Abstract
Secreted small cysteine‐rich proteins (SCPs) play a critical role in modulating host immunity in plant–pathogen interactions. Bioinformatic analyses showed that the fungal pathogen Verticillium dahliae encodes more than 100 VdSCPs, but their roles in host–pathogen interactions have not been fully characterized. Transient expression of 123 VdSCP‐encoding genes in Nicotiana benthamiana identified three candidate genes involved in host–pathogen interactions. The expression of these three proteins, VdSCP27, VdSCP113, and VdSCP126, in N. benthamiana resulted in cell death accompanied by a reactive oxygen species burst, callose deposition, and induction of defence genes. The three VdSCPs mainly localized to the periphery of the cell. BAK1 and SOBIR1 (associated with receptor‐like protein) were required for the immunity triggered by these three VdSCPs in N. benthamiana. Site‐directed mutagenesis showed that cysteine residues that form disulphide bonds are essential for the functioning of VdSCP126, but not VdSCP27 and VdSCP113. VdSCP27, VdSCP113, and VdSCP126 individually are not essential for V. dahliae infection of N. benthamiana and Gossypium hirsutum, although there was a significant reduction of virulence on N. benthamiana and G. hirsutum when inoculated with the VdSCP27/VdSCP126 double deletion strain. These results illustrate that the SCPs play a critical role in the V. dahliae–plant interaction via an intrinsic virulence function and suppress immunity following infection.
Highlights
Some fungi have evolved the ability to cause plant diseases and are a threat to many economically important crops
Three small cysteine-rich proteins (SCPs), VdSCP27, VdSCP113, and VdSCP126, which act as PAMPs, were identified, and BRI1-associated receptor kinase1 (BAK1) and Suppressor of BIR1-1 (SOBIR1) are required for these VdSCPs to induce cell death and trigger immunity in N. benthamiana
To surmount these defence responses, V. dahliae employs other VdSCP effectors to suppress the immunity triggered by VdSCP27, VdSCP113, and VdSCP126
Summary
Some fungi have evolved the ability to cause plant diseases and are a threat to many economically important crops. Asterisks (∗) and double asterisks (∗∗) represent statistical significance at p < .05 and p < .01, respectively, based on unpaired Student's t tests the VdSCP27, VdSCP113 or VdSCP126 constructs into N. benthamiana leaves resulted in a significant reduction of lesion area 48 hr after inoculation with Botrytis cinerea, compared to agroinfiltration of GFP alone (Figure 4f,g) Together, these results suggested that VdSCP27, VdSCP113, and VdSCP126 can each act as elicitors that induce cell death by triggering defence responses during host–pathogen interaction in N. benthamiana. Immunoblotting analysis confirmed that all the mutant alleles of these three VdSCPs were successfully expressed 3 days after infiltration in N. benthamiana leaves (Figure 6b,d,f) These results suggested that cysteine residues are critical for inducing cell death in VdSCP126, but not in VdSCP27 and VdSCP113. These results indicate that VdSCP27 and VdSCP126 act together as virulence factors in N. benthamiana and cotton interactions with V. dahliae
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