Project description:Posttranscriptional and posttranslational modifications play crucial roles in plant immunity. However, how plant fine-tune these two modifications to activate antiviral immunity remains unknown. Here, we report that the m6A methyltransferase TaHAKAI is utilized by wheat yellow mosaic virus (WYMV) to increase viral genomic m6A modification and promotes viral replication. However, TaHAKAI also functions as an E3 ligase that targets the viral RNA silencing suppressor P2 for degradation and inhibits viral infection. A major allele of TaHAKAI in susceptible cultivar reduced the E3 ligase activity but not m6A methyltransferase activity, promoting viral infection. Interestingly, TaHAKAIR attenuates the mRNA stability of TaWPS1, the negative regulator of spike development, to increase panicle length and spikelet number by modulating its m6A modification. Our study reveals a new mechanisms of balancing disease resistance and yield by fine-tuning m6A modification and ubiquitination.
Project description:au09-04_geminisilsup2 - gemsupv2 - Determine the level of expression in transgenic plants expressing geminviral gene silencing suppressors. One transgenic line of Arabidopsis overexpressing the geminiviral silencing suppressor V2 was grown in addition to a control, and their total RNA was extracted in order to study their genetic expression profiles.
Project description:au10-09_gemsupv2ts-7 - gemsupv2ts-7 - Determine the level of expression in transgenic plants expressing geminiviral gene silencing suppressors. - One transgenic line of Arabidopsis overexpressing the geminiviral silencing suppressor V2TS was grown in adition to a control and their total RNA was extracted in order to study its genetic expression profile.
Project description:The 35S::GFP fluorescence was silenced 6-day after infiltration to Nicotiana benthamiana leaves due to the post-transcriptional gene silencing, but became stable by adding the viral suppressor 2b which repressed RNA silencing in plants. We performed the small RNA high throughput sequencing to test whether WUS can inhibit 2b functions in repressing plant RNA silencing.
Project description:White clover mosaic virus (WCMV) is a major pathogen of white clover (Trifolium repens L.), with significant effects on yield and persistence. Due to the absence of natural sources of WCMV resistance a transgenic strategy has been employed to produce plants constitutively expressing WCMV replicase gene derivatives, designed to inhibit the propagation of WCMV through an RNA silencing mechanism. A 12,000 feature oligonucleotide microarray has been used to identify global changes in host plant, in addition to virus genome-encoded gene expression associated with WCMV infection in non-transgenic and transgenic WCMV-resistant white clover. Pairwise comparison between the transcriptome of mock-inoculated non-transgenic and WCMV-inoculated transgenic plants provides clear evidence for substantial equivalence between these two genotype/treatments, and demonstrate the efficacy of the transgenic strategy. WCMV- inoculated non-transgenic plants exhibit elevated abundance of many virus-encoded, and host immune response-specific transcripts compared to the transgenic resistant plants or mock-inoculated non-transgenic plants. By contrast, relative to inoculated sensitive plants, the majority of significantly up-regulated genes in mock-inoculated non-transgenic plants or WCMV-inoculated transgenic plants are markers of healthy cellular function. These results, and the occurrence of levels of WCMV-encoded transcripts in inoculated transgenic plants equivalent to those in virus-free plants, confirm the validity of the transgenic RNA silencing approach.<br>
Project description:au09-04_geminisilsup2 - gemsupv2 - Determine the level of expression in transgenic plants expressing geminviral gene silencing suppressors. One transgenic line of Arabidopsis overexpressing the geminiviral silencing suppressor V2 was grown in addition to a control, and their total RNA was extracted in order to study their genetic expression profiles. 3 dye-swaps - gene knock-in (transgenic).
Project description:Transgenic tobacco plants expressing begomoviral AC2 RNA silencing suppressor were used to compare transcriptional changes in the transcriptome between transgenic and wild type tobacco plants. Transcriptional analysis using Agilent 4x44k tobacco array was performed of six week-old leaves and 3-5 month-old flowers taken from the same plants as leaf samples.
Project description:RNA silencing is one of the main defense mechanisms employed by plants to fight viruses. In change, viruses have evolved silencing suppressor proteins to neutralize antiviral silencing. Since the endogenous and antiviral functions of RNA silencing pathway rely on common components, it was suggested that viral suppressors interfere with endogenous silencing pathway contributing to viral symptom development. In this work, we aimed to understand the effects of the tombusviral p19 suppressor on endogenous and antiviral silencing during genuine virus infection. We showed that ectopically expressed p19 sequesters endogenous small RNAs (sRNAs) in the absence, but not in the presence of virus infection. Our presented data question the generalized model in which the sequestration of endogenous sRNAs by the viral suppressor contributes to the viral symptom development. We further showed that p19 preferentially binds the perfectly-paired ds-viral small interfering RNAs (vsiRNAs) but does not select based on their sequence or the type of the 5’ nucleotide. Finally, co-immunoprecipitation of sRNAs with AGO1 or AGO2 from virus-infected plants revealed that p19 specifically impairs vsiRNA loading into AGO1 but not AGO2. Our findings, coupled with the fact that p19-expressing wild type Cymbidium ringspot virus (CymRSV) overcomes the Nicotiana benthamiana silencing based defense killing the host, suggest that AGO1 is the main effector of antiviral silencing in this host-virus combination. To further support our hypothesis we investigate whether the ability of p19 to bind endogenous sRNA without virus infection has biological important impact on endogenous pathways and is this reduced if the virus is present. To asses this we made mRNA sequencing from mock inoculated and Cym19stop infected p19syn plants. Cym19stop infected wild type plant was sequenced as a control. The sequencing data results supports our claims. An increase in transcriptional levels were found in case of genes known to be under small RNA regulation in uninfected p19syn plants and expressional levels return to normal Cym19stop p19syn plants.
Project description:au10-09_gemsupv2ts-7 - gemsupv2ts-7 - Determine the level of expression in transgenic plants expressing geminiviral gene silencing suppressors. - One transgenic line of Arabidopsis overexpressing the geminiviral silencing suppressor V2TS was grown in adition to a control and their total RNA was extracted in order to study its genetic expression profile. 3 dye-swap - gene knock in (transgenic)
