Rhizoviticin is an alphaproteobacterial tailocin that mediates biocontrol of grapevine crown gall disease.

Tailocins are headless phage tail structures that mediate interbacterial antagonism. Although the prototypical tailocins, R- and F-pyocins, in Pseudomonas aeruginosa, and other predominantly R-type tailocins have been studied, their presence in Alphaproteobacteria remains unexplored. Here, we report the first alphaproteobacterial F-type tailocin, named rhizoviticin, as a determinant of the biocontrol activity of Allorhizobium vitis VAR03-1 against crown gall. Rhizoviticin is encoded by a chimeric prophage genome, one providing transcriptional regulators and the other contributing to tail formation and cell lysis, but lacking head formation genes. The rhizoviticin genome retains a nearly intact early phage region containing an integrase remnant and replication-related genes critical for downstream gene transcription, suggesting an ongoing transition of this locus from a prophage to a tailocin-coding region. Rhizoviticin is responsible for the most antagonistic activity in VAR03-1 culture supernatant against pathogenic A. vitis strain, and rhizoviticin deficiency resulted in a significant reduction in the antitumorigenic activity in planta. We identified the rhizoviticin-coding locus in eight additional A. vitis strains from diverse geographical locations, highlighting a unique survival strategy of certain Rhizobiales bacteria in the rhizosphere. These findings advance our understanding of the evolutionary dynamics of tailocins and provide a scientific foundation for employing rhizoviticin-producing strains in plant disease control.

A fungal microRNA-like RNA subverts host immunity and facilitates pathogen infection by silencing two host receptor-like kinase genes.

Small RNAs (sRNAs) play important roles in various biological processes by silencing their corresponding target genes in most eukaryotes. However, cross-kingdom regulation mediated by fungal microRNA-like RNAs (milRNAs) in plant-pathogen interactions is still largely unknown. Using molecular, genetic, histological, and biochemical approaches, we found that the apple tree Valsa canker pathogen Valsa mali milRNA Vm-milR1 could suppress the host immunity by silencing two host receptor-like kinase genes, MdRLKT1 and MdRLKT2. Vm-milR1 was highly induced during V. mali infection. Deletion of Vm-milR1 precursor abolished the generation of Vm-milR1 and reduced the virulence of V. mali. Inoculation of Vm-milR1 deletion mutants induced the host defence responses, including reactive oxygen species (ROS) accumulation, callose deposition, and high expression of defence-related genes. Furthermore, Vm-milR1 was confirmed to be able to suppress the expression of MdRLKT1 and MdRLKT2 in a sequence-specific manner. Moreover, overexpression of either MdRLKT1 or MdRLKT2 enhanced apple resistance to V. mali by activating the host defence responses. Furthermore, knockdown of MdRLKT1 or MdRLKT2 compromised the host resistance to V. mali. Our study revealed that V. mali was equipped with Vm-milR1 as an sRNA effector to silence host receptor-like kinase genes, suppress the host defence responses, and facilitate pathogen infection.

Adaptive regulation of virulence genes by microRNA-like RNAs in Valsa mali.

MicroRNAs play important roles in the regulation of gene expression in plants and animals. However, little information is known about the action mechanism and function of fungal microRNA-like RNAs (milRNAs). In this study, combining deep sequencing, molecular and histological assays, milRNAs and their targets in the phytopathogenic fungus Valsa mali were isolated and identified. A critical milRNA, Vm-milR16, was identified to adaptively regulate the expression of virulence genes. Fourteen isolated milRNAs showed high expression abundance. Based on the assessment of a pathogenicity function of these milRNAs, Vm-milR16 was found to be a critical milRNA in V. mali by regulating sucrose non-fermenting 1 (VmSNF1), 4,5-DOPA dioxygenase extradiol (VmDODA), and a hypothetical protein (VmHy1). During V. mali infection, Vm-milR16 is downregulated, while its targets are upregulated. Overexpression of Vm-milR16, but not mutated Vm-milR16, significantly reduces the expression of targets and virulence of V. mali. Furthermore, deletion of VmSNF1, VmDODA and VmHy1 significantly reduce virulence of V. mali. All three targets seem to be essential for oxidative stress response and VmSNF1 is required for expression of pectinase genes during V. mali-host interaction. Our results demonstrate Vm-milRNAs contributing to the infection of V. mali on apple trees by adaptively regulating virulence genes.