The ABI5-dependent down-regulation of mitochondrial ATP synthase OSCP subunit facilitates apple necrotic mosaic virus infection.

Apple necrotic mosaic virus (ApNMV) is associated with apple mosaic disease in China. However, the mechanisms of ApNMV infection, as well as host defence against the virus, are still poorly understood. Mitochondrial ATP synthase plays a fundamental role in the regulation of plant growth and development. However, mitochondrial ATP synthase function in response to virus infection remains to be defined. In the present study, a yeast two-hybrid (Y2H) screening revealed that the apple mitochondrial ATP synthase oligomycin sensitivity-conferring protein (OSCP) subunit (MdATPO) interacts with ApNMV coat protein (CP). It was further verified that overexpression of MdATPO in Nicotiana benthamiana inhibited viral accumulation. In contrast, silencing of NbATPO facilitated viral accumulation, indicating that ATPO plays a defensive role during ApNMV infection. Further investigation demonstrated that ApNMV infection accelerated abscisic acid (ABA) accumulation, and ABA negatively regulated ATPO transcription, which was related to the ability of ABA insensitive 5 (ABI5) to bind to the ABA-responsive elements (ABREs) of the ATPO promoter. Taken together, our results indicated that transcription factor ABI5 negatively regulated ATPO transcription by directly binding to its promoter, leading to the susceptibility of apple and N. benthamiana to ApNMV infection. The current study facilitates a comprehensive understanding of the intricate responses of the host to ApNMV infection.

The coat protein of the ilarvirus prunus necrotic ringspot virus mediates long-distance movement.

The coat protein (CP) of plant viruses generally has multiple functions involving infection, replication, movement and pathogenicity. Functions of the CP of prunus necrotic ringspot virus (PNRSV), the causal agent of several threatening diseases of Prunus fruit trees, are poorly studied. Previously, we identified a novel virus in apple, apple necrotic mosaic virus (ApNMV), which is phylogenetically related to PNRSV and probably associated with apple mosaic disease in China. Full-length cDNA clones of PNRSV and ApNMV were constructed, and both are infectious in cucumber (Cucumis sativus L.), an experimental host. PNRSV exhibited higher systemic infection efficiency with more severe symptoms than ApNMV. Reassortment analysis of genomic RNA segments 1-3 found that RNA3 of PNRSV could enhance the long-distance movement of an ApNMV chimaera in cucumber, indicating the association of RNA3 of PNRSV with viral long-distance movement. Deletion mutagenesis of the PNRSV CP showed that the basic motif from amino acids 38 to 47 was crucial for the CP to maintain the systemic movement of PNRSV. Moreover, we found that arginine residues 41, 43 and 47 codetermine viral long-distance movement. The findings demonstrate that the CP of PNRSV is required for long-distance movement in cucumber, which expands the functions of ilarvirus CPs in systemic infection. For the first time, we identified involvement of Ilarvirus CP protein during long-distance movement.

Selection and Validation of Reference Genes for Gene Expression Studies Using Quantitative Real-Time PCR in Prunus Necrotic Ringspot Virus-Infected Cucumis sativus.

Several members of the genus Ilarvirus infect fruit trees and are distributed worldwide. Prunus necrotic ringspot virus (PNRSV) is one of the most prevalent viruses, causing significant losses. Cucumissativus can be infected by several ilarviruses, leading to obvious symptoms, including PNRSV, which suggests that cucumbers could be good hosts for the study of the pathogenesis of ilarviruses. Real-time quantitative PCR is an optimal choice for studying gene expression because of its simplicity and its fast and high sensitivity, while its accuracy is highly dependent on the stability of the reference genes. In this study, we assessed the stability of eleven reference genes with geNorm, NormFinder, ΔCt method, BestKeeper, and the ranking software, RefFinder. The results indicated that the combined use of EF1α and F-BOX was the most accurate normalization method. In addition, the host genes AGO1, AGO4, and RDR6 were selected to test the reliability of the reference genes. This study provides useful information for gene expression analysis during PNRSV infection and will facilitate gene expression studies associated with ilarvirus infection.

Identification of Silencing Suppressor Protein Encoded by Strawberry Mottle Virus.

Strawberry mottle virus (SMoV) is associated with strawberry decline disease, causing losses to fruit yield and quality. In this study, using a screening system that enables detection of both local and systemic plant host (RNA silencing) defense responses, we found that Pro2Glu and P28, encoded by SMoV RNA2 genome, functioned to suppress local and systemic RNA silencing triggered by single- but not double-stranded GFP RNA. Subcellular localization assay revealed that both Pro2Glu and P28 were localized to nucleus and cytoplasm. The deletion of 11 amino acid residues at the C-terminus destabilized Pro2Glu protein, and the disruption of two conserved GW motifs deprived Pro2Glu of ability to suppress RNA silencing. Additionally, SMoV Pro2Glu and P28 enhanced the accumulation of potato virus X (PVX) in Nicotiana benthamiana 22 days post-infiltration, and P28 exacerbated significantly the symptoms of PVX. Collectively, these data indicate that the genome of SMoV RNA2 encodes two suppressors of RNA silencing. This is the first identification of a stramovirus suppressor of RNA silencing.

A novel pathogenicity determinant hijacks maize catalase 1 to enhance viral multiplication and infection.

Pathogens have evolved various strategies to overcome host immunity for successful infection. Maize chlorotic mottle virus (MCMV) can cause lethal necrosis in maize (Zea mays) when it coinfects with a virus in the Potyviridae family. However, the MCMV pathogenicity determinant remains largely unknown. Here we show that the P31 protein of MCMV is important for viral accumulation and essential for symptom development. Ectopic expression of P31 using foxtail mosaic virus or potato virus X induced necrosis in systemically infected maize or Nicotiana benthamiana leaves. Maize catalases (CATs) were shown to interact with P31 in yeast and in planta. P31 accumulation was elevated through its interaction with ZmCAT1. P31 attenuated the expression of salicylic acid (SA)-responsive pathogenesis-related (PR) genes by inhibiting catalase activity during MCMV infection. In addition, silencing of ZmCATs using a brome mosaic virus-based gene silencing vector facilitated MCMV RNA and coat protein accumulation. This study reveals an important role for MCMV P31 in counteracting host defence and inducing systemic chlorosis and necrosis. Our results have implications for understanding the mechanisms in defence and counter-defence during infection of plants by various pathogens.