TOR信号通路通过CIPK23调控马铃薯和拟南芥的钾离子吸收和生长

发布时间：2018-03-23 18:34

本文选题：马铃薯　切入点：拟南芥　出处：《西南大学》2017年硕士论文

【摘要】：钾离子是植物生长所必需的营养元素之一,参与到多种生理过程中,包括渗透调节,糖类的协同运输以及光合作用等。TOR是在真核生物中进化上保守的丝氨酸/苏氨酸蛋白激酶,属于PI3K蛋白激酶家族,可以响应能量、营养等信号调控细胞的生长发育,TOR激酶功能缺失时,导致植物叶片发黄,同时伴随有光合效率下降以及叶绿素降解。TOR信号通路是否通过调控钾素营养的吸收代谢从影响植物的光合作用、叶绿素生物合成等还未见报道,因此在本研究中通过药理学和遗传学试验研究了TOR信号通路与钾离子吸收的关系,得出以下四点结论:1)在低钾培养基上,马铃薯外植体迅速地变黄且不能正常完成根系再生过程;这种表型与在培养基中加入TOR抑制剂(KU、AZD)后植物的表型相似,均出现黄化的表型。同时通过钾离子吸收实验,测定了在TOR抑制时马铃薯对于钾离子的吸收能力,结果显示TOR的二代抑制剂能够明显抑制马铃薯中钾离子的吸收。2)本研究发现拟南芥低钾敏感型突变体cbl1cbl9,lks1-2,lks1-3以及akt1均对TOR抑制剂表现出敏感性。TOR抑制剂可以有效抑制野生型拟南芥和突变体的生长,这种抑制作用在突变体植株中表现尤为明显。测定钾离子浓度后发现,TOR抑制剂可以显著下调拟南芥对于钾离子的吸收;然而对比发现在钾离子吸收方面,TOR抑制剂对于野生型拟南芥植株的影响比突变体植株要更显著。3)本研究拟南芥中扩增了At CBL9,At CIPK23,At AKT1基因,构建了携带GFP标签的过表达载体,将此载体在拟南芥转基因植株BP12-2(雷帕霉素敏感的拟南芥转基因材料)中过表达,获得了At CBL9-GFP/BP12,At CIPK23-GFP/BP12以及At AKT1-GFP/BP12过表达株系。通过雷帕霉素敏感性试验确定了TOR与At CBL9,At CIPK23,At AKT1之间的关系。发现在雷帕霉素处理下At CIPK23-GFP/BP12过表达株系的生长情况要优于拟南芥BP12-2株系。在雷帕霉素和KU双抑制处理下,At CIPK23-GFP/BP12过表达株系可以继续生长,而拟南芥BP12-2株系则表现为叶片发黄,停止生长。通过钾离子浓度测定及叶绿素含量测定试验发现,在TOR抑制剂处理下,与BP12-2相比,过表达At CIPK23基因可以提高植株对于钾离子的吸收和叶绿素含量,这些结果表明CIPK23可能在TOR信号通路影响钾素吸收过程中具有重要的作用。4)在马铃薯基因组中,通过序列比对,发现存在At CBL1,At CIPK23以及At AKT1的同源基因;同时通过q PCR试验确定了基因在马铃薯中的表达模式为组成性表达。扩增了马铃薯中St CIPK23基因,并构建了马铃薯St CIPK23-GFP过表达株系;通过试验发现在幼苗生长中St CIPK23过表达株系对于TOR抑制剂具有抗性。这些数据表明马铃薯中CBL1-CIPK23-AKT1信号通路同样参与植物对低钾环境的响应。综合这些结果表明TOR信号通路可能通过调控CIPK23的功能来调节植物钾离子的吸收。之前以及最近的研究都表明TOR信号通路作为核心调控元件来调控植物的光合生长,细胞增殖以及细胞伸长等生理过程,然而这些研究大多均集中与对拟南芥的研究。在本研究中发现在马铃薯中TOR的功能同在拟南芥中同样保守,通过对马铃薯中TOR信号通路功能的研究将有助于提高马铃薯的产量。
[Abstract]:Potassium is one of the essential nutrients for plant growth, to participate in a variety of physiological processes, including osmotic adjustment, sugar transport and Photosynthesis of collaborative.TOR is an evolutionary conserved in eukaryotes on serine / threonine protein kinase, belongs to the PI3K protein kinase family, can respond to energy and nutrition regulation of cellular signal growth of TOR kinase function loss, resulting in plant leaf yellow, accompanied by the photosynthetic efficiency decreased and chlorophyll degradation of.TOR signaling pathway by regulating potassium absorption and metabolism from affecting plant photosynthesis, chlorophyll biosynthesis has not been reported, so in this study by pharmacological and genetic tests on the relationship between TOR signal pathway and potassium uptake, draw the following four conclusions: 1) in low potassium medium, potato explants quickly turn yellow and not just To complete the regeneration process of root; this phenotype and the TOR inhibitor was added into the culture medium (KU, AZD) after the phenotypic similarity of plants showed chlorosis phenotype. The potassium ion absorption experiments in TOR inhibition of potato for K + absorption was measured, the results showed the uptake of.2 TOR two generation inhibitor can obviously inhibit the growth of potassium ions in Arabidopsis) this study found that low potassium sensitive mutants cbl1cbl9, lks1-2, lks1-3 