外源茉莉酮酸甲酯通过调节相关基因表达诱导光温敏雄性不育小麦BS366离体花药开裂

植物育性与花药开裂性状相关.茉莉酮酸甲酯(methyl jasmonic acid,MeJA)是一种植物激素,广泛参与植物的胁迫应答和生长发育过程.植物体内存在6类基因所编码的酶共同维持茉莉酮酸(JA)代谢途径的畅通,并与植物花药开裂有关.本研究以小麦光温敏不育系BS366为材料,于抽穗期—开花期进行穗部离体48 h的MeJA外源处理.分别于0、1、3、6、12、24和48 h时间点收集花药并进行RNA提取.通过半定量RT-PCR分析发现,在内参基因甘油醛-3-磷酸脱氢酶(glyceraldehyde-3-phosphate dehydrogenase,GAPDH)表达基本一致的情况下,BS366的JA代谢途径中参与调控的6种基因均存在不同程度的诱导表达.经MeJA的48 h处理后的离体花药开裂程度明显增大.本研究获得了BS366离体环境下JA代谢途径基因经MeJA诱导后表达水平的概况,并为诱导BS366花药开裂及人工调控育性,达到高效制种及繁种的目的提供了初步的理论依据.     

茉莉酸调控花药开裂的研究进展

茉莉酸(JA)是广泛存在于植物中的生长调节物质,JA及其衍生物茉莉酸甲酯(Me JA)在植物生命活动中起着重要作用。JA参与调控雄蕊发育,影响花药开裂,从而影响植物育性。就JA的生物合成及相关基因的表达调控、JA在植物花药发育尤其是后期花药开裂过程中相关基因以及信号转导的分子机制研究进行回顾总结,并对JA调控花药开裂的分子机理研究提出展望。     

茉莉酸甲酯对广藿香JA信号转导途径及倍半萜合成途径关键基因表达的影响

分析外源性茉莉酸甲酯对广藿香JA信号转导途径关键基因JAZ2、MYC2、COI1及倍半萜合成途径关键基因PTS、FPPS、SQLE表达的影响,为深入研究茉莉酸甲酯调控广藿香JA信号转导途径及倍半萜合成途径的分子机制奠定基础。该文分别用0.10和0.25 mmol·L~(-1)的MeJA喷施广藿香叶片,于处理后的0、2、6、12、24、48、72 h摘取叶片,运用实时荧光定量PCR对JAZ2、MYC2、COI1、PTS、FPPS、SQLE基因的表达量进行检测。结果表明:0.10和0.25 mmol·L~(-1)的MeJA对广藿香JA信号转导途径JAZ2、MYC2、COI1及倍半萜合成途径PTS、FPPS、SQLE基因表达均有不同程度的促进作用,其中对JAZ2基因表达影响最显著。0.10mmol·L~(-1)MeJA溶液处理2 h时,JAZ2表达量上调13.52倍; 0.25 mmol·L~(-1)MeJA溶液处理48 h时,JAZ2表达量上调19.09倍。JA信号转导途径关键基因JAZ2与倍半萜合成途径关键基因FPPS存在极显著正相关关系。综上结果表明MeJA溶液可诱导广藿香JAZ2、MYC2、COI1、PTS、FPPS、SQLE基因的表达,且不同浓度MeJA对基因表达有着不一样的影响; JAZ2是JA信号转导途径里响应MeJA诱导的主要基因,其可激活倍半萜合成途径FPPS基因的协同表达,进而影响广藿香醇等倍半萜合成。     

茉莉酸和乙烯信号途径参与了油菜菌核病防卫反应

茉莉酸(JA)和乙烯(ET)介导的信号防卫反应在植物抗病性方面具有重要作用。用核盘菌(Sclerotinia sclerotiorum)分别接种甘蓝型油菜(Brassica napus)抗病品种中双9号和感病品种84039,通过荧光定量RT-PCR方法探测JA和ET的关键合成基因及其信号途径标志基因的相对表达水平。结果显示,S.sclerotiorum在这两个品种中都能够诱导这些基因的表达,并且这些基因的相对表达水平在中双9号中显著高于84039;甲基茉莉酸(MeJA)和1-氨基环丙烷-1-羧酸(ACC)的药理学实验结果显示JA和ET信号反应能够相互促进,并且JA和ET信号反应本身存在自我放大的功能。这些结果表明茉莉酸和乙烯信号转导途径参与了甘蓝型油菜菌核病防卫反应。     

