, U. Ramakrishnan et al., “Production of siRNA targeted against TYLCV coat protein transcripts leads to silencing of its expression and resistance to the virus,” Transgenic Research, vol. 16, no. 3, pp. 385–398, 2007.
[52]
S. V. Ramesh, A. K. Mishra, and S. Praveen, “Hairpin RNA-mediated strategies for silencing of tomato leaf curl virus AC1 and AC4 genes for effective resistance in plants,” Oligonucleotides, vol. 17, no. 2, pp. 251–257, 2007.
[53]
R. . Gorovitsa, A. Moshea, M. Kolotb, I. Sobola, and H. Czosneka, “Progressive aggregation of Tomato yellow leaf curl virus coat protein in systemically infected tomato plants, susceptible and resistant to the virus,” Virus Research. In press.
