HAR1 mediates systemic regulation of symbiotic organ development

Symbiotic root nodules are beneficial to leguminous host plants; however, excessive nodulation damages the host because it interferes with the distribution of nutrients in the plant. To keep a steady balance, the nodulation programme is regulated systemically in leguminous hosts. Leguminous mutants that have lost this ability display a hypernodulating phenotype. Through the use of reciprocal and self-grafting studies using Lotus japonicus hypernodulating mutants, har1 (also known as sym78), we show that the shoot genotype is responsible for the negative regulation of nodule development. A map-based cloning strategy revealed that HAR1 encodes a protein with a relative molecular mass of 108,000, which contains 21 leucine-rich repeats, a single transmembrane domain and serine/threonine kinase domains. The har1 mutant phenotype was rescued by transfection of the HAR1 gene. In a comparison of Arabidopsis receptor-like kinases, HAR1 showed the highest level of similarity with CLAVATA1 (CLV1). CLV1 negatively regulates formation of the shoot and floral meristems through cell-cell communication involving the CLV3 peptide. Identification of hypernodulation genes thus indicates that genes in leguminous plants bearing a close resemblance to CLV1 regulate nodule development systemically, by means of organ-organ communication.     

Symbiotic host-specificity of Rhizobium meliloti is determined by a sulphated and acylated glucosamine oligosaccharide signal

Rhizobia are symbiotic bacteria that elicit the formation on leguminous plants of specialized organs, root nodules, in which they fix nitrogen. In various Rhizobium species, such as R. leguminosarum and R. meliloti, common and host-specific nodulation (nod) genes have been identified which determine infection and nodulation of specific hosts. Common nodABC genes as well as host-specific nodH and nodQ genes were shown recently, using bioassays, to be involved in the production of extracellular Nod signals. Using R. meliloti strains overproducing symbiotic Nod factors, we have purified the major alfalfa-specific signal, NodRm-1, by gel permeation, ion exchange and C18 reverse-phase high performance liquid chromatography. From mass spectrometry, nuclear magnetic resonance, (35)S-labelling and chemical modification studies, NodRm-1 was shown to be a sulphated beta-1,4-tetrasaccharide of D-glucosamine (Mr 1,102) in which three amino groups were acetylated and one was acylated with a C16 bis-unsaturated fatty acid. This purified Nod signal specifically elicited root hair deformation on the homologous host when added in nanomolar concentration.     

贵州省豆科结瘤固氮植物资源调查

通过对贵州省豆科植物结瘤固氮资源的调查,共采集根瘤标本242份,分属豆科植物42属103种及变种,其中蝶形花亚科、含羞草亚科、云实亚科植物的结瘤率分别为95%,75%和5%;采集的根瘤中,96%生于寄主的侧根,形状以球形、长柱形为主,颜色多为白色、褐色.对分离纯化的55株根瘤菌进行了回接试验,回接结瘤率为33%.生态条件是影响根瘤的大小、数量以及有效性的原因之一.     

黄腐酸对大豆内源激素的调控及结瘤能力的影响

为探讨黄腐酸提高大豆结瘤能力的作用机理,以辽宁慢生根瘤菌(Bradyrhizobium liaoningense)CCBAU 05525和冀豆17为材料,采用高效液相色谱法(HPLC)探究黄腐酸对大豆结瘤初期内源激素含量、分布及结瘤的影响.结果表明:黄腐酸质量浓度为500 mg/L时大豆结瘤数量最多,比对照组显著提高73.1%,同时该条件下激素含量和分布变化最为明显,具体表现为大豆根、茎、叶中玉米素(ZT)、生长素(IAA)、赤霉素(GA3)含量增加,根中脱落酸(ABA)含量减少,叶中ABA含量增加.而黄腐酸质量浓度为1 000 mg/L时则表现为抑制大豆结瘤. 上述结果为解析黄腐酸提高大豆结瘤固氮的机理、增加大豆产量提供了理论依据.     

Sinorhizobium meliloti lsrB is involved in alfalfa root nodule development and nitrogen-fixing bacteroid differentiation

Rhizobia interact with host legumes to induce the formation of nitrogen-fixing nodules, which is very important in agriculture and ecology. The development of nitrogen-fixing nodules is stringently regulated by host plants and rhizobial symbionts. In our previous work, a new Sinorhizobium meliloti LysR regulator gene (lsrB) was identified to be essential for alfalfa nodulation. However, how this gene is involved in alfalfa nodulation was not yet understood. Here, we found that this gene was associated with prevention of premature nodule senescence and abortive bacteroid formation. Heterogeneous deficient alfalfa root nodules were induced by the in-frame deletion mutant of lsrB (lsrB1-2), which was similar to the plasmid-insertion mutant, lsrB1. Irregular senescence zones earlier appeared in these nodules where bacteroid differentiation was blocked at different stages from microscopy observations. Interestingly, oxidative bursts were observed in these nodules by DAB staining. The decreased expression of lipopolysaccharide core genes (lpsCDE) was correspondingly determined in these nodules. S. meliloti lipopolysaccharide is required for suppression of oxidative bursts or host cell defense. These findings demonstrate that the S. meliloti lsrB gene is involved in alfalfa root nodule development and bacteroid differentiation by suppressing oxidative bursts or defense responses in host cells.     

