ABSTRACT
Three ourmia-like viruses, designated Pyricularia oryzae ourmia-like virus (PoOLV) 1 to 3, were identified in a wheat-infecting isolate of P. oryzae. The sizes of the full-length PoOLV1-3 genomes were determined to be 2,528, 1,671, and 2,557â¯nt. Interestingly, we also found two abundant single-stranded RNAs sharing their 5' terminal 25 and 255â¯nt with PoOLV1 RNA and PoOLV3 RNA, respectively. The PoOLV1- and PoOLV3-associated RNAs (ARNA1 and ARNA3) were 639 and 514â¯nt in length, and possessed one and two small ORFs, respectively. In the field isolates of P. oryzae, PoOLVs and ARNAs were detectable at varying levels, and the levels of PoOLV1 and ARNA1 as well as those of PoOLV3 and ARNA3, were tightly correlated. In addition, gene silencing of PoOLV1 and PoOLV3 resulted in a reduction of ARNA1 and ARNA3, respectively. There results indicated that replication of ARNA1 and ARNA3 was associated with PoOLV1 and PoOLV3, respectively.
Subject(s)
Ascomycota/virology , Fungal Viruses/isolation & purification , Plant Diseases/microbiology , RNA, Viral/metabolism , Fungal Viruses/classification , Fungal Viruses/genetics , Genome, Viral , Open Reading Frames , Phylogeny , RNA, Viral/genetics , Triticum/microbiology , Viral Proteins/genetics , Viral Proteins/metabolismABSTRACT
Small RNA (sRNA)-mediated gene silencing phenomena, exemplified by RNA interference (RNAi), require a unique class of proteins called Argonautes (AGOs). An AGO protein typically forms a protein-sRNA complex that contributes to gene silencing using the loaded sRNA as a specificity determinant. Here, we show that MoAGO2, one of the three AGO genes in the fungus Pyricularia oryzae (Magnaporthe oryzae) interferes with RNAi. Gene knockout (KO) studies revealed that MoAGO1 and MoAGO3 additively or redundantly played roles in hairpin RNA- and retrotransposon (MAGGY)-triggered RNAi while, surprisingly, the KO mutants of MoAGO2 (Δmoago2) showed elevated levels of gene silencing. Consistently, transcript levels of MAGGY and mycoviruses were drastically reduced in Δmoago2, supporting the idea that MoAGO2 impeded RNAi against the parasitic elements. Deep sequencing analysis revealed that repeat- and mycovirus-derived small interfering RNAs were mainly associated with MoAGO2 and MoAGO3, and their populations were very similar based on their size distribution patterns and positional base preference. Site-directed mutagenesis studies indicated that sRNA binding but not slicer activity of MoAGO2 was essential for the ability to diminish the efficacy of RNAi. Overall, these results suggest a possible interplay between distinct sRNA-mediated gene regulation pathways through a competition for sRNA.