基于蛋白质组学对螺旋藻砷胁迫响应机制的研究

通过研究螺旋藻(Spirulina sp.)在砷离子胁迫下的蛋白质组变化,从蛋白质表达水平解释螺旋藻对砷离子胁迫的响应机理。螺旋藻经过不同浓度砷离子胁迫7 d后,提取蛋白质进行凝胶电泳,并对差异蛋白进行质谱分析。结果表明螺旋藻在2.0 ppm砷酸盐中暴露10 min光合放氧速率降低27.3%,培养24 h后细胞内的金属硫蛋白、叶绿素、类胡萝卜素及藻胆蛋白相对含量均明显降低。蛋白组学共鉴定出75个差异蛋白,其中26个显著上调,49个呈现下调。这些差异蛋白表明砷离子主要通过破坏螺旋藻光合色素蛋白,干扰电子传递过程,导致能量合成受损,使得依赖光合作用产生能量进行的跨膜运动、蛋白质合成等相关过程受到影响;同时,活性氧清除与防御相关蛋白呈现上调,螺旋藻细胞内抗氧化系统被激活。     

Proteomic Analysis of Rice Seedlings Under Cold Stress

Low temperature can greatly restrict the growth and development of rice. The rice seedlings show growth retardation, lamina wrap, and part of blade even died under the condition of low temperature. In order to get more information about cold stress responses in rice, two dimensional electrophoresis and bioinformatics analysis of mass spectrometry were used to preliminary survey the cold tolerance of cold sensitive line 9311 and cold resistance variety Fujisaka 5 under cold stress. Two dimensional electrophoresis maps of 9311 and Fujisaka 5 were established under cold treatment. With analysis of bioinformation, the proteins were found involve in many aspects of rice development. The largest category of proteins is functioning on metabolism. By comparing the proteins from the two varieties, it can be found that most proteins from 9311 were down-regulated and were up-regulated in Fujisaka 5. The results showed that the membrane composition and structure were damaged, metabolism changed dramatically and rice defense system was activated under the cold stimulation. Fifty-nine proteins related to the resistance of cold stress were identified in our study, and we have investigated and classified all of their biological functions. The importance of our study are providing some conduct for the research of rice resistant to cold stress, supporting auxiliary technique for rice varieties and widening the search field of cold tolerance in plants.     

Proteomic Analysis of the Response of Liangyoupeijiu （Super High-Yield Hybrid Rice） Seedlings to Cold Stress

Liangyoupeijiu is a super high-yield hybrid rice. Despite its advantages with respect to yield and grain quality, it is sensitive to cold, which keeps it from being widely cultivated. We subjected Liangyoupeijiu seedlings to 4 ℃ cold treatment, then extracted the leaf proteins. After 2-D gel electrophoresis separation and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis, a series of differentially displayed proteins were identified. Some metabolism-associated proteins were found among the downregulated proteins, such as carbamoyl phosphate synthetase, transketolase 1, dihydrolipoamide dehydrogenase and glyceraldehyde 3-phosphate dehydrogenase. The upregulated proteins included both stress-resistance proteins such as nucleoside diphosphate kinase I and proteins that are negative for rice growth, such as FtsH-like protein, plastid fusion and/or translocation factor （Pftf） and actin. Our results indicate that cold may inhibit Liangyoupeijiu growth through decreasing metabolic activity and damaging cell structure.     

Proteomic Analysis of Trypanosoma cruzi Response to Ionizing Radiation Stress

Trypanosoma cruzi, the causative agent of Chagas disease, is extremely resistant to ionizing radiation, enduring up to 1.5 kGy of gamma rays. Ionizing radiation can damage the DNA molecule both directly, resulting in double-strand breaks, and indirectly, as a consequence of reactive oxygen species production. After a dose of 500 Gy of gamma rays, the parasite genome is fragmented, but the chromosomal bands are restored within 48 hours. Under such conditions, cell growth arrests for up to 120 hours and the parasites resume normal growth after this period. To better understand the parasite response to ionizing radiation, we analyzed the proteome of irradiated (4, 24, and 96 hours after irradiation) and non-irradiated T. cruzi using two-dimensional differential gel electrophoresis followed by mass spectrometry for protein identification. A total of 543 spots were found to be differentially expressed, from which 215 were identified. These identified protein spots represent different isoforms of only 53 proteins. We observed a tendency for overexpression of proteins with molecular weights below predicted, indicating that these may be processed, yielding shorter polypeptides. The presence of shorter protein isoforms after irradiation suggests the occurrence of post-translational modifications and/or processing in response to gamma radiation stress. Our results also indicate that active translation is essential for the recovery of parasites from ionizing radiation damage. This study therefore reveals the peculiar response of T. cruzi to ionizing radiation, raising questions about how this organism can change its protein expression to survive such a harmful stress.     

