Efeito do silenciamento dos genes DnaJ e TCTP na infecção de tomateiro e Nicotiana benthamiana pelo potyvírus Pepper yellow mosaic virus (PepYMV)

• Autor(a) principal: Xavier, André da Silva
• Data de Publicação: 2012
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/4413
• topic: Doença; Infecção viral; Plantas não-transformadas; Plantas transgênicas; Disease; Viral infection; Non-transformed plants; Transgenic plants; CNPQ::CIENCIAS AGRARIAS::AGRONOMIA::FITOSSANIDADE::FITOPATOLOGIA

During coevolution, plant viruses have developed the ability to modulate the expression of several host genes, or to alter the function of cognate protein to succeed in their multiplication and perpetuation. These virus-induced changes might lead to a high level of dependency, creating an indissoluble link between virus and host. The objective of this study was to investigate the contribution of a DnaJ of Nicotiana benthamiana (homologous of the tomato protein) and of the TCTP protein from tomato during PepYMV infection. These two genes were identified as differentially expressed in a cDNA library constructed from tomato plants infected by PepYMV. Kinetic studies of DnaJ expression in PepYMV-infected N. benthamiana demonstrated that the induction occurs at both 72 hours post-inoculation (hpi) and 14 days post-inoculation (dpi). Plants of N. benthamiana silenced for DnaJ by means of VIGS and tomato plants cv. Moneymaker silenced by transgenesis for TCTP were obtained and mechanically inoculated with PepYMV. Viral infection was confirmed by ELISA and viral load determined by qRT-PCR. Silencing of the DnaJ gene in N. benthamiana interfered with the early stages of viral infection (72 hpi) but its effect on established infections (14dpi) was inconclusive. Non-transformed tomato plants showed severe symptoms of the disease, while TCTP-silenced transgenic plants showed greatly attenuated symptoms or remained asymptomatic. Viral load was dramatically reduced in silenced plants. The subcellular localization of a TCTP-GFP fusion protein in healthy or PepYMV-infected N. benthamiana plants was analyzed by confocal microscopy. In healthy plants TCTP was nuclear and cytoplasmic, while in infected plants at 14 dpi, its subcellular localization was exclusively cytoplasmic. Together, these results suggest that both TCTP and DnaJ are proteins which positively regulate the infection cycle of PepYMV, being required for disease development in the case of TCTP or for the rapid establishment of viral infection in the case of DnaJ. Further studies should be conducted in order to unravel the mechanisms by which these host factors are used to benefit the viral infection.