巴西橡胶树转录中介体HbMed25基因克隆与表达分析

转录中介体(mediator,Med)中的Med25基因在植物的组织发育、器官发生、胁迫响应以及调控茉莉酸(JA)信号途径等方面起重要调控作用。该文在巴西橡胶树基因组序列中找到了橡胶树Med25基因的序列,通过RT-qPCR和RACE技术克隆HbMed25基因的全长序列并进行生物信息学预测分析结果显示,HbMed25基因包含2 655 bp的开放阅读框,编码884个氨基酸,蛋白分子量为95 kD,理论等电点为8.68,疏水性蛋白,亚细胞定位最可能为细胞核。Real-time PCR分析结果显示,HbMed25基因在树皮中表达量最高,在植物激素JA处理的形成层和胶乳材料中HbMed25基因在处理早期(1 h和2～4 h)上调表达,受割胶处理HbMed25基因在处理早期上调表达且对高产的橡胶树品系(CATAS 8-79和CATAS 7-33-97)的割胶处理响应更明显。综上结果认为,HbMed25基因极大可能参与了JA信号途径的调控,对阐明JA诱导橡胶树次生乳管分化的分子调控机制具有促进作用。     

COI1参与茉莉酸调控拟南芥吲哚族芥子油苷生物合成过程

芥子油苷是一类具有防御作用的植物次生代谢产物,外源激素茉莉酸对吲哚族芥子油苷的合成具有强烈的诱导作用,但茉莉酸调控吲哚族芥子油苷生物合成的分子机制并不清楚。以模式植物拟南芥(Arabidopsis thaliana)的野生型和coi1-22、coi1-23两种突变体为研究材料,通过茉莉酸甲酯(MeJA)处理,比较了拟南芥野生型和coi1突变体植株吲哚族芥子油苷含量、吲哚族芥子油苷合成前体色氨酸的生物合成基因(ASA1、TSA1和TSB1)、吲哚族芥子油苷生物合成基因(CYP79B2、CYP79B3和CYP83B1)及调控基因(MYB34和MYB51)的表达对MeJA的响应差异,由此确定茉莉酸信号通过COI1蛋白调控吲哚族芥子油苷生物合成,即茉莉酸信号通过信号开关COI1蛋白作用于转录因子MYB34和MYB51,进而调控吲哚族芥子油苷合成基因CYP79B2、CYP79B3、CYP83B1和前体色氨酸的合成基因ASA1、TSA1、TSB1。并且推断,COI1功能缺失后,茉莉酸信号可能通过其他未知调控因子或调控途径激活MYB34转录因子从而调控下游基因表达。     

茉莉酮酸酯和水杨酸酯对草食性昆虫细胞色素P450基因表达的诱导

茉莉酮酸酯和水杨酸酯是植物遭受昆虫或病原物攻击时 ,植物产生的激活本身防卫基因的信号物质。这些信号物质的增加 ,可以导致植物他感化合物和防卫蛋白合成增加。旱芹 (ApiumgraveolensCelery)是棉铃虫的寄主植物之一。用茉莉酮酸酯和甲基茉莉酮酸酯诱导旱芹体内花椒毒素和另一种类似的呋喃香豆素化合物香柠檬烯 ,发现处理后2 4h两种毒素开始积累 ,4～ 6d后到达最大值。用人工饲料添加茉莉酮酸酯和水杨酸酯的方法 ,研究棉铃虫 4种P45 0基因 (CYP6B8,CYP6B9,CYP6B2 7andCYP6B2 8)表达。用RT_PCR和XmnI消化及凝胶斑点分析检测处理…     

甘草JAZ基因家族的鉴定及其在干旱胁迫下的表达分析

甘草(Glycyrrhiza uralensis)是一味大宗中药材，黄酮类活性成分是其主要的药物活性成分之一。茉莉酸是甘草响应非生物胁迫调控类黄酮类活性成分的主要内源激素。JAZ是茉莉酸信号转导途径中的一个重要节点，其通过负向调节茉莉酸的信号转导，参与植物发育、非生物胁迫和对植物激素处理的响应的调节。本研究通过测定聚乙二醇(polyethylene glycol, PEG) 6000诱导的干旱胁迫对乌拉尔甘草(Glycyrrhiza uralensis Fisch.)中茉莉酸(jasmonic acid, JA)含量和JAZ基因家族表达水平以及JAZ基因家族的生物信息学分析，筛选受干旱胁迫诱导JAZ基因。研究结果显示：JA主要在甘草地下部分中积累，干旱胁迫明显提升地下部分JA含量，地下和地上部分中PEG处理2h时JA表达量最高。基于对上述材料的转录组测序数据分析，有10个GuJAZ基因具有完整的读码框，且这些JAZ基因具有时空表达差异性。干旱胁迫诱导地下部分中的GuJAZ基因上调，GuJAZ1、GuJAZ3、GuJAZ4、GuJAZ5、GuJAZ6和GuJAZ10在处理2h组上调最明显...     