蛋白质组学在细菌应激反应研究中的应用

当外部生存环境发生变化时,细菌会在短时间内发生应激反应。利用双向凝胶电泳技术结合生物质谱鉴定的方法对细菌蛋白表达谱变化进行研究,是细菌转录谱变化研究的深入和扩展,是细菌应激反应研究中的新热点。综述了蛋白质组学在细菌应激反应研究中的应用现状和存在问题。     

植物响应低温胁迫的应答机制

介绍了植物响应低温胁迫的生理生化和分子机制及其信号转导途径的研究进展。并对今后这一领域研究前景作了展望和讨论。     

脱水素在植物低温胁迫响应中的作用

脱水素(dehydrin)是一类胚胎发育后期丰富蛋白(LEA.late embryogenesis abundant proteins),含有富含赖氨酸的K片段,属于具有高度热稳定性的亲水性蛋白,在植物脱水条件下能保护细胞内蛋白质和膜结构免受破坏.低温胁迫下,耐低温植物细胞内部会发生一系列的生理生化反应来抵御低温所造成的伤害.很多研究表明植物脱水素的表达和积累与多种双子叶植物(包括草本和木本植物)以及冬季栽培的禾本科植物品种(特别是小麦和大麦)的耐低温能力密切相关.本文对近年来国内外关于脱水素的结构、功能以及内源ABA(abscisie acid)含量、拟南芥CBF(C-repeat binding factor)同源转录激活因子、春化基因、光周期信号等对脱水素基因的表达调控机制进行综述.     

Proteomic Analysis Identifies Ribosome Reduction as an Effective Proteotoxic Stress Response

Stress responses are adaptive cellular programs that identify and mitigate potentially dangerous threats. Misfolded proteins are a ubiquitous and clinically relevant stress. Trivalent metalloids, such as arsenic, have been proposed to cause protein misfolding. Using tandem mass tag-based mass spectrometry, we show that trivalent arsenic results in widespread reorganization of the cell from an anabolic to a catabolic state. Both major pathways of protein degradation, the proteasome and autophagy, show increased abundance of pathway components and increased functional output, and are required for survival. Remarkably, cells also showed a down-regulation of ribosomes at the protein level. That this represented an adaptive response and not an adverse toxic effect was indicated by enhanced survival of ribosome mutants after arsenic exposure. These results suggest that a major source of toxicity of trivalent arsenic derives from misfolding of newly synthesized proteins and identifies ribosome reduction as a rapid, effective, and reversible proteotoxic stress response.     

木薯MeMYC2.2基因耐低温功能研究

MYC2（MYeloCytomatosis）转录因子是植物应对逆境胁迫过程中茉莉酸信号传导相关的核心转录因子。本研究旨在初步分析木薯MeMYC2.2基因在低温胁迫响应中的功能。利用生物信息学分析木薯MeMYC2.1和MeMYC2.2基因的结构及其编码蛋白的理化性质;通过定量PCR分析了上述2个基因在木薯组培苗叶片中对低温胁迫的响应;通过转基因拟南芥研究MeMYC2.2的抗冻功能。木薯组培苗叶片中2个MeMYC2基因的表达均在低温胁迫早期被诱导,其中,与MeMYC2.1相比,MeMYC2.2差异表达更显著。MeMYC2.2蛋白主要定位于细胞核中,且在酵母中具有明显转录自激活功能,表明该蛋白具有转录因子特性。与野生型相比,过表达MeMYC2.2的转基因拟南芥抗冻能力显著提高。在低温处理下,CBF3基因在转基因拟南芥中的表达量要明显高于其在野生型的表达量,但另外3个CBF基因在转基因拟南芥中的表达量明显下降。木薯MeMYC2.2的表达受低温和茉莉酸调控,可以提高植物的抗冻性,且可能影响CBF基因对低温的响应。本研究为进一步利用MeMYC2基因改良木薯的低温耐受性奠定了理论基础。     

Comparative Physiological and Proteomic Analysis Reveals the Leaf Response to Cadmium-Induced Stress in Poplar (Populus yunnanensis)