and AKT1 were on TOR inhibitors showed sensitivity to.TOR inhibitors can effectively inhibit the Arabidopsis wild-type and mutant growth, the inhibition performance in mutant plants is particularly obvious. Determination of potassium ion concentration found TOR inhibitors can significantly down regulated in Arabidopsis for potassium ion absorption; however, compared the absorption of potassium ion, TOR inhibitors for the wild-type Arabidopsis plants. Ring than the mutant plants more significant.3) this study in Arabidopsis thaliana was amplified At CBL9, At CIPK23, At AKT1 gene, constructed over expression vector carrying the GFP tag, this vector in Arabidopsis transgenic plants BP12-2 (rapamycin sensitive Arabidopsis transgenic materials) in over expression, won the At CBL9-GFP/BP12, At CIPK23-GFP/BP12 and At over expression of AKT1-GFP/BP12 and At strains. TOR CBL9 was determined by At CIPK23 At rapamycin sensitivity test, AKT1. The relationship of overexpression lines in rapamycin treated CIPK23-GFP/BP12 growth of At is superior to BP12-2 in Arabidopsis lines. Rapamycin and KU double inhibition, At CIPK23-GFP/BP12 overexpression strains can continue to grow, and Arabidopsis BP12-2 strains showed the yellow leaves, stop growing. Through the determination of the chlorophyll content and the potassium ion concentration test found in TOR The inhibitor treatment, compared with BP12-2, expression of At CIPK23 gene can improve plant for absorption and chlorophyll content of potassium ions, these results suggest that CIPK23 may affect the potassium in the signal pathway of TOR uptake has an important role for.4 in the process) in potato genome, found by sequence alignment, At CBL1, At CIPK23 and At AKT1 homologous genes; at the same time through the Q test to determine the expression pattern of PCR gene in potato is constitutively expressed in St. The amplification of potato CIPK23 gene, and constructed the potato St CIPK23-GFP overexpression strains; the experiment found that overexpression strains with resistance to TOR inhibitors in seedling growth in St CIPK23 these data. Show that the CBL1-CIPK23-AKT1 signaling pathway in potato is also involved in plant response to low potassium environment. These results indicated that TOR signaling pathway may be regulated by CIPK2 3 functions to regulate plant potassium absorption. Before, recent studies have indicated that TOR signaling pathway as a key regulatory element to regulate plant photosynthesis and growth, cell proliferation, cell elongation and other physiological processes, however, most of these studies are focused on the research and the Arabidopsis. In this study, found in potato TOR the function in Arabidopsis as conservative through the research on the function of TOR pathway in potato will help to increase the yield of potato.

【学位授予单位】：西南大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S532;Q943.2

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