| Cotton(Gossypium hirsutum) is one of the most important economic crops in China. However, the yield and quality of cotton are often affected by various stresses, such as drought, low temperature, high salinity and pathogen attack. In the process of long-term evolution, plants have developed a series of sophisticated defense mechanisms to deal with diverse unfavorable environmental stresses. And transcription factors play very important roles in these processes. WRKY transcription factors as unique to plants are widely involved in various biological processes, such as growth, development, senescence, metabolism, biotic and abiotic stress responses. However, in contrast to well-characterized group ? and ? WRKY proteins, the expression patterns and functions of group ? WRKY transcription factors in abiotic and biotic stress responses are poorly understood in cotton. In this study, a group ? WRKY gene from cotton, GhWRKY27 a, was isolated. And its expression patterns and functions in abiotic and biotic stress responses were characterized. The main results were as follows:(1) The full-length cDNA of the GhWRKY27 a sequence consisted of 1513 nucleotides, including a 1068-bp open reading frame(ORF), a 319-bp 5'-untranslated region(5'-UTR) and a 126-bp 3'-UTR. The ORF encoded a protein composed of 356 amino acid residues with a predicted molecular mass and isoelectric point of 40.062 kDa and 5.46, respectively. Amino acid sequence alignment and a phylogenetic analysis derivated by GhWRKY27 a showed that GhWRKY27 a is a member of group ? WRKY proteins. In addition, subcellular localization indicated that the GhWRKY27 a protein localized to the nucleus.(2) Utilizing PlantCARE and PLACE databases to analyse the promoter sequence of GhWRKY27 a, many cis-elements related to the stress responses were found. The expression pattern analysis indicated that GhWRKY27 a expression could be induced by various abiotic stesses, such as drought, high salinity, low temperature; diverse signalling molecules, such as hydrogen peroxide(H2O2), abscisic acid(ABA), salicylic acid(SA), methyl jasmonate(MeJA), ethylene(ET); and the fungal pathogen Rhizoctonia solani(R. solani).(3) Using virus-induced gene silencing technique, cotton GhWRKY27a-silencing plants were obtained. Some physiological parameters related to drought stress resistance indicated that silencing GhWRKY27 a enhanced drought tolerance in cotton. Using Agrobacterium-mediated transformation method, GhWRKY27a-overexpressing plants were obtained. GhWRKY27 a overexpression in Nicotiana benthamiana markedly enhanced plant sensitivity to drought stress, and the transgenic plants had a higher water loss rate. These results indicated that Gh WRKY27 a is a negative regulator of tolerance to drought stress. Moreover, we further found that GhWRKY27a-overexpressing plants exhibited reduced accumulation of ABA under drought condition, and stomatal closing was insensitive to ABA.(4) GhWRKY27a-overexpressing plants exhibited reduced resistance to R. solani infection, mainly demonstrated by the transgenic lines exhibiting more severe disease symptoms, accompanied by attenuated expression of defense-related genes. These findings indicated that GhWRKY27 a may play a negative role in the response to R. solani infection.(5) We found that GhWRKY27a-overexpressing plants accumulated more reactive oxygen species(ROS) and exhibited decreased expression of stress-related genes after drought treatment and inoculation with R. solani. Moreover, GhWRKY27a-overexpressing plants showed reduced tolerance to oxidative stress. These observations demonstrated that overexpression of GhWRKY27 a in transgenic plants enhance stress sensitivity by affecting ROS scavenging system. |
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