| Cotton is one of the most important commercial crops all over the world. However, cotton production is influenced by Verticillium wilt every year. Verticillium wilt is caused by Verticillium dahliae, which is a soil-borne fungi caused vascular disease. The quality and yield of cotton will be subjected to serious reduction, even more than30%reduction of production when the outbreak of Verticillium wilt. So Verticillium wilt is called the cancer of cotton. It is a challenge to explore resistance genes, because of lacking cotton germplasm with high resistance to V. dahliae. On the other hand, it is difficult to uncover the molecular mechanism of cotton resistance to V. dahliae, because the race of V. dahliae mutates rapidly in nature. Sea island cotton exhibit more resistance to V. dahliae compared with upland cotton, therefore there may be some resistance genes or resistance mechanisms in sea island cotton. We used sea island cotton ’7124’ with high resistance to V. dahliae as the research materials, and utilized comparative proteomics to discover resistance mechanisms during the interaction of cotton and V. dahliae.By two-dimensional electrophoresis and mass spectrometry analysis, totally188differentially expressed proteins were identified from ’7124’ roots after infection by V. dahliae. In these differentially expressed proteins,47%of them was down regulated. According to gene ontology analysis, these differentially expressed proteins can be divided into a total of17categories. The number of proteins which involved in cell processes, protein metabolic processes or stress responses is more than others. These proteins may participate in the response of cotton to V. dahliae. After cotton infection by V. dahliae, cellular metabolism and physiological processes were influenced, and it may be the response to pathogen invasion.Gossypol is one of cotton-specific secondary metabolites. GbCAD1and GbdHG-6-OMT, two enzymes involved in gossypol metabolism, were found to differentially expressed in our protein profile. GbCADl, a cadinene synthase gene, is a key regulator in gossypol synthesis. After silencing GbCADl by VIGS, the gossypol content of GbCAD1-silenced plants was reduced compared with mock. Meanwhile, the resistance to V. dahliae was significantly reduced after GbCAD1was silenced. So gossypol may affect the cotton resistance to V. dahliae. Some other gossypol metabolism-related genes were found to up regulated in cotton roots after infection by V. dahliae, suggesting that gossypol metabolism was activated in cotton after infection. As a negative regulator of the activity of gossypol, GbdHG-6-OMT encodes desoxyhemigossypol-6-O-methyltransferase, and was down regulated after pathogen infection. By histochemical analysis, gossypol was accumulated in cotton after infection by V. dahliae. It is suggested that the accumulation of gossypol in cotton may enhance the resistance to V. dahliae. Gossypol metabolism-related genes (GbFPS, GbCAD1,WRKY1) were found to up regulated in cotton plants with methyl jasmonate treatment, indicating that the metabolism of gossypol may be regulated by jasmonate signal.In addition to playing an important role in plant growth and development, BR-mediated signal pathway is also reported to involve in plant immunity. Four Gb14-3-3proteins, the homologous genes of which in Arabidopsis were reported to interact with the BIN2-phosphorylated targets in the BZR1protein, were all down-regulated significantly in the protein expression profile. According to RT-PCR analysis, the positive regulated genes (GbBRI1and GbBZR1) in BR signal pathway were up regulated in cotton after infection with V. dahliae, indicating BR signal pathway was activated in infected cotton. Application of exogenous BL or silencing of Gb14-3-3by VIGS can enhance cotton disease resistance to V. dahliae, suggesting that BR signal pathway may contribute to cotton disease resistance. After treated cotton with BL, genes related to JA signal pathway were up regulated using qRT analysis, indicating that the BR signal pathway might be upstream JA signal pathway in cotton, and JA signal pathway might be have crosstalk with BR signal pathway.SA and JA are two very important small molecule hormones in plant disease resistance responses. Some genes located at the junction of the SA and JA signal pathway play important roles in regulating crosstalk of these two hormones. In our study, GbSSI2was isolated from protein profile, which was up regulated in the transcript and protein levels in cotton after infection with V. dahliae. After silencing GbSSI2by VIGS, GbSSI2-silenced plants appeared necrosis on stems and leaves; severe necrosis even caused plant death. After ROS detection, the H2O2was accumulated in the GbSSI2-silenced plants; the SA content was accumulated and SA signal related genes were up regulated; while JA content was decreased and JA signal-related genes were suppressed in the silenced cotton plants. After the leaves of silenced plants infected by V. dahliae, the GbSSI2-silenced leaves were more susceptive to V. dahliae. All the mentions above indicate that GbSSI2may influence the cotton resistance by altering SA-and JA-mediated defense signaling. By RT-PCR analysis, JA signal related genes were up regulated in cotton after infection with V. dahliae, indicating JA signal pathway might contribute to cotton disease resistance to V. dahliae.The molecular mechanism that underlies Verticillium dahliae resistance in cotton is poorly understood at present. Here we characterize HDTF1, which is cloned from transcription expression profile of cotton response to V. dahliae. HDTF1was predicted to encode a nuclear homeodomain transcription factor and HDTF1expression was down-regulated in cotton upon Verticillium dahliae and Botrytis cinerea inoculation. To elucidate the possible involvement of HDTF1in cotton-pathogen interactions, we employed virus-induced gene silencing to generate HDTF1-silenced cotton. Silencing HDTF1significantly enhanced cotton resistance to the fungal pathogens V. dahliae and B. cinerea. In addition, HDTF1silencing caused the accumulation of the phytohormone JA in addition to the activation of a JA-related signal pathway. We could not detect altered SA levels or differential expression in the SA-related genes of HDTFl-silenced plants. Our studies suggest that silencing of HDTF1transcripts in cotton resulted in improved resistance to V. dahliae and B. cinerea, and HDTF1was involved in regulating the JA signaling pathway. |
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