Project description:<p>Rust fungi are plant pathogens that cause epidemics which threatens the production of many important plant species, such as wheat, soy, coffee and poplar. Melampsora larici-populina (Mlp) causes the poplar rust and encodes at least 1184 candidate effectors (CEs), however their functions are poorly known. In this study, we analysed the transcriptome and metabolome of Arabidopsis constitutively expressing CEs of Mlp to discover processes targeted by these fungal proteins. For this purpose, we sequenced the transcriptome and used mass spectrometry to analyse the metabolome of Arabidopsis plants expressing one of 14 selected CEs and of a control line. We found 2299 deregulated genes in at least one of the 14 transgenic lines. Among the down-regulated genes, the KEGG pathways “MAPK signaling pathway” and “Plant-pathogen interaction” were over-represented in six and five of the 14 transgenic lines, respectively. Moreover, the genes down-regulated across the fourteen transgenic lines are related to hormone response and defense. </p><p>Regarding the metabolome, there were 680 metabolites deregulated across the 14 transgenic lines, with highly unsaturated and phenolic compounds and peptides enriched among down-regulated and up-regulated metabolites, respectively, in almost all transgenic lines. Interestingly, we found that some transgenic lines expressing CEs with no similarity in amino acid sequence had similar patterns of gene and metabolite deregulation, while plants expressing CEs from the same family deregulated different genes and metabolites. Taken together, our results indicate that the sequence of effectors may not be a good predictor of its impact in the plant.</p>
Project description:Small RNAs play essential regulatory roles in genome stability, development and stress responses in most eukaryotes. Plants encode DICER-LIKE (DCL) RNaseIII enzymes, including DCL1, which produces miRNAs, and DCL2, DCL3 and DCL4, which produce diverse size classes of siRNA. Plants also encode RNASE THREE-LIKE (RTL) enzymes that lack DCL-specific domains and whose function is largely unknown. Small RNA sequencing in plants over-expressing RTL1 or RTL2 or lacking RTL2 revealed that RTL1 over-expression inhibits the accumulation of all types of small RNAs produced by DCL2, DCL3 and DCL4, indicating that RTL1 is a general suppressor of plant siRNA pathways. By contrast, RTL2 plays minor, if any, role in the small RNA repertoire. In vivo and in vitro assays revealed that RTL1 prevents siRNA production by degrading dsRNA before they are processed by DCL2, DCL3 and DCL4. The substrate of RTL1 cleavage is likely long perfect (or near-perfect) dsRNA, consistent with the RTL1-insensitivity of miRNAs, which derive from short imperfect dsRNA. RTL1 is naturally expressed only weakly in roots, but virus infection strongly induces its expression in leaves, suggesting that RTL1 induction is a general strategy used by viruses to counteract the siRNA-based plant antiviral defense. Accordingly, transgenic plants over-expressing RTL1 are more sensitive to TYMV infection than wild-type plants, likely because RTL1 prevents the production of antiviral siRNAs. However, TCV, TVCV and CMV, which encode stronger suppressors of RNA silencing (VSR) than TYMV, are insensitive to RTL1 over-expression. Indeed, TCV VSR inhibits RTL1 activity, suggesting that inducing RTL1 expression and dampening RTL1 activity is a dual strategy used by viruses to establish a successful infection. These results reveal another level of complexity in the evolutionary battle between viruses and plant defenses.
Project description:We used Illumina-based RNA-seq to identify differentially expressed genes in wild-type Arbaidopsis or in transgenic plants over-expressing HSFA1b-RFP. These data were interescted with ChIP-seq experiments conducted under the same conditions. The aim of the combined analyses was to identify HSFA1b-regulated genes involved in growth to stress transitions.
Project description:A novel cold-inducible GSK3/Shaggy-like kinase cDNA (TaSK5) was isolated from winter wheat by a macroarray-based differential screening approach. Sequence analysis of TaSK5 revealed high similarity to Arabidopsis subgroup I GSK3/Shaggy-like kinases ASK-alpha, ASK-gamma and ASK-epsilon. Transgenic Arabidopsis plants overexpressing TaSK5 cDNA under the control of CaMV 35S promoter showed enhanced tolerance to salt and drought stresses. In contrast, the tolerance of the transgenic plants to freezing stress was not altered. To identify genes which are differentially regulated in the 35S:TaSK5 over-expressing Arabidopsis plants under non-stress conditions, we compared the genome-wide expression profiles of Col-0 and plants over-expressing TaSK5 using DNA microarrays. Sixty seven genes were found to be expressed at least 2-fold more strongly in 35S:TaSK5 plants than in Col-0, and 17 genes were found to be expressed at least 2-fold more strongly in Col-0 than in 35S:TaSK5 plants. Most of the TaSK5 up-regulated genes were also induced by abiotic stresses, including cold, salt and drought. These results support the involvement of TaSK5 in abiotic stress signal transduction. Keywords: transgenic vs wt Col.-0 comparison Total RNA was extracted from rosette leaves of two independent 3-week-old T1 Arabidopsis Col-0 plants over-expressing TaSK5 driven by 35S promoter and the whole transcriptome was compared with that of wild-type plant. Sample pairs in two independent transformants (OX7 and OX17), referred as biological replicates, were analyzed by the two-color method. Dye-swapped hybridizations were performed in every replicate.
Project description:To obtain better insight into the underlying mechanism of melatonin in switchgrass growth, flowering and defence, we performed proteomics analyses between transgenic lines overexpressing melatonin biosynthesis genes and transgenic control (expressing only the empty vector) plants.