Cotton Employs Multiple Signal Pathways for Resistance to V. dahliae

Plants evolve effective mechanisms to protect themselves against multiple stresses and employ jasmonates (JA) as vital defense signals to defend against pathogen infection. Accumulation of JA induced by signals from biotic and abiotic stresses, results in degradation of Jasmonate-ZIM-Domain (JAZ) proteins, and then de-repressed the JAZ-repressed transcription factors (such as MYC2) to activate defense responses and developmental processes. Here, we characterized a JAZ family protein, GhJAZ2, from cotton (Gossypium hirsutum) which was induced by methyl jasmonate (MeJA) and inoculation of Verticillium dahliae. Over-expression of GhJAZ2 in cotton impairs the sensitivity to JA, decreases the expression level of JA-response genes (GhPDF1.2 and GhVSP) and enhances more susceptibility to V. dahliae and insect herbivory. Yeast two-hybrid and BiFC assays showed that GhJAZ2 may be involved in regulation of cotton disease resistance by interacting with more disease response proteins, like pathogenesis-related protein GhPR10, dirigent-like protein GhD2, NBS-LRR disease resistant protein GhR1, and a basic helix-loop-helix transcription factor bHLH171. Unlike MYC2, over-expression of bHLH171 in cotton activates the JA synthesis and signaling pathway, and improves plant tolerance to fungus V. dahliae. Molecular and genetic evidences showed that GhJAZ2 could interact with bHLH171 and inhibit its transcriptional activity, as a result, restrain the JA-mediated defense response. Recent studies have revealed that the SUPPRESSOR OF BIR1-1 (SOBIR1) can interact with multiple receptor-like proteins (RLPs) and is required for resistance against fungal pathogens. We also find that GbSOBIR1 gene could be induced by Verticillium dahliae inoculation. Knock-down of GbSOBIR1 resulted in attenuated resistance of cotton plants to V. dahliae, while heterologous overexpression of GbSOBIR1 in Arabidopsis improves the resistance. We also found that the kinase region of GbSOBIR1 could interact with bHLH171 and contributes to the resistance of cotton against V. dahliae. And the transcriptional activity of bHLH171 is significantly improved when co-expressed with GbSOBIR1 in tobacco. To identify the GbSOBIR1-mediated phosphorylation site of bHLH171, a spectrometric analysis was performed, and phosphorylation defective forms of the bHLH171 protein with serine to alanine mutations were assayed. The results showed that phosphorylation at Ser413 is essential for the physiological function of bHLH171. Collectively, these results demonstrate that multiple signal pathways are employed by cotton during cotton resistance to V. dahliae.