硫化氢在白菜根部抵抗镉胁迫过程中的作用

发布时间：2018-06-12 13:45

本文选题：硫化氢 + Cd胁迫　；参考：《山西大学》2016年博士论文

【摘要】：近年来,H_2S作为生理信号分子得到了研究者越来越多的关注,已有的报道表明在植物中它具有促进根生长和形态建成、增强光合作用、促进种子萌发、刺激开花、延迟衰老等多方面生理功能。H_2S在植物抵抗各种环境胁迫过程中,也扮演了积极的角色。其中,在植物适应重金属胁迫的过程中发挥如下方面的作用：H_2S可以促进还原型谷胱甘肽、抗坏血酸的合成,并提高一系列抗氧化酶的活性来维持细胞内氧化还原状态的平衡,降低胁迫造成的氧化损伤；研究表明H_2S提高植物中Na+-ATPase、 K+-ATPase和H+-ATPase的活性来抑制对重金属离子的吸收,同时增加植物对硫、磷、钙、镁和铁的吸收,H_2S还可以抑制受H202调控的Ca通道来减少Cd进入质膜,并且激活液泡膜上的转运通道,将胞质中Cd转运至液泡,从而降低胞质内的重金属离子的浓度；外源H_2S通过提高植物的叶绿素含量,光合作用强度,以及促进细胞内叶绿体内囊的发育,增加线粒体数目来缓解Cd对植物造成的毒害：H_2S还会和其他信号分子、多种激素相互作用来缓解重金属对植物造成的伤害。本研究以重要的十字花科芸薹属作物大白菜(Brassica rapa L. ssp. Pekinensis)为材料,运用药理学、细胞生物学、生理生化等实验技术,从表型、细胞、分子多个水平对H_2S在提高白菜根部耐受重金属Cd胁迫过程中的生理作用进行系统研究。1.H_2S对Cd胁迫下白菜根部细胞死亡的影响。Cd胁迫下,随着处理浓度的增加,白菜幼苗根的生长受到抑制,同时根部细胞的死亡率也迅速提高,且根长和根部细胞死亡率呈负相关。Cd胁迫下白菜幼苗根部内源H_2S产率和H_2S生成关键酶的编码基因表达量都显著上调。外源H_2S处理(NaHS为供体),可以缓解Cd对白菜幼苗根的生长抑制,并且显著减少根部细胞的死亡和DNA的降解。进一步研究表明,H_2S信号可以减少白菜幼苗根中Cd胁迫诱导的活性氧(ROS)积累、抑制ROS诱导的根部膜脂过氧化损伤并维护质膜结构的完整,从而抑制根部细胞死亡。2.H_2S对Cd胁迫下白菜根部抗氧化系统的影响。对白菜根部抗氧化系统指标检测表明,H_2S可以降低Cd胁迫下根部H202的含量,减少细胞内脂质过氧化产物丙二醛(MDA)和蛋白质羰基化产物(PCO)的积累。进一步对ROS产生和清除系统进行研究后发现：H_2S抑制了质膜上02·-的产生酶NADPH氧化酶(NOX)编码基因BraRohD、BraRohF的表达,同时对NOX的活性也有明显抑制；Cd胁迫中ROS清除系统中超氧化物歧化酶(SOD)和过氧化氢酶(CAT)也受到H_2S的调节,转录水平和酶活水平都有不同程度的增加,而过氧化物酶(POD)和抗坏血酸过氧化物酶(APX)则没有明显变化；H_2S预处理可以促进Cd胁迫下抗氧化剂谷胱甘肽(GSH)的合成,减少了氧化型谷胱甘肽(GSSG)的产生,这有效地提高了GSH/GSSG比值,相应地检测到GSH-AsA循环系统中抗坏血酸(AsA)含量的显著增加。这些结果说明H_2S信号可从酶和非酶系统多方面调节植物体内ROS的平衡,降低植物在Cd胁迫下的氧化损伤程度。3.H_2S对Cd胁迫白菜幼苗根尖细胞遗传损伤的影响。Cd胁迫可使白菜幼苗根的生长受到严重抑制,根尖细胞的有丝分裂也受到了抑制。生理浓度H_2S处理可以促进Cd胁迫中根尖细胞有丝分裂的发生,而且可以显著降低有丝分裂过程中染色体的断裂、滞后、染色体桥等异常现象,细胞的微核发生率也显著降低。H_2S对Cd胁迫造成的DNA损伤也有明显的抑制作用,DNA-蛋白质交联程度降低,基因组DNA的断裂减少,与DNA损伤修复相关的多聚腺苷二磷酸-核糖聚合酶(PARP)家族中的PARP1和PARP2编码基因显著上调。这些结果表明H_2S在抑制Cd胁迫对白菜根尖细胞造成的遗传损伤过程中发挥了积极作用。4.H_2S对Cd胁迫下白菜幼苗根系渗透胁迫的影响在Cd胁迫过程中,随处理时问的延长,白菜根系的膜脂过氧化产物MDA含量增加,根细胞的相对电导率增大,细胞膜透性增大；H_2S预处理可使Cd胁迫引起的白菜根相对电导率显著下降,提高根部的相对含水量；进一步研究表明,H_2S增加Cd胁迫下幼苗根部可溶性糖和脯氨酸的含量来抵抗渗透胁迫,但可溶性蛋白和甜菜碱含量无明显变化。综上所述：Cd胁迫下白菜幼苗根部的内源H_2S合成增加；H_2S预处理可以减少Cd胁迫下根部细胞ROS诱导的细胞死亡；根部细胞ROS的积累减少源自H_2S从多方面调节ROS的产生和清除；H_2S可以降低Cd胁迫对根部细胞造成的遗传损伤,促进根尖细胞有丝分裂；H_2S通过调节Cd胁迫过程中白菜根部多种渗透调节物质的变化来抵抗Cd胁迫。因此,H_2S提高白菜幼苗根部耐受Cd胁迫的能力是通过多层次的防御和调节机制实现的。
[Abstract]:In recent years, more and more attention has been paid to H_2S as a physiological signal molecule. Reports have shown that in plants it has many physiological functions, such as promoting root growth and morphogenesis, enhancing photosynthesis, promoting seed germination, stimulating flowering, delayed senescence, and so on,.H_2S also plays a role in the resistance of plants to various environmental stresses. Positive roles. Among them, plants play the following role in the process of plant adaptation to heavy metal stress: H_2S can promote the synthesis of reduced glutathione, ascorbic acid, and increase the activity of a series of antioxidant enzymes to maintain the balance of redox state in cells and reduce oxidative damage caused by stress; studies show that H_2S increases The activities of Na+-ATPase, K+-ATPase and H+-ATPase in plants inhibit the absorption of heavy metal ions and increase the absorption of sulfur, phosphorus, calcium, magnesium and iron in plants. H_2S also inhibits the Ca