棉花多逆境响应基因的挖掘和功能验证
发布时间：2018-11-07 06:56
【摘要】：植物通过复杂而精细的调控网络感知外界的各种环境变化,激活相关信号路径,抵御逆境胁迫,但同时往往伴随着生长发育进程的改变并影响产量。由于基因调控网络的复杂性,部分基因位于多个信号路径的交叉点形成枢纽基因,并在多个信号转导中具有不同的功能从而形成一因多效。系统研究多效基因在各种逆境胁迫应答中的具体功能及其对生长发育的影响对于未来鉴定作物广谱抗逆调控因子具有重要作用。本研究通过生物信息学相关手段筛选出一批棉花响应多逆境的重要候选基因,并对其中三个基因进行了功能鉴定。取得的主要结果如下:1.利用STIFDB和GENEVESTIGATOR V3等数据库,筛选出826个拟南芥广谱响应逆境的调节因子。通过同源序列法进一步分离筛选出38个棉花广谱响应逆境的候选基因。对该38个棉花候选基因在盐胁迫、冷胁迫以及ABA处理下的表达模式进行了分析,有37个基因至少对一种处理有响应。为了进一步验证该策略的可靠性,我们选择其中三个基因(GhATAF1、GhJAZ2、GhHB12)进行了系统的功能研究。2.研究发现GhATAF1编码一个有自激活活性的NAC转录因子,不仅受到ABA、盐胁迫和冷胁迫等非生物胁迫的诱导表达,还强烈受到MeJA、SA和棉花黄萎病菌V991的诱导表达,正调控棉花的耐盐性,同时负调控棉花对黄萎病菌的抗性。GhATAF1能通过激活棉花钾离子/钠离子共转运子GhHKT1的表达,同时还能激活ABA信号关键应答基因GhABI4,液泡氢离子-焦磷酸磷酸酶GhAVP1,晚期胚胎丰富蛋白GhLEA3,GhLEA6,干旱应答基因GhRD22和GhDREB2A的表达,减少棉花组织钠离子的积累,减轻细胞的损伤,从而提高棉花的耐盐性;此外我们发现GhATAF1能够激活棉花水杨酸信号路径,同时抑制茉莉酸信号路径,最终削弱棉花对大丽轮枝菌和灰霉菌的抗性。3.研究发现GhJAZ2基因同时受到ABA、MeJA、SA、盐胁迫、冷胁迫以及大丽轮枝菌V991的诱导上调表达,是一个重要的调控棉花防御反应和生长发育的节点基因。GhJAZ2作为一个棉花茉莉酸信号的抑制子,通过与GhJAZ12互作,形成异源二聚体,直接抑制GhMYC2-like和GhPR10,GhR1,GhD2等抗性相关蛋白,负调控棉花抗病性和抗虫性;可能通过与DELLA蛋白的互作,间接激活赤霉素信号路径,调控棉花花期、种子萌发和果枝夹角等生长发育过程。此外GhJAZ2还参与了高温胁迫和低温胁迫下棉花雄蕊的发育。4.研究发现GhHB12基因同时受到ABA、MeJA、SA、盐胁迫、冷胁迫以及大丽轮枝菌V991的诱导上调表达,是一个重要的调控棉花防御反应和生长发育的多效基因。GhHB12负调控植物ABA信号路径,负调控植物对非生物逆境的抗性;同时负调控棉花JA信号路径,削弱棉花的抗病性和抗虫性;此外GhHB12通过与GhTCP1互作,协同负调控生长素信号路径,抑制棉花株高,促进叶枝的发生;同时可以与GhSPLs互作,相互拮抗,直接调控GhFT、GhSOC1、GhFUL等开花基因的表达,调控棉花花期、叶枝和果枝的发育。此外GhHB12还参与对棉花叶形的调控。我们还发现GhHB12受到光周期和节律的调控,GhHB12调控棉花株型的功能在短日照条件下会丧失。以上结果表明利用生物信息学分析能提高鉴定棉花响应多重逆境关键调控因子的有效性,以上结果为棉花响应多重逆境反应的生理机制提供了参考。
[Abstract]:The plant can sense various environmental changes of the outside through the complex and fine control network, activate the relevant signal path, resist the stress of adversity, but at the same time, it is often accompanied by the change of the growth and development process and the yield. Due to the complexity of the gene regulation network, some genes are located at the intersection of multiple signal paths to form the hub gene and have different functions in multiple signal transduction to form a multi-effect. The specific function of the multi-effect gene in various stress-stress responses and its influence on the growth and development of the system have an important role in the future identification of crop broad-spectrum anti-reverse regulation factors. In this study, a number of important candidate genes in response to multiple adversity were selected by means of bioinformatics, and the three genes were identified. The main results are as follows: 1. By using the database of STIFDB and GENVESTIGATOR V3, the regulation factors of the broad-spectrum response of 826 arabidopsis thaliana were selected. and the candidate genes of the 38 cotton broad-spectrum response adversity are further separated and screened by the homologous sequence method. The expression patterns of 38 cotton candidate genes under salt stress, cold stress and ABA treatment were analyzed. To further verify the reliability of the strategy, we selected three of the genes (GhATAF1, GJAK2, GHB12) to perform the functional study of the system. It is found that GhATAF1 encodes a NAC transcription factor with self-activation activity, not only is induced by abiotic stress such as ABA, salt stress and cold stress, but also is strongly