茉莉酸类物质在草莓果实发育中的作用及其分子机理分析

该研究以草莓‘红颜’(Fragaria ananassa Duch.‘Benihoppe’)为试材,于草莓花后15d采用注射法开始注射茉莉酸甲酯(MeJA,浓度为400μmol/L),分析MeJA对草莓果实发育进程的影响及其相关基因的表达,以揭示MeJA在草莓果实发育和成熟调控中的作用及其分子机理。结果表明:(1)MeJA处理草莓果实后,果实变红成熟期比对照显著提前,平均提前4d;(2)随着草莓果实发育成熟,MeJA处理的茉莉酸(JA)合成基因FaOPDA1的表达量迅速升高;(3)FaOPDA1基因在草莓果实中的超表达能够促进草莓果实提前成熟3~5d,且FaOPDA1基因的超表达能够诱导与草莓果实成熟相关的一系列基因的表达量升高,从而促进草莓果实提前成熟。     

基于转录组分析鉴定苹果茉莉素响应基因

为阐明外源茉莉酸甲酯(MeJA)诱导的苹果(Malus domestica)抗病分子机制, 以生长30天的Gala组培苗为试材, 用100 μmol?L ^-1MeJA处理叶片12小时, 通过转录组测序, 结合生物信息学分析鉴定出苹果叶片中受MeJA诱导表达的基因。结果表明, 外源MeJA主要影响苹果叶片倍半萜类、三萜和类黄酮的生物合成, 以及芸薹素(BR)信号转导途径间接诱导的抗病性; 倍半萜类、三萜及类黄酮生物合成途径中的关键基因为MDP0000702120和MDP0000692178; MDP0000123379是联系芸薹素信号转导途径和植物-病原菌互作途径的关键调控基因。     

Characterization of two subclasses of PR-10 transcripts in lily anthers and induction of their genes through separate signal transduction pathways

The lily PR-10 belongs to a family of intracellular pathogenesis-related (IPR) proteins. Genomic Southern analysis indicates that the PR-10 is encoded by a family of multiple genes. Seven heterogeneous cDNA clones encoding lily PR-10 from Lilium longiflorum are divided into two subclasses based on sequence comparison and Southern hybridization. A 82% overall sequence similarity was found between the two subclasses (represented by PR-10c and d). The two cDNAs include an open reading frame of 474 bp encoding 157 amino acids. 5'- and 3'-untranslated regions exhibit low similarity, but similarity is high in the coding region. The lily PR-10 genes are induced by abscisic acid (ABA) and methyl jasmonate (MeJA) in the anther and various other organs of lily plants. The induction of PR-10 genes by ABA and MeJA in lily anthers occurs by two separate signal transduction pathways. The protein phosphatase inhibitor okadaic acid inhibits the MeJA-induced expression of PR-10 genes downstream of MeJA. In addition, the protein kinase inhibitor staurosporine inhibits the MeJA-induced expression of PR-10 genes, implying that an activity of staurosporine-sensitive protein kinases exists downstream of MeJA in the anther. However, okadaic acid does not inhibit the ABA-induced expression of PR-10 genes whereas staurosporine does. These observations suggest that, in addition to the known pathway that ABA induces gene expression by activating JA or MeJA, a MeJA-independent pathway of ABA induction exists in the anther. The alternative pathway of ABA induction involves a staurosporine-sensitive protein kinase activity downstream of ABA.     

Microarray Analysis of Gene Expression Involved in Anther Development in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

In flowering plants, anthers bear male gametophytes whose development is regulated by the elaborate coordination of many genes. In addition, both gibberellic acid (GA₃) and jasmonic acid (JA) play important roles in anther development and pollen fertility. To facilitate the analysis of anther development genes and how GA₃ and JA regulate anther development, we performed microarray experiments using a 10-K cDNA microarray with probes derived from seedlings, meiotic anthers, mature anthers and GA₃- or JA-treated suspension cells of rice. The expression level change of 2155 genes was significantly (by 2-fold or greater) detected in anthers compared with seedlings. Forty-seven genes, representing genes with potential function in cell cycle and cell structure regulation, hormone response, photosynthesis, stress resistance and metabolism, were differentially expressed in meiotic and mature anthers. Moreover, 314 genes responded to either GA₃ or JA treatment, and 24 GA₃- and 82 JA-responsive genes showed significant changes in expression between meiosis and the mature anther stages. RT-PCR demonstrated that gene y656d05 was not only highly expressed in meiotic anthers but also induced by GA₃. Strong RNA signals of y656d05 were detected in pollen mother cells and tapetum in in situ hybridization. Further characterization of these candidate genes can contribute to the understanding of the molecular mechanism of anther development and the involvement of JA and GA₃ signals in the control of anther development in rice.     

