The co-infection of Maize chlorotic mottle virus (MCMV) and Sugarcane mosaic virus (SCMV) can cause maize lethal necrosis. However, the mechanism underlying the synergistic interaction between these two viruses remains elusive. In this study, we found that the co-infection of MCMV and SCMV increased the accumulation of MCMV. Moreover, the profiles of virus-derived siRNAs (vsiRNAs) from MCMV and SCMV in single- and co-infected maize plants were obtained by high-throughput sequencing. Our data showed that synergistic infection of MCMV and SCMV increased remarkably the accumulation of vsiRNAs from MCMV, which were mainly 22 and 21 nucleotides in length. The single-nucleotide resolution maps of vsiRNAs revealed that vsiRNAs were almost continuously but heterogeneously distributed throughout MCMV and SCMV genomic RNAs, respectively. Moreover, we predicted and annotated dozens of host transcript genes targeted by vsiRNAs. Our results also showed that maize DCLs and several AGOs RNAs were differentially accumulated in maize plants with different treatments (mock, single or double inoculations), which were associated with the accumulation of vsiRNAs. Our findings suggested possible roles of vsiRNAs in the synergistic interaction of MCMV and SCMV in maize plants.

RNA silencing is a conserved surveillance mechanism against viruses in plants. It is mediated by Dicer-like (DCL) proteins producing small interfering RNAs (siRNAs), which guide specific Argonaute (AGO)-containing complexes to inactivate viral genomes and may promote the silencing of host mRNAs. In this study, we obtained the profile of virus-derived siRNAs (vsiRNAs) from Sugarcane mosaic virus (SCMV) in infected maize (Zea mays L.) plants by deep sequencing. Our data showed that vsiRNAs which derived almost equally from sense and antisense SCMV RNA strands accumulated preferentially as 21- and 22-nucleotide (nt) species and had an adenosine bias at the 5′-terminus. The single-nucleotide resolution maps revealed that vsiRNAs were almost continuously but heterogeneously distributed throughout the SCMV genome and the hotspots of sense and antisense strands were mainly distributed in the HC-Pro coding region. Moreover, dozens of host transcripts targeted by vsiRNAs were predicted, several of which encode putative proteins involved in ribosome biogenesis and in biotic and abiotic stresses. We also found that ZmDCL2 mRNAs were up-regulated in SCMV-infected maize plants, which may be the cause of abundant 22-nt vsiRNAs production. However, ZmDCL4 mRNAs were down-regulated slightly regardless of the most abundant 21-nt vsiRNAs. Our results also showed that SCMV infection induced the accumulation of AGO2 mRNAs, which may indicate a role for AGO2 in antiviral defense. To our knowledge, this is the first report on vsiRNAs in maize plants.

Cucumber green mottle mosaic virus (CGMMV) belongs to the Tobamovirus genus and is a major global plant virus on cucurbit plants. It causes severe disease symptoms on infected watermelon plants (Citrullus lanatus), particularly inducing fruit decay. However, little is known about the molecular mechanism of CGMMV-induced watermelon fruit decay. For this study, comparative analysis of transcriptome profiles of CGMMV-inoculated and mock-inoculated watermelon fruits were conducted via RNA-Seq. A total of 1,621 differently expressed genes (DEGs) were identified in CGMMV-inoculated watermelon, among which 1,052 were up-regulated and 569 were down-regulated. Functional annotation analysis showed that several DEGs were involved in carbohydrate metabolism, hormone biosynthesis and signaling transduction, secondary metabolites biosynthesis, and plant-pathogen interactions. We furthermore found that some DEGs were related to cell wall components and photosynthesis, which may directly be involve in the development of the symptoms associated with diseased watermelons. To confirm the RNA-Seq data, 15 DEGs were selected for gene expression analysis by qRT-PCR. The results showed a strong correlation between these two sets of data. Our study identified many candidate genes for further functional studies during CGMMV-watermelon interactions, and will furthermore help to clarify the understanding of pathogenic mechanism underlying CGMMV infection in cucurbit plants.

Sugarcane mosaic virus (SCMV) is the most important cause of maize dwarf mosaic disease. To identify maize genes responsive to SCMV infection and that may be involved in pathogenesis, a comparative proteomic analysis was performed using the first and second systemically infected leaves (termed 1 SL and 2 SL, respectively). Seventy-one differentially expressed proteins were identified in 1 SL and 2 SL upon SCMV infection. Among them, eight proteins showed the same changing patterns in both 1 SL and 2 SL. Functional annotations of regulated proteins and measurement of photosynthetic activity revealed that photosynthesis was more inhibited and defensive gene expression more pronounced in 1 SL than in 2 SL. Knockdown of regulated proteins in both 1 SL and 2 SL by a brome mosaic virus-based gene silencing vector in maize indicated that protein disulfide isomerase-like and phosphoglycerate kinase were required for optimal SCMV replication. By contrast, knockdown of polyamine oxidase (ZmPAO) significantly increased SCMV accumulation, implying that ZmPAO activity might contribute to resistance or tolerance. The results suggest that combining comparative proteomic analyses of different tissues and virus-induced gene silencing is an efficient way to identify host proteins supporting virus replication or enhancing resistance to virus infection.

Background/Aim: Unilateral ureteral obstruction (UUO) is a well-established model for tubulointerstitial fibrosis. During the progression of tubulointerstitial fibrosis, upregulation of collagen synthesis and subsequent accumulation of collagen were observed in the tubulointerstitial area. Heat shock protein 47 (HSP47) is a collagen-specific molecular chaperone and plays an essential role in regulating collagen synthesis. We designed small interfering RNA (siRNA) sequences for HSP47 mRNA to examine whether HSP47 is involved in the progression of renal tubulointerstitial fibrosis in a mouse UUO model. Methods: The HSP47 siRNA was injected once via the ureter at the time of UUO preparation. We also applied a new gene delivery system for siRNA using cationized gelatin microspheres. The kidneys were harvested 7 and 14 days after UUO. The HSP47 and type I, III, and IV collagen expression levels were analyzed by immunohistochemistry and Western blotting. Results: Seven days after UUO, the expression levels of HSP47 and type I, III, and IV collagens were markedly upregulated in obstructed kidneys or green fluorescent protein siRNA treated obstructed kidneys. HSP47 siRNA injection significantly reduced the protein expression levels and significantly diminished the accompanying interstitial fibrosis. Moreover, cationized gelatin microspheres as a delivery system enhanced and lengthened the antifibrotic effect of HSP47 siRNA. Conclusions: Our results indicate that HSP47 is a candidate target for the prevention of tubulointerstitial fibrosis and that selective blockade of the HSP47 expression by using siRNA could be a potentially useful therapeutic approach for patients with renal disease.