channel regulated by H202 to reduce Cd into the plasma membrane and activates the transport channel on the vacuolar membrane and transtransport Cd in the cytoplasm to the vacuole, thus reducing the cytoplasm in the cytoplasm. The concentration of heavy metal ions; exogenous H_2S can alleviate the toxic effects of Cd on plants by increasing the chlorophyll content of plants, the intensity of photosynthesis, and promoting the development of the internal capsule of the cells in the cells, and increasing the number of mitochondria to alleviate the damage to plants. H_2S will also interact with other signaling molecules and various hormones to alleviate the damage caused by heavy metals to plants. In this study, Brassica rapa L. ssp. Pekinensis, an important cruciferous Brassica plant, was used as the material, using pharmacological, cell biology, physiological and biochemical techniques to systematically study the physiological role of H_2S in improving the physiological role of Cd stress tolerance in the root of Chinese cabbage from the phenotypic, cell and molecular levels of H_2S in the process of increasing the tolerance to heavy gold in the root of Chinese cabbage. The effects of Cd stress on the cell death of cabbage root cells under.Cd stress, with the increase of treatment concentration, the root growth of cabbage seedlings was inhibited and the mortality of root cells increased rapidly. The root length and root cell mortality were negatively correlated with the endogenous H_2S yield in root of cabbage seedlings and the encoding genes of key enzymes of H_2S generation under.Cd stress. Exogenous H_2S treatment (NaHS as donor) could alleviate the growth inhibition of Cd on the root of cabbage seedlings, and significantly reduce the death of root cells and the degradation of DNA. Further research shows that H_2S signal can reduce the accumulation of active oxygen (ROS) induced by Cd stress in the root of cabbage seedlings and inhibit the membrane lipid peroxidation induced by ROS. Damage and maintenance of the integrity of the plasma membrane structure, thus inhibiting the effect of cell death.2.H_2S on the antioxidant system in root of Chinese Cabbage under Cd stress. The detection of antioxidant system in the root of Chinese cabbage shows that H_2S can reduce the content of H202 at the root of Cd stress, reduce the malondialdehyde (MDA) and protein carbonylation products of the lipid peroxylation products in the cells. After further study on the ROS production and scavenging system, it was found that H_2S inhibited the 02. - producing enzyme NADPH oxidase (NOX) encoding gene BraRohD, the expression of BraRohF, and the activity of NOX in the membrane of the plasma membrane, and also inhibited the activity of NOX, and the superoxide dismutase (SOD) and catalase (CAT) in the ROS scavenging system in Cd stress were also affected. The regulation of _2S, transcriptional level and enzyme activity level increased in varying degrees, while peroxidase (POD) and ascorbic acid peroxidase (APX) did not change significantly; H_2S pretreatment could promote the