The Myosin5‐mediated actomyosin motility system is required for Verticillium pathogenesis of cotton

Abstract

The vascular wilt fungus Verticillium dahliae is one of the most destructive pathogens of cotton (Gossypium hirsutum) and many other economically important dicot plants. Fungal pathogens require Myosin-mediated actomyosin motility system for colonization of their host plants; however, the mechanisms underlying this process have not been fully characterized for V. dahliae. Here, in a knock-out experiment, we characterized the role of VdMyo5, a member of the Myosin V family, before and during infection of cotton and Arabidopsis thaliana. The VdMyo5 deletion mutant (ΔVdmyo5) fungi showed obvious defects in the development of conidia and the polarized elongation of vegetative hyphae, but no inhibition of host root penetration. Overall, the ΔVdmyo5 fungi exhibited dramatically reduced virulence in cotton and Arabidopsis, with almost no colonization in sections of host vascular tissue. We found labelled Myosin5-GFP to be specifically enriched at the hyphal tip, co-localized with FM4-64 labelled Spitzenkörper, which is the vesicle supply centre in filamentous fungi. Comparative secretome analysis revealed that proteins associated with cell wall modification and degradation of reactive oxygen species were significantly altered in mutant strains. Our results indicate that Myosin5 is required for vegetative growth and full virulence, possibly by regulating vesicle transport. The findings provide important insight into the cellular mechanisms of Verticillium pathogenesis.

The Cotton Apoplastic Protein CRR1 Stabilizes Chitinase 28 to Facilitate Defense against the Fungal Pathogen Verticillium dahliae

“…The upstream and downstream genomic sequences of VdSSEP1 and the hygromycin resistance cassette were amplified and fused in order into the pGKO vector via homologous recombination cloning (ClonExpressMultiS One Step Cloning Kit, Vazyme Biotech, C112; Wang et al, 2016;Zhou et al, 2017). A. tumefaciens-mediated transformation was performed as previously described with some modifications (Feng et al, 2018). A. tumefaciens cells (strain AGL1) carrying pGKO-VdSSEP1 were grown in YEP medium, the cultures were then transferred to minimal medium at a 1:100 ratio and grown at 28°C for 24 h. The A. tumefaciens cells were collected and diluted to OD 600 = 0.2 in induction medium (1/2 minimal medium containing 5 mL/L glycerol) supplemented with 200 mM acetosyringone (AS).…”

Verticillium dahliae VdBre1 is required for cotton infection by modulating lipid metabolism and secondary metabolites

“…The Agrobacterium tumefacines ‐mediated transformation‐based gene‐disruption method was used to generate the knockout VdBre1 mutant as previously described (Wang et al ., 2016; Feng et al ., 2018). Transformants were verified by hygromycin B and PCR using the primer set Hyg–F/R.…”

“…Gossypium hirsutum cultivar TM‐1 was used as host to perform pathogenicity assays. Two‐week‐old cotton seedlings were inoculated with conidial suspension (1 × 10 7 cfu/ml) by the unimpaired root‐dip inoculation method (Feng et al ., 2018). The method to calculate the disease index as previously described (Liu et al ., 2014 b).…”

“…Fungal cell walls were stained with 0.5 mg/ml calcofluor white as described previously (Feng et al ., 2018). The image of fungi was taken under a spinning disk confocal microscope (UltraView Vox, PerkinElmer, Beaconsfield, Buckinghamshire, UK).…”

Host-Induced Gene Silencing of an Adenylate Kinase Gene Involved in Fungal Energy Metabolism Improves Plant Resistance to Verticillium dahliae

“…VdMcm1, a MADS-box transcription factor, has multiple biological functions during spore and microsclerotial production, secondary metabolism, and virulence [15]. As a member of the Myosin V family, VdMyo5 may participate in vesicle transport to regulate vegetative growth and is essential for full virulence [16]. Although multiple genes playing a role in signal pathways, development, and nutrition have been reported [17][18][19][20], few have been confirmed as feasible candidates for the control of Vd in plants.…”