Studies On GhBZR3 Gene Involved In Cotton Response To Verticillium Wilt And Heat Stress

Plants are subjected to multiple biotic and abiotic stresses during their growth and development.Disease and high temperature stress are the two key factors,which seriously endanger the growth and development of crops.Among them,verticillium wilt,known as the "cancer of cotton",is closely related to temperature during the onset period,which can seriously cause de-etiolation and wilt of leaves,square and boll shedding and even plant death.The GhBZR3 gene was cloned in upland cotton in the our previous work.The results showed that the GhBZR3 gene participates in the regulation of fiber growth and development of cotton by BR,but its functions in the plant response to adversity stress is still very limited.Here,the inoculation test with LX2-1,a strong pathogen of verticillium wilt,and the high temperature stress treatment with 42℃ were carried out respectively on Col-0 and GhBZR3-overexpressed transgenic Arabidopsis.Meanwhile,the upland cotton genetic standard line TM-1was used as the material to silence the expression of GhBZR3 gene by virus-induced gene silencing(VIGS)method,then subject to inoculation with Verticillium dahliae and high temperature stress treatment respectively.Analysis of phenotype plant death,plant height and cotton fiber elongation were conducted.RT-q PCR was used to detect the expression of lignin-related synthesis genes,heat shock transcription factors(HSFs)and heat shock proteins(HSPs).And the chlorophyll fluorescence-imaging-gas exchange synchronous measurement system was used to analyze the chlorophyll fluorescence parameters of Arabidopsis leaves treated with high temperature.The main results are as follows:1.After inoculation with verticillium wilt,phenotypic analysis results showed that GhBZR3–over pressed Arabidopsis was more susceptible to disease compared with Col.RT-q PCR results showed that the expression of lignin synthesis-related genes At PAL4 and At CCo AOMT1 were significantly reduced in GhBZR3-overexpressed Arabidopsis compared to Col.2.In t cotton TM-1,the results of GhBZR3 silenceing and inoculation test showed that inhibition of GhBZR3 gene expression enhanced the resistance of cotton to verticillium wilt.and RT-q PCR experimental results showed that cotton lignin biosynthesis-related genes Gh F5H1,Gh CAD6 and Gh CCo AOMT1 were significantly increased after silencing GhBZR3 gene,compared with control.3.Under high temperature stress,wilt mortality of GhBZR3-overexpressed Arabidopsis was higher and the heat tolerance of plants was reduced than Col.RT-q PCR results showed that the expression levels of HSFs and HSPs genes HSP90,HSFA2 and DREB2 A in GhBZR3-overexpressed Arabidopsis were significantly lower than those of Col.Meantime,t chlorophyll fluorescence parameters of leaves were also significantly reduced than control.4.Silencing GhBZR3 gene with VIGS can improve cotton survival rate and reduce cotton shoot tip mortality under high temperature stress.5.The gene editing vector was transformed into cotton by Agrobacterium-mediated method,and 3 gene editing cotton lines(CRISPR-BZR3)have been obtained so far.In conclusion,GhBZR3 may play a negative regulatory role in plant response to verticillium wilt by regulating the expression of genes related to plant lignin synthesis.In plant response to high temperature stress,overexpression of GhBZR3 in transgenic Arabidopsis may reduce the transcriptional levels of HSFs and HSPs,to increase plant sensitivity to high temperature,and negatively regulate plant response to high temperature stress.The interaction between GhBZR3 and other regulatory factors and the specific molecular mechanism still need further study.The research results provide very important theoretical and practical significance for the further molecular improvement breeding of cotton resistance for disease and high temperature stress.