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Accumulating evidence indicates that plant cell wall-associated receptor-like kinases (WAKs) involve in defense against pathogen attack, but their related signaling processes and regulatory mechanism remain largely unknown. We identified a WAK-like kinase (GhWAKL) from upland cotton (Gossypium hirsutum) and characterized its functional mechanism. Expression of GhWAKL in cotton plants was induced by Verticillium dahliae infection and responded to the application of salicylic acid (SA). Knockdown of GhWAKL expression results in the reduction of SA content and suppresses the SA-mediated defense response, enhancing cotton plants susceptibility to V. dahliae. And, ecotopic overexpression of GhWAKL in Arabidopsis thaliana conferred plant resistance to the pathogen. Further analysis demonstrated that GhWAKL interacted with a cotton DnaJ protein (GhDNAJ1) on the cell membrane. Silencing GhDNAJ1 also enhanced cotton susceptibility to V. dahliae. Moreover, the mutation of GhWAKL at site Ser628 with the phosphorylation decreased the interaction with GhDNAJ1 and compromised the plant resistance to V. dahliae. We propose that GhWAKL is a potential molecular target for improving resistance to Verticillium wilt in cotton.

Verticillium wilt is a plant vascular disease caused by the soilborne fungus Verticillium dahliae that severely limits cotton production. In a previous study, we screened Bacillus cereus YUPP‐10, an efficient antagonistic bacterium, to uncover mechanisms for controlling verticillium wilt. Here, we report a novel antimicrobial cyclodextrin glycosyltransferase (CGTase) from YUPP‐10. Compared to other CGTases, six different conserved domains were identified, and six mutants were constructed by gene splicing with overlap extension PCR. Functional analysis showed that domain D was important for hydrolysis activity and domains A1 and C were important for inducing disease resistance. Direct effects of recombinant CGTase on V. dahliae included reduced mycelial growth, spore germination, spore production, and microsclerotia germination. In addition, CGTase also elicited cotton's innate defence reactions. Transgenic Arabidopsis thaliana lines that overexpress CGTase showed higher resistance to verticillium wilt. Transgenic CGTase A. thaliana plants grew faster and resisted disease better. CGTase overexpression enabled a burst of reactive oxygen species production and activated pathogenesis‐related gene expression, indicating that the transgenic cotton was better prepared to protect itself from infection. Our work revealed that CGTase could inhibit the growth of V. dahliae, activate innate immunity, and play a major role in the biocontrol of fungal pathogens., Cyclodextrin glycosyltransferase, a hydrolytic protease from Bacillus cereus YUPP‐10, inhibits the growth of Verticillium dahliae and activates cotton innate immunity. The domains for hydrolysis activity and disease resistance induction were identified.

Background Verticillium wilt, caused by the soil-borne fungus of Verticillium dahliae Kleb., is one of the most devastating diseases of cotton. The complex mechanism underlying cotton resistance to Verticillium wilt remains uncharacterized. Identifying an endogenous resistance gene may be helpful to control this disease. Previous studies revealed that succinate dehydrogenase (SDH) is involved in reactive oxygen species (ROS)-induced stress signaling pathway that is likely to be triggered by salicylic acid (SA). Here, through the metabolomics and differential expression analyses in wilt-inoculated cotton (Gossypium hirsutum), we noticed that GhSDH1–1 gene in cotton may play an important role in the resistance to V. dahlia. Then we reported GhSDH1–1 gene and its functional analysis in relation to the resistance of cotton to V. dahliae. Results The GhSDH1–1 gene in cotton root was significantly up-regulated after V. dahlia inoculation, and its expression level peaked at 12 and 24 h post-infection. SA can also induce the up-regulation of GhSDH1–1. Additionally, the functional analysis showed that GhSDH1–1-silenced cotton was more susceptible to V. dahliae than the control because of the significant decrease in abundance of immune-related molecules and severe damage to the SA-signaling pathway. In Arabidopsis thaliana, high expression of GhSDH1–1 conferred high resistance to V. dahliae. Arabidopsis that overexpressed GhSDH1–1 had higher resistance to V. dahliae infection compared with the wild-type. Conclusions Our findings provide new insights into the role of GhSDH1–1; it positively regulates cotton resistance to Verticillium wilt. The regulatory mechanism of GhSDH1–1 is closely related to SA-related signaling pathway.

Verticillium wilt, mainly caused by a soil-inhabiting fungus Verticillium dahliae, can seriously reduce the yield and quality of cotton. The complex mechanism underlying cotton resistance to Verticillium wilt remains largely unknown. In plants, reactive oxygen species (ROS) mediated by Rbohs is one of the earliest responses of plants to biotic and abiotic stresses. In our previous study, we performed a time-course phospho-proteomic analysis of roots of resistant and susceptible cotton varieties in response to V. dahliae, and found early differentially expressed protein burst oxidase homolog protein D (GhRbohD). However, the role of GhRbohD-mediated ROS in cotton defense against V. dahliae needs further investigation. In this study, we analyzed the function of GhRbohD-mediated resistance of cotton against V. dahliae in vitro and in vivo. Bioinformatics analysis showed that GhRbohD possessed the conservative structural attributes of Rbohs family, 12 members of RbohD out of 57 Rbohs in cotton. The expression of GhRbohD was significantly upregulated after V. dahliae inoculation, peaking at 6 hpi, and the phosphorylation level was also increased. A VIGS test demonstrated that ROS production, NO, H2O2 and Ca2+ contents of GhRbohD-silenced cotton plants were significantly reduced, and lignin synthesis and callose accumulation were damaged, important reasons for the impairment of GhRbohD-silenced cotton’s defense against V. dahliae. The expression levels of resistance-related genes were downregulated in GhRbohD-silenced cotton by qRT-PCR, mainly involving the lignin metabolism pathway and the jasmonic acid signaling pathway. However, overexpression of GhRbohD enhanced resistance of transgenic Arabidopsis to V. dahliae challenge. Furthermore, Y2H assays were applied to find that GhPBL9 and GhRPL12C may interact with GhRbohD. These results strongly support that GhRbohD activates ROS production to positively regulate the resistance of plants against V. dahliae.

Background: Understanding the extent of heritability of plant-associated microbiome (phytobiome) is critical for exploitation of phytobiome in agriculture. Two crosses were made between pairs of cotton cultivars with differential resistance to the Verticillium wilt. F2 plants were grown together with the four parents to study the heritability of cotton rhizosphere microbiome. Amplicon sequencing was used to profile bacterial and fungal communities.Results: F2 hybrid offspring of both crosses had higher alpha diversity indices as well as greater variability than the two parents. Two types of data were used generated to study the heritability of rhizosphere microbiome: principal components (PCs) and individual top microbial OTUs. For the L1×Z49 cross, the variance among the 100 progeny genotypes (VT) was significantly greater than the random variability (VE) for 12 and 34 out of top 100 fungal and bacterial PCs, respectively. For the Z2×J11 cross, the corresponding values were 10 and 20 PCs. Out of the top 100 OTUs, 29 fungal OTUs and 10 bacterial OTUs had VT that was significantly greater than VE for the L1×Z49 cross; the corresponding value for the Z2×J11 cross were 24 and one. The estimated heritability was mostly in the range of 40% to 60%.Conclusions: These results suggested the existence of genetic control in terms of rhizosphere microbiome in cotton.