Deregulation of a Ca2+/calmodulin-dependent kinase leads to spontaneous nodule development

Induced development of a new plant organ in response to rhizobia is the most prominent manifestation of legume root-nodule symbiosis with nitrogen-fixing bacteria. Here we show that the complex root-nodule organogenic programme can be genetically deregulated to trigger de novo nodule formation in the absence of rhizobia or exogenous rhizobial signals. In an ethylmethane sulphonate-induced snf1 (spontaneous nodule formation) mutant of Lotus japonicus, a single amino-acid replacement in a Ca2+/calmodulin-dependent protein kinase (CCaMK) is sufficient to turn fully differentiated root cortical cells into meristematic founder cells of root nodule primordia. These spontaneous nodules are genuine nodules with an ontogeny similar to that of rhizobial-induced root nodules, corroborating previous physiological studies. Using two receptor-deficient genetic backgrounds we provide evidence for a developmentally integrated spontaneous nodulation process that is independent of lipochitin-oligosaccharide signal perception and oscillations in Ca2+ second messenger levels. Our results reveal a key regulatory position of CCaMK upstream of all components required for cell-cycle activation, and a phenotypically divergent series of mutant alleles demonstrates positive and negative regulation of the process.     

百脉根Rac1基因启动子的克隆与表达分析

百脉根小G蛋白Rac1基因促进共生结瘤过程,但其转录调控机制尚不明确.采用生物信息学方法对百脉根Rac1基因的启动子序列进行了预测,并对该启动子中含有的顺式调控元件进行了统计分析.克隆了约1.8kb的启动子片段,并构建了GUS融合的重组质粒p1391Z-Rac1Pro.通过百脉根毛根转化法获得转基因毛根,进一步利用组织化学染色法对Rac1基因在阳性毛根中的表达部位进行了研究.结果显示:该启动子除含有常见的转录调控保守元件TATA-box和CAAT-box外,还含有调控防御和胁迫、激素、光照等信号的应答元件.组织化学染色发现,Rac1基因在接种根瘤菌的根毛、根尖、根瘤原基皮层中表达量较高.     

A receptor kinase gene regulating symbiotic nodule development

Leguminous plants are able to establish a nitrogen-fixing symbiosis with soil bacteria generally known as rhizobia. Metabolites exuded by the plant root activate the production of a rhizobial signal molecule, the Nod factor, which is essential for symbiotic nodule development. This lipo-chitooligosaccharide signal is active at femtomolar concentrations, and its structure is correlated with host specificity of symbiosis, suggesting the involvement of a cognate perception system in the plant host. Here we describe the cloning of a gene from Medicago sativa that is essential for Nod-factor perception in alfalfa, and by genetic analogy, in the related legumes Medicago truncatula and Pisum sativum. The identified 'nodulation receptor kinase', NORK, is predicted to function in the Nod-factor perception/transduction system (the NORK system) that initiates a signal cascade leading to nodulation. The family of 'NORK extracellular-sequence-like' (NSL) genes is broadly distributed in the plant kingdom, although their biological function has not been previously ascribed. We suggest that during the evolution of symbiosis an ancestral NSL system was co-opted for transduction of an external ligand, the rhizobial Nod factor, leading to development of the symbiotic root nodule.     

Sinorhizobium meliloti nifA gene exerts a pleiotropic effect on nodulation through the enhanced plant defense response

Sinorhizobium meliloti nifA gene is required for the expression of a bunch of nif and fix genes. Here, we report its pleiotropic effects on the nodule formation. Compared with wild type strain, nifA mutant sig- nificantly reduced nodule suppression rate in split-root system. The plants inoculated with mutant strain produced lower amount of daidzein and less necrotic cells on their roots. In addition, the defense genes failed to be evoked by nifA mutant at the early nodulation stage. These findings indicated that host defense response was one of the mechanisms mediated by nifA gene to regulate nodule formation during symbiosis. Even though nifA mutant could increase the number of nodules in host plant, it synthesized lower Nod factors than wild type. This suggested that nifA gene mediated multiple and diverse instances in nodulation formation.     