influenced by the induced expression of MeJA, SA and Verticillium dahliae V991, and the salt tolerance of cotton is being regulated. and meanwhile, the resistance of the cotton to the Verticillium dahliae is negatively regulated. The GhATAF1 can activate the expression of the cotton potassium ion/ sodium ion cotransporter GhHKT1, and can also activate the expression of the ABA signal key response gene GhABI4, the vacuole hydrogen ion-pyrophosphatase GhAVP1, the late embryo-rich protein GhLEA3, GhLEA6, the drought response gene GhR22 and the GhDREB2A, the accumulation of sodium ions in the cotton tissue is reduced, the damage of the cells is reduced, and the salt tolerance of the cotton is improved; in addition, the GhATAF1 is found to be capable of activating the path of the cotton salicylic acid signal, and meanwhile, the signal path of the jasmonic acid is inhibited, and the resistance of the cotton to the large-lily and the gray mold is finally weakened. It was found that the GhJAK2 gene was induced up-regulated by ABA, MeJA, SA, salt stress, cold stress and V991, which is an important node gene for regulating and controlling the defense response and growth of cotton. GhJAK2 is used as a suppressor of a cotton jasmonic acid signal, and through interaction with the GhJZ12, the heterogenous dimer is formed to directly inhibit the resistance-related proteins such as GhMYC2-like and GhPR10, GhR1, GGD2 and the like, and the resistance and the insect resistance of the cotton are negatively controlled; and the gibberellin signal path can be indirectly activated through the interaction with the DELLA protein, the growth and development process of controlling the flowering stage, the seed germination and the fruit branch included angle and the like of the cotton. In addition, GJAK2 is involved in the development of the stamen of the cotton under high-temperature stress and low-temperature stress. It was found that the GHB12 gene was induced up-regulated by ABA, MeJA, SA, salt stress, cold stress and V991, which is an important multi-effect gene for regulating and controlling the defense response and growth of cotton. the GhHB12 negative control plant ABA signal path and the negative control plant resistance to abiotic stress; and meanwhile, the cotton JA signal path is negatively regulated, the disease resistance and the insect resistance of the cotton are reduced, and the expression of the flowering genes such as the GhFT, the GhSOC1 and the GhFUL can be directly controlled, and the development of the flowering, branch and fruit branches of the cotton can be controlled. In addition, the GhHB12 is involved in the control of the leaf shape of the cotton. We also found that the GhHB12 was regulated by the photoperiod and the rhythm, and the function of the GhHB12 in the control of the cotton plant type was lost under short-day conditions. The above results show that bioinformatics analysis can improve the effectiveness of the key control factors for identifying the response of the cotton in response to the multiple adversity, and the above results provide a reference for the physiological mechanism of the response of the cotton to the multiple adversity.