植物花药开裂的细胞学和分子生物学机制

植物花药开裂具有重要的生物学意义,花药开裂异常所导致的最直接后果为花粉粒不能正常散粉,影响到植物受精过程。现从细胞和分子生物学角度综述了植物花药开裂过程中花药组织的细胞结构和生理变化及调控花药开裂相关基因的分离和克隆。     

Functional Analysis of Wheat <Emphasis Type="Italic">TaPaO1</Emphasis> Gene Conferring Pollen Sterility Under Low Temperature

Thermosensitive male sterility plays an important role in wheat fertility and production. As a key enzyme for chlorophyll degradation, pheophorbide a oxygenase (PaO) can suppress cell death in plants. We cloned the wheat gene TaPaO1 from the thermosensitive genetic male sterile (TGMS) line BS366; it encodes a typical PaO protein, containing a conserved Rieske [2Fe-2S] iron–sulphur motif, a mononuclear non-heme iron-binding motif, and a C-terminal CxxC motif. TaPaO1 was expressed in all tissues and was upregulated during the meiosis stage of BS366 anthers under low temperature. Subcellular localization of TaPaO1 specifically labelled the surrounding of chloroplasts. TaPaO1 regulated by RD29A promoter which responded to low temperature led to pollen sterility in transgenic tobacco. Expression analysis showed that TaPaO1 exhibited a higher level of expression in the anther than in other tissues in transgenic tobacco plants during low temperature treatment. We propose that the higher senescence-related activity of TaPaO1 may lead to the cell death of anthers, which happens at an early developmental stage under low temperature. These results provide new insights into the function of PaO during the early developmental stage of anthers. PaO is closely related to cell death regardless of whether it exhibits increased activity or inactive.     

Epidermal jasmonate perception is sufficient for all aspects of jasmonate‐mediated male fertility in Arabidopsis

Jasmonate (JA) signaling is essential for several environmental responses and reproductive development in many plant species. In Arabidopsis thaliana, the most obvious phenotype of JA biosynthetic and perception mutants is profound sporophytic male sterility characterized by failure of stamen filament elongation, severe delay of anther dehiscence and pollen inviability. The site of action of JA in the context of reproductive development has been discussed, but the ideas have not been tested experimentally. To this end we used targeted expression of a COI1‐YFP transgene in the coi1‐1 mutant background. As COI1 is an essential component of the JA co‐receptor complex, the null coi1‐1 mutant is male sterile due to lack of JA perception. We show that expression of COI1‐YFP in the epidermis of the stamen filament and anther in coi1 mutant plants is sufficient to rescue filament elongation, anther dehiscence and pollen viability. In contrast, filament expression alone or expression in the tapetum do not restore dehiscence and pollen viability. These results demonstrate that epidermal JA perception is sufficient for anther function and pollen viability, and suggest the presence of a JA‐dependent non‐autonomous signal produced in the anther epidermis to synchronize both anther dehiscence and pollen maturation.     

小麦光温敏雄性不育系BS366铜锌超氧化物歧化酶基因的克隆及表达分析

超氧化物歧化酶(SOD,superoxide dismutase)是植物中一种主要的抗氧化酶,在植物应对逆境胁迫及抗衰老中起重要作用。本研究从基因芯片数据中筛选获得小麦Cu/Zn-SOD基因的EST序列,通过序列比对后拼接得到小麦Cu/Zn-SOD的候选基因,利用PCR技术在小麦光温敏雄性不育材料BS366中克隆并获得该基因。通过对Cu/Zn-SOD基因序列进行生物信息学分析,结果表明,该基因拥有连续且完整的开放阅读框,长495bp,编码164个氨基酸。氨基酸序列分析发现,该蛋白具有保守的Cu/Zn-SOD功能结构域与典型的Cu/Zn-SOD三维结构,且定位于细胞质中。通过同源进化分析表明,该蛋白与二穗短柄草(Brachypodium distachyon(L.)Beauv.)和大麦(Hordeum vulgare L.)的Cu/Zn-SOD蛋白亲缘关系较近,相似度分别为89%和94%。利用实时荧光定量PCR技术对其在小麦不同组织的表达特异性及不同逆境胁迫下的表达模式进行分析,结果表明,该基因在根、茎、叶、雌蕊、雄蕊、颖壳中均有表达,属于组成型表达,且在小麦的地上部含叶绿体的组织中含量较高;同时受多种胁迫诱导,可能参与了多种胁迫诱导调控途径。通过对该基因在不同育性环境中BS366育性转换期花药中的表达模式分析,发现可育环境下,在小孢子母细胞时期和减数分裂期的表达量分别约为对照的8倍与16倍;而不育环境下,该基因表达水平无明显变化。因而推测,小麦Cu/Zn-SOD基因可能参与了光温敏雄性不育系BS366的育性调控。本研究为深入研究Cu/Zn-SOD基因在小麦中的作用机理奠定了重要基础。