Project description:The objective of this study was on the one hand to compare the transcriptional dynamics of mutualistic Colletotrichum tofieldiae and pathogenic Colletotrichum incanum during colonization of Arabidopsis roots and on the other hand also examine the corresponding host responses Arabidopsis seeds that were either untreated (mock) or inoculated with Colletotrichum tofieldiae (isolate 0861) or Colletotrichum incanum (isolate MAFF230704) were grown in normal (P+) or low (P-) phosphate conditions. Roots (with/without fungal colonization) were collected at 6, 10, 16, and 24 days post inoculation (dpi). Additionally, C. tofieldiae and C. incanum in vitro hyphae were collected after two days growth in liquid Mathurâs medium. For each condition three replicates were obtained.
Project description:Here we describe the identification and regulation of a novel dsRNA virus in Colletotrichum higginsianum. High throughput sequencing of small RNAs and strand-specific RNA-seq was performed on single gene knock-out mutants created for each RNAi component gene: rdr1, rdr2, rdr3, dcl1, dcl2, ago1, and ago2, and the double mutant: ∆dcl1∆dcl2. De novo assembly of the ∆dcl1 RNA-seq data identified two contigs that represented the forward and reverse strands of an uncharacterized dsRNA virus, designated here as Colletotrichum higginsianum non-segmented dsRNA virus 1 (ChNRV1). We found increased presence of the viral RNA in the RNA-seq datasets of the ∆dcl1, ∆dcl1dcl2, and ∆ago1 strains, suggesting that these genes are required for control of the virus. We show that viral small RNAs co-immunoprecipitate with a 6xFLAG-3xHIS-tagged AGO1 protein by sequencing the small RNAs from immunoprecipitated fractions. Additionally, analyses of the small RNA datasets from the RNAi mutants revealed control of the virus through small RNA-mediated silencing required both AGO1 and DCL1.