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Dicer proteins are mainly responsible for generating small RNAs (sRNAs), which are involved in gene silencing in most eukaryotes. In previous research, two DCL proteins in  Valsa mali , the pathogenic fungus causing apple tree  Valsa  canker, were found associated with both the pathogenicity and generation of sRNAs. In this study, the differential expression of small interfering RNAs (siRNAs) and miRNA-like RNAs (milRNAs) was analyzed based on the deep sequencing of the wild type and  Vm - DCL2  mutant, respectively. Overall, the generation of 40 siRNAs and 18 milRNAs was evidently associated with  Vm - DCL2.  The target genes of milRNAs were then identified using degradome sequencing; according to the prediction results, most candidate targets are related to pathogenicity. Further, expression of  Vm -PC-3p-92107_6 was confirmed in the wild type but not in the  Vm - DCL2  mutant. Moreover, the pathogenicity of  Vm -PC-3p-92107_6 deletion mutants (Δ Vm -PC-3p-92107_6) and the over-expression transformants ( Vm -PC-3p-92107_6-OE) was significantly increased and decreased, respectively. Based on those degradome results, vacuolar protein sorting 10 ( Vm - VPS10 ) was identified as the target of  Vm -PC-3p-92107_6. Co-expression analysis in tobacco leaves further confirmed that  Vm -PC-3p-92107_6 could suppress the expression of  Vm - VPS10 . Meanwhile, the expression levels of  Vm -PC-3p-92107_6 and  Vm - VPS10  displayed divergent trends in Δ Vm -PC-3p-92107_6 and  Vm -PC-3p-92107_6-OE, respectively. Perhaps most importantly, Δ Vm - VPS10  featured a significant reduction in pathogenicity. Taken together, our results indicate that a DCL2-dependent milRNA  Vm -PC-3p-92107_6 plays roles in pathogenicity by regulating the expression of  Vm - VPS10 . This study lays a foundation for the comprehensive analysis of pathogenic mechanisms of  V. mali  and deepens our understanding of the generation and function of fungal sRNA.

A Novel DCL2-Dependent Micro-Like RNA Vm-PC-3p-92107_6 Affects Pathogenicity by Regulating the Expression of Vm-VPS10 in Valsa mali