Excess amounts of heavy metals are important environmental pollutants with significant ecological and nutritional effects. Cdmium (Cd) is of particular concern because of its widespread occurrence and high toxicity. We conducted physiological and proteomic analyses to improve our understanding of the responses of Populus yunnanensis to Cd stress. The plantlets experienced two apparent stages in their response to Cd stress. During the first stage, transiently induced defense-response molecules, photosynthesis- and energy-associated proteins, antioxidant enzymes and heat shock proteins (HSPs) accumulated to enhance protein stability and establish a new cellular homeostasis. This activity explains why plant photosynthetic capability during this period barely changed. During the second stage, a decline of ribulose-1, 5-bisphosphate carboxylase (RuBisCO) and HSP levels led to imbalance of the plant photosynthetic system. Additionally, the expression of Mitogen-activated protein kinase 3 (MPK3), Mitogen-activated protein kinase 6 (MPK6) and a homeobox-leucine zipper protein was higher in the second stage. Higher expression of caffeoyl-CoA O-methyltransferase (CCoAOMT) may regulate plant cell wall synthesis for greater Cd storage. These genes may be candidates for further research and use in genetic manipulation of poplar tolerance to Cd stress.     

外源H2S影响黄瓜幼苗响应高盐胁迫的蛋白质组学分析

为了阐明外源H₂S对高盐胁迫下黄瓜蛋白质表达的影响,以盐敏感型黄瓜栽培种‘春夏秋王’为材料,通过双向电泳（2 DE）技术分离叶片总蛋白质,采用基质辅助激光解析飞行时间串联质谱（MALDI TOF/TOF MS）技术对差异表达蛋白质进行质谱鉴定,并利用NCBI及SwissProt数据库检索进行差异表达蛋白质功能注释和代谢通路分析。结果表明：(1) 共检测到约2 490个蛋白质,其中蛋白质表达差异在1.5倍以上且差异显著(P<0.05)的有45个,其中有24个蛋白质表达上调,21个蛋白质表达下调。(2) 成功鉴定蛋白质26个,这些蛋白质参与了光合作用（26.92%）、蛋白质代谢（23.08%）、能量与碳水化合物代谢（11.54%）、氨基酸生物合成（11.54%）、细胞结构相关蛋白（7.69%）、抗氧化作用（3.85%）、信号转导（3.85%）及未知功能蛋白（11.54%）。(3) GO注释表明差异表达的蛋白质分子功能主要以蛋白质结合与水解酶活性为主,生物学过程涉及应激反应、有机物代谢过程、胁迫响应和细胞分化等,细胞组成集中分布在细胞部分和一些胞内细胞器。KEGG代谢通路分析表明,差异表达蛋白质主要参与了肌动蛋白细胞骨架调控、细胞凋亡、碳代谢和光和调控等代谢途径。研究表明,外源H₂S诱导了高盐胁迫下黄瓜叶片蛋白质的差异表达,为后续探索外源H₂S对黄瓜的抗盐分子机制研究提供了理论依据。     

杨树HDA902基因在低温胁迫应答反应中的功能

组蛋白去乙酰化酶在植物非生物胁迫应答反应中具有重要的调控作用。利用RT-PCR的方法从毛果杨中克隆了组蛋白去乙酰化酶基因HDA902。利用农杆菌介导法将其遗传转化到烟草中,并对转基因植株进行低温耐受性分析。研究结果表明,HDA902在烟草中的表达显著提高了转基因株系对低温的耐受性。叶片NBT和DAB染色结果表明,在低温处理后转基因烟草比野生型烟草产生较少的活性氧。丙二醛和脯氨酸含量测定结果表明,在低温条件下,转基因烟草叶片的脯氨酸含量显著高于野生型烟草,而丙二醛含量显著低于野生型烟草。这些研究结果表明,HDA902参与低温胁迫应答反应,其过量表达提高了植株耐低温的能力。     

木薯亲本正反交亲和力与授粉柱头的蛋白质组学分析

为研究木薯不同亲本杂交亲和力的差异,分别以华南5号、华南6号、华南7号为亲本进行正反交,统计不同组合间稔实率的差异。同时对受精柱头蛋白质进行分析,研究柱头对杂交亲和力影响的蛋白质调控水平。结果表明:SC5(♀)×SC7(♂)平均稔实率最高为26.22%,SC7(♀)×SC5(♂)平均稔实率最低为11.13%。通过双向电泳和质谱法分析母本SC5和SC7授粉柱头的蛋白质变化,得到27个差异蛋白质点,其中19个蛋白质出现上调表达,这些差异表达蛋白质分别参与碳代谢及能量代谢、分子伴侣和氨基酸代谢;而8个蛋白质下调表达,它们分别参与蛋白质合成及氢氰酸代谢;表明这些代谢途径对木薯亲和力高低有一定的影响。     

Proteomic Analysis of the Systemic Response to Radiographic Contrast Media

Introduction  

Radiographic contrast media (RCM) have numerous effects on the hemostatic system, inflammatory pathways, and vascular endothelium. Given the increasing number of high-risk patients undergoing radiographic procedures, more information regarding the systemic effects of RCM is needed.