Li-Juan Chen

Involvement of PHYB in resistance to Cucumber mosaic virus in Nicotiana tabacum

AbstractPlants are exposed to a diversity of environmental stresses throughout their life cycles and need to constantly regulate their developmental and physiological processes to respond various external stimuli. It is well known that the interaction between light and plant defence is multifaceted and complicated. Here, the recombinant vector pRNAi-LIC-PHYB was used for achieving tobacco (Nicotiana tabacum) PHYB defective mutant via the Agrobacterium-mediated transformation method. Furthermore, we investigated molecular and physiological principles in PHYB defective N. tabacum plants in response to Cucumber mottle virus (CMV) infection. Our results indicated that PHYB defective plants were more susceptible to CMV infection. There were higher viral replication level, more impetuous viral symptoms and more serious stress damages in PHYB defective mutants along the time going when compared with wild-type (wt) plants. Moreover, the transcripts of defence-related genes and activities of antioxidant enzymes in phyB mutants were down-regulated when compared with wt plants after CMV infection. Overall, our results provided new information about the molecular link between PHYB and plant immune system in N. tabacum.

Characterisation of the dark green islands of cucumber mosaic virus infected Nicotiana tabacum

AbstractKey messageThere are significant differences between the DGIs and LGTs. Additionally, most of the characteristics indicate that the DGIs are more similar to recovered tissue and can resist viral attacks.AbstractDark green islands (DGIs) surrounded by light green tissues (LGTs) are common leaf symptoms of plants that are systemically infected by various mosaic viruses. We performed cytological, physiological and molecular biological analyses of the DGIs and LGTs in cucumber mosaic virus-infected Nicotiana tabacum leaves. Our results indicated that the DGIs contained less virus than did the LGTs. Compared to the LGTs, the DGIs contained higher levels of the metabolites involved in plant defence. The contents of reduced glutathione and ascorbic acid were increased in the DGIs to reach levels that were even higher than those of control plants. Moreover, hormone measurements and quantitative real-time PCR analysis revealed that the endogenous salicylic acid, ethylene and defence genes mediated these elevations by playing positive roles in the regulation of the DGIs responses to viral infection. The accumulation of cytokinin was also much greater in the DGIs than in the LGTs. Finally, northern blotting analysis indicated that the accumulation of viral small interfering RNAs was decreased in the DGIs compared to the LGTs. Taken together, these results suggest that DGIs might represent leaf areas that have recovered from viral infection due to locally enhanced defence responses.

The Roles of Alpha-Momorcharin and Jasmonic Acid in Modulating the Response of Momordica charantia to Cucumber Mosaic Virus.

Alpha-momorcharin (α-MMC) is a type-I ribosome inactivating protein with a molecular weight of 29 kDa that is found in Momordica charantia, and has been shown to be effective against a broad range of human viruses as well as having anti-tumor activities. However, the role of endogenous α-MMC under viral infection and the mechanism of the anti-viral activities of α-MMC in plants are still unknown. To study the effect of α-MMC on plant viral defense and how α-MMC increases plant resistance to virus, the M. charantia–cucumber mosaic virus (CMV) interaction system was investigated. The results showed that the α-MMC level was positively correlated with the resistance of M. charantia to CMV. α-MMC treatment could alleviate photosystem damage and enhance the ratio of glutathione/glutathione disulfide in M. charantia under CMV infection. The relationship of α-MMC and defense related phytohormones, and their roles in plant defense were further investigated. α-MMC treatment led to a significant increase of jasmonic acid (JA) and vice versa, while there was no obvious relevance between salicylic acid and α-MMC. In addition, reactive oxygen species (ROS) were induced in α-MMC-pretreated plants, in a similar way to the ROS burst in JA-pretreated plants. The production of ROS in both ibuprofen (JA inhibitor) and (α-MMC+ibuprofen)-pretreated plants was reduced markedly, leading to a greater susceptibility of M. charantia to CMV. Our results indicate that the anti-viral activities of α-MMC in M. charantia may be accomplished through the JA related signaling pathway.

Tobacco alpha-expansin EXPA4 plays a role in Nicotiana benthamiana defence against Tobacco mosaic virus

AbstractMain conclusionTobaccoEXPA4plays a role inNicotiana benthamianadefence against virus attack and affects antioxidative metabolism and phytohormone-mediated immunity responses in tobacco. Expansins are cell wall-loosening proteins known for their endogenous functions in cell wall extensibility during plant growth. The effects of expansins on plant growth, developmental processes and environment stress responses have been well studied. However, the exploration of expansins in plant virus resistance is rarely reported. In the present study, virus-induced gene silencing (VIGS) and Agrobacterium-mediated transient overexpression were conducted to investigate the role of Nicotiana tabacum alpha-expansin 4 (EXPA4) in modulating Tobacco mosaic virus (TMV-GFP) resistance in Nicotiana benthamiana. The results indicated that silencing of EXPA4 reduced the sensitivity of N. benthamiana to TMV-GFP, and EXPA4 overexpression accelerated virus reproduction on tobacco. In addition, our data suggested that the changes of virus accumulation in response to EXPA4 expression levels could further affect the antioxidative metabolism and phytohormone-related pathways in tobacco induced by virus inoculation. EXPA4-silenced plants with TMV-GFP have enhanced antioxidant enzymes activities, which were down-regulated in virus-inoculated 35S:EXPA4 plants. Salicylic acid accumulation and SA-mediated defence genes induced by TMV-GFP were up-regulated in EXPA4-silenced plants, but depressed in 35S:EXPA4 plants. Furthermore, a VIGS approach was used in combination with exogenous phytohormone treatments, suggesting that EXPA4 has different responses to different phytohormones. Taken together, these results suggested that EXPA4 plays a role in tobacco defence against viral pathogens.