【学位授予单位】：华中农业大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S562
本文编号：2315565
[Abstract]:The plant can sense various environmental changes of the outside through the complex and fine control network, activate the relevant signal path, resist the stress of adversity, but at the same time, it is often accompanied by the change of the growth and development process and the yield. Due to the complexity of the gene regulation network, some genes are located at the intersection of multiple signal paths to form the hub gene and have different functions in multiple signal transduction to form a multi-effect. The specific function of the multi-effect gene in various stress-stress responses and its influence on the growth and development of the system have an important role in the future identification of crop broad-spectrum anti-reverse regulation factors. In this study, a number of important candidate genes in response to multiple adversity were selected by means of bioinformatics, and the three genes were identified. The main results are as follows: 1. By using the database of STIFDB and GENVESTIGATOR V3, the regulation factors of the broad-spectrum response of 826 arabidopsis thaliana were selected. and the candidate genes of the 38 cotton broad-spectrum response adversity are further separated and screened by the homologous sequence method. The expression patterns of 38 cotton candidate genes under salt stress, cold stress and ABA treatment were analyzed. To further verify the reliability of the strategy, we selected three of the genes (GhATAF1, GJAK2, GHB12) to perform the functional study of the system. It is found that GhATAF1 encodes a NAC transcription factor with self-activation activity, not only is induced by abiotic stress such as ABA, salt stress and cold stress, but also is strongly influenced by the induced expression of MeJA, SA and Verticillium dahliae V991, and the salt tolerance of cotton is being regulated. and meanwhile, the resistance of the cotton to the Verticillium dahliae is negatively regulated. The GhATAF1 can activate the expression of the cotton potassium ion/ sodium ion cotransporter GhHKT1, and can also activate the expression of the ABA signal key response gene GhABI4, the vacuole hydrogen ion-pyrophosphatase GhAVP1, the late embryo-rich protein GhLEA3, GhLEA6, the drought response gene GhR22 and the GhDREB2A, the accumulation of sodium ions in the cotton tissue is reduced, the damage of the cells is reduced, and the salt tolerance of the cotton is improved; in addition, the GhATAF1 is found to be capable of activating the path of the cotton salicylic acid signal, and meanwhile, the signal path of the jasmonic acid is inhibited, and the resistance of the cotton to the large-lily and the gray mold is finally weakened. It was found that the GhJAK2 gene was induced up-regulated by ABA, MeJA, SA, salt stress, cold stress and V991, which is an important node gene for regulating and controlling the defense response and growth of cotton. GhJAK2 is used as a suppressor of a cotton jasmonic acid signal, and through interaction with the GhJZ12, the heterogenous dimer is formed to directly inhibit the resistance-related proteins such as GhMYC2-like and GhPR10, GhR1, GGD2 and the like, and the resistance and the insect resistance of the cotton are negatively controlled; and the gibberellin signal path can be indirectly activated through the interaction with the DELLA protein, the growth and development process of controlling the flowering stage, the seed germination and the fruit branch included angle and the like of the cotton. In addition, GJAK2 is involved in the development of the stamen of the cotton under high-temperature stress and low-temperature stress. It was found that the GHB12 gene was induced up-regulated by ABA, MeJA, SA, salt stress, cold stress and V991, which is an important multi-effect gene for regulating and controlling the defense response and growth of cotton. the GhHB12 negative control plant ABA signal path and the negative control plant resistance to abiotic stress; and meanwhile, the cotton JA signal path is negatively regulated, the disease resistance and the insect resistance of the cotton are reduced, and the expression of the flowering genes such as the GhFT, the GhSOC1 and the GhFUL can be directly controlled, and the development of the flowering, branch and fruit branches of the cotton can be controlled. In addition, the GhHB12 is involved in the control of the leaf shape of the cotton. We also found that the GhHB12 was regulated by the photoperiod and the rhythm, and the function of the GhHB12 in the control of the cotton plant type was lost under short-day conditions. The above results show that bioinformatics analysis can improve the effectiveness of the key control factors for identifying the response of the cotton in response to the multiple adversity, and the above results provide a reference for the physiological mechanism of the response of the cotton to the multiple adversity.
【学位授予单位】：华中农业大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S562
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