Functional Analysis Of NbH2B And NbFD1 In Potato Virus X Infection

Plant virus is a kind of important plant pathogen,which cause serious disease on crops with huge economic loss.During the process of viral infection,various resistant pathways in plants would be activated to antagonize virus infection and attenuate viral symptom.It is significantly important to investigate the interaction between plants and virus to understand resistant mechanisms in viral infection and offer new strategies to control virus infection.Based on mass spectrometry analysis of proteins pull-down by expressed potato virus X(PVX)p25 in Nicotiana Benthamiana,we found that genes histones H2B and ferredoxin 1(FD1)were involved in the process of virus infectionHistone H2B protein is not only structurally important for chromosomal DNA packaging but is also involved in the regulation of gene expression,including the immune response of plants against pathogens.In this study,we show that the PVX infection resulted in the reduced expression of H2B at both the mRNA and protein level in Nicotiana bentharniana.Tobacco rattle virus(TRV)-based virus-induced gene silencing(VIGS)was then used to down-regulate the expression of H2B in N.benthamiana and tests showed that the titre of TRV was similar in these plants to that in control treated plants.When these H2B-silenced plants were inoculated with PVX,the virus spread more slowly through the plant and there was a lower titre of PVX compared to non-silenced plants Abnormal leaf development and stem necrosis were observed in the H2B-silenced plants,which were alleviated in H2B-silenced NahG transgenic plants suggesting the involvement of salicylic acid(SA)in the production of these symptoms.Indeed,quantitative reverse transcription(qRT)-PCR and liquid chromatography tandem mass spectroscopy(LC-MS)results showed that endogenous SA is increased in H2B-silenced N.benthamiana.Thus,downregulation of H2B induced the accumulation of endogenous SA,which was correlated with stem necrosis and a decreased accumulation of PVX in N.benthamianaThe chloroplast is a primary site of infection by some plant viruses and can undergo profound structural and functional turbulence during the infection process.Mechanism of viral targeting chloroplast protein promoting viral infection remains to be further studied.Here,we report that the PVX p25 protein interacted with the chloroplast protein FD1,whose mRNA and protein levels were reduced by PVX infection or by transient expression of p25.Silencing of FD1 by TRV-based VIGS promoted the local and systemic infection of plants by PVX.Use of a drop-and-see assay(DANS)and Callose staining revealed that the permeability of PD was increased in FD1-silenced plants together with a consistently low level of PD callose deposition.After FD1 silencing,qRT-PCR analysis and LC-MS revealed these plants to have a low accumulation of the phytohormones ABA and SA,which contributed to the decreased callose deposition at PD.Addition of exogenous ABA and SA overcame the reduction in deposition of callose at PD in FD1-silenced plants Overexpression of FD1 in transgenic plants manifested resistance to PVX infection,but the contents of ABA,SA and the PD callose deposition were not increased in transgenic plants.The interference of overexpressed FD1 to suppressor function of p25 probably caused the alleviated infection of PVX in FD1 overexpressed plants.All in all,by silencing or overexpressing FD1 in N.benthamiana induced multiple responses to infection by PVXTaken together,our study revealed that histone H2B participates in the process of plant resistance to PVX infection through negatively regulating SA pathway,and FD1 participates in the process of plant virus infection by regulating multiple responsesn in N.benthamiana to PVX infection.This study provided us a further insight to understand the mechanism of plant-virus interaction.