The capsid protein p38 of turnip crinkle virus is associated with the suppression of cucumber mosaic virus in Arabidopsis thaliana co-infected with cucumber mosaic virus and turnip crinkle virus

Infection of plants by multiple viruses is common in nature. Cucumber mosaic virus (CMV) and Turnip crinkle virus (TCV) belong to different families, but Arabidopsis thaliana and Nicotiana benthamiana are commonly shared hosts for both viruses. In this study, we found that TCV provides effective resistance to infection by CMV in Arabidopsis plants co-infected by both viruses, and this antagonistic effect is much weaker when the two viruses are inoculated into different leaves of the same plant. However, similar antagonism is not observed in N. benthamiana plants. We further demonstrate that disrupting the RNA silencing-mediated defense of the Arabidopsis host does not affect this antagonism, but capsid protein (CP or p38)-defective mutant TCV loses the ability to repress CMV, suggesting that TCV CP plays an important role in the antagonistic effect of TCV toward CMV in Arabidopsis plants co-infected with both viruses.

Phytochromes play a positive role in Nicotiana tabacum against Cucumber mosaic virus via a salicylic acid-dependent pathway

Previous literature indicates that both light and photoreceptors may play key roles in host susceptibility to viruses. Our previous studies have suggested that phytochromeB is essential for the defense strategies of -Nicotiana tabacum against Cucumber mosaic virus. Here, we further investigated the roles of phytochromes in N. tabacum against CMV. The recombinant vector pRNAi-LIC-PHYA/PHYB was used to produce tobacco phyA and phyAphyB mutants via the Agrobacterium-mediated transformation method. The results suggested that both phyA and phyB deficiency increased the susceptibility of N. tabacum to CMV infection and showed more obvious CMV-induced symptoms and higher transcriptional levels of the CMV-coat protein gene than wild plant, and the effect was clearly synergistic in the phyAphyB double mutant. Furthermore, our data showed that mutations in phyA/B up-regulated the sensitivity of N. tabacum to CMV by repressing the SA-mediated defence pathway in tobacco plants. Additionally, the results in the present study indicated that phytochrome signalling strongly modulates the response of N. tabacum to endogenous SA but not to endogenous JA. Taken together, these results reinforced the idea that an intact phytochrome signalling plays a positive role in the ability of N. tabacum to resist CMV and is required for the induction and development of SA-mediated defence responses in CMV-inoculated N. tabacum. This article is protected by copyright. All rights reserved.

Temperature dependent defence of Nicotiana tabacum against Cucumber mosaic virus and recovery occurs with the formation of dark green islands

Temperature is an important environmental factor controlling plant growth, development, and immune response. However, the role of temperature in plant disease resistance is still elusive. In the present study, the potential effects of temperature on the interaction between Nicotiana tabacum and Cucumber mosaic virus (CMV) were investigated. Our results indicated that N. tabacum plants displayed severe symptoms at early stage of post inoculation at high temperature (HT, 28°C), associated with higher viral replication level, more serious stress damages. By contrast, low temperature (LT, 18°C) effectively delayed the replication of CMV compared with elevated temperatures. Additionally, quantitative real-time PCR analyses revealed that lower temperatures (≤ 24°C) promote salicylic acid (SA) dependent responses, whereas higher temperatures (> 24°C) activate the genes expression of jasmonic acid (JA) pathway. Interestingly, the dark green islands (DGIs) appeared much earlier in CMV-inoculated plants grown at HT compared with those at LT and the accumulation of virus small interfering RNAs in plants were significantly up-regulated under elevated temperatures at early stage of post inoculation. Taken together, these results indicated that temperature changes had important effects on plant defence response, and different temperatures could induce different immune pathways of N. tabacum against CMV infection.

α-Expansin EXPA4 Positively Regulates Abiotic Stress Tolerance but Negatively Regulates Pathogen Resistance in Nicotiana tabacum

Since they function as cell wall-loosening proteins, expansins can affect plant growth, developmental processes and environmental stress responses. Our previous study demonstrated that changes in Nicotiana tabacum α-expansin 4 (EXPA4) expression affect the sensitivity of tobacco to Tobacco mosaic virus [recombinant TMV encoding green fluorescent protein (TMV-GFP)] infection by Agrobacterium-mediated transient expression. In this study, to characterize the function of tobacco EXPA4 further, EXPA4 RNA interfernce (RNAi) mutants and overexpression lines were generated and assayed for their tolerance to abiotic stress and resistance to pathogens. First, the differential phenotypes and histomorphology of transgenic plants with altered EXPA4 expression indicated that EXPA4 is essential for normal tobacco growth and development. By utilizing tobacco EXPA4 mutants with abiotic stress, it was demonstrated that RNAi mutants have increased hypersensitivity to salt and drought stress. In contrast, the overexpression of EXPA4 in tobacco conferred greater tolerance to salt and drought stress, as indicated by less cell damage, higher fresh weight, higher soluble sugar and proline accumulation, and higher expression levels of several stress-responsive genes. In addition, the overexpression lines were more susceptible to the viral pathogen TMV-GFP when compared with the wild type or RNAi mutants. The induction of the antioxidant system, several defense-associated phytohormones and gene expression was down-regulated in overexpression lines but up-regulated in RNAi mutants when compared with the wild type following TMV-GFP infection. In addition, EXPA4 overexpression also accelerated the disease development of Pseudomonas syringae DC3000 on tobacco. Taken together, these results suggested that EXPA4 appears to be important in tobacco growth and responses to abiotic and biotic stress.