synthesis of antioxidant glutathione (GSH) under Cd stress and reduced the production of oxidized glutathione (GSSG), which effectively improved GSH/GSS. The G ratio, correspondingly detected the significant increase in the content of ascorbic acid (AsA) in the GSH-AsA cycle system. These results show that H_2S signals can regulate the balance of ROS in plants in various aspects of enzymes and non enzyme systems, and reduce the oxidative damage degree of plants under Cd stress.3.H_2S on the effect of.Cd stress on the genetic damage of the root tip cells of the cabbage seedlings under the stress of Cd. The root growth of cabbage seedlings was severely inhibited and the mitosis of root tip cells was inhibited. Physiological concentration H_2S treatment could promote the mitosis of root tip cells in Cd stress, and could significantly reduce chromosome breakage, lag, chromosome bridge and other abnormal phenomena in the process of mitosis, and the incidence of micronucleus in cells. It also significantly reduced the inhibitory effect of.H_2S on the DNA damage caused by Cd stress, the decrease of DNA- protein crosslinking degree and the decrease of genomic DNA. The PARP1 and PARP2 encoding genes in the polyadenosine two phosphoric ribose polymerase (PARP) family associated with DNA damage repair were significantly up-regulated. These results indicate that H_2S is inhibiting Cd stress. In the process of genetic damage caused by root tip cells of Chinese cabbage, the effect of.4.H_2S on the root osmotic stress of Cabbage Seedlings under the stress of Cd stress under the stress of Cd stress in the process of Cd stress, with the prolongation of treatment time, the content of MDA content of membrane lipid peroxidation product in root of Chinese cabbage increased, the relative conductivity of root cells increased, cell membrane permeability increased; H_2S pretreatment The relative conductivity of cabbage root caused by Cd stress decreased significantly and increased the relative water content of root. Further studies showed that H_2S increased the content of soluble sugar and proline in root of seedling under Cd stress to resist osmotic stress, but there was no obvious change in soluble protein and betaine content. Under Cd stress, the root of Cabbage Seedlings under Cd stress The endogenous H_2S synthesis increased; H_2S pretreatment could reduce the cell death induced by ROS in root cells under Cd stress; the accumulation of ROS in root cells decreased from H_2S to regulate the production and removal of ROS in many ways; H_2S could reduce the genetic damage caused by Cd stress to root cells and promote mitosis in the root tip cells; H_2S by regulating Cd. In the course of stress, the changes of various osmotic regulators in the root of Chinese cabbage resist Cd stress. Therefore, the ability of H_2S to improve the tolerance to Cd stress in the roots of Chinese cabbage is achieved through a multi-level defense and regulation mechanism.
【学位授予单位】：山西大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S634.1

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