MtROP8 is involved in root hair development and the establishment of symbiotic interaction between Medicago truncatula and Sinorhizobium meliloti

Emerging evidence has demonstrated that ROP GTPases play important roles in symbiosis, but the molecular mechanisms on their regulation in symbiosis are largely unknown. In this study, we showed that MtROP8 is involved in the symbiotic interaction between Medicago truncatula and Sinorhizobium meliloti. Expression analyses showed that MtROP8 was down-regulated in the early infected roots, but significantly up-regulated in nodules compared to the roots. Phenotypic analysis of RNA inter- ference （RNAi）-mediated silencing of MtROP8 revealed that knock-down of MtROP8 expression resulted in various developmental defects of root hairs, including branched hairs, short bulbous root hairs, and even root hairs with apparent swollen bases, which were caused by the modi- fication of the distribution and the levels of reactive oxygen species （ROS）. Moreover, infection events were increased in transgenic roots harboring the MtROP8 RNAi construct in response to S. meliloti inoculation, concomitant with enhanced nodulation. These results indicate that MtROP8 participates in root hair development and the establishmentof the symbiotic interaction by regulating ROS production and distribution.     

豆科植物-根瘤菌共生固氮的分子机理

与豆科植物－根瘤菌共生固氮有关的基因涉及根瘤菌基因和宿主基因，根瘤菌基因有结瘤基因（nodD,nodAB-CIJ和hsn基因），根瘤菌细胞表面结构基因（exs,lps和ndv基因）和固氮基因（nif和fix基因）；宿主基因主要是结瘤素基因（ENOD和NOD基因）。根瘤菌结瘤基因表达后诱导产生结瘤因子。在根瘤发育过程中，这些基因在根瘤菌与植物之间进行着信息交换，并且具有不同的表达水平。结瘤因子和植物激素对它们进行着调节。     

Identification of genes induced during Medicago sativa nodule development by using the cDNA-AFLP technique

Under limited nitrogen conditions, rhizobia are ableto induce the formation of nitrogen-fixing root nodules on their leguminous plant host. This organogenetic process is triggered by a complex exchange of molecu- lar signals between the host plant and bac…     

A plant receptor-like kinase required for both bacterial and fungal symbiosis

Most higher plant species can enter a root symbiosis with arbuscular mycorrhizal fungi, in which plant carbon is traded for fungal phosphate. This is an ancient symbiosis, which has been detected in fossils of early land plants. In contrast, the nitrogen-fixing root nodule symbioses of plants with bacteria evolved more recently, and are phylogenetically restricted to the rosid I clade of plants. Both symbioses rely on partially overlapping genetic programmes. We have identified the molecular basis for this convergence by cloning orthologous SYMRK ('symbiosis receptor-like kinase') genes from Lotus and pea, which are required for both fungal and bacterial recognition. SYMRK is predicted to have a signal peptide, an extracellular domain comprising leucine-rich repeats, a transmembrane and an intracellular protein kinase domain. Lotus SYMRK is required for a symbiotic signal transduction pathway leading from the perception of microbial signal molecules to rapid symbiosis-related gene activation. The perception of symbiotic fungi and bacteria is mediated by at least one common signalling component, which could have been recruited during the evolution of root nodule symbioses from the already existing arbuscular mycorrhiza symbiosis.     

诱导不结瘤豆科树种结瘤固氮的研究

根据国内外有关诱导非豆科作物结瘤固氮的现状,结合作者几年来的工作,对诱导不结瘤豆科树种结瘤固氮的可行性进行了探讨,并报道了作者近年来人工诱导不结瘤的豆科树种—山皂荚(Gledtsia melanacantha)的试验结果。     

氮_磷_钾对异果山绿豆结瘤固氮特性的影响

本试验发现磷对异果山绿豆结瘤、固氮和生长都起到极其重要的作用.含磷者结瘤多、根瘤发育正常固氮活性高,植株生长良好;而缺磷者根瘤小而白、植株生长差.缺磷钾时,少量化合态氮[0.61mM Ca(NO_3)_2或1.87mM NH_4Cl]对植物生长结瘤都引起有害的作用.在磷钾水平一致时,低水平化合态氮〔0.61mM Ca(NO_3)_2或1.87mM NH_4Cl〕对异果山绿豆结瘤固氮及生长都是有益的;而中水平氮〔3.05mM Ca(NO_3)_2或9.35mM NH_4Cl〕则不利;高水平氮〔6.1m MCa(NO_3)_2或18.7mM NH_4Cl〕有明显的抑制作用.植株含氮量随着化合态氮的水平提高而降低.     

Extra-copy nifA enhances the nodulation efficiency of Sinorhizobium fredii

Previous investigations have shown that nifA gene is involved in nodulation and symbiotic nitrogen fixation regulation of Rhizobium. We study the role of nifA on nodulation of leguminous plants. We found that Sinorhizobium fredii harboring multi-copy plasmid carrying the constitutively expressed Klebsiella pneumoniae nifA exhibited an increase of noduiation activity and nodulation competitiveness on soybean plants. The Nod-factor secreted by the rhizobia cells containing the multi-copied nifA was assayed,and preliminary results showed that S. fredii containing the multi-copy plasmid carrying nifA produced higher strength of Nod-factor than the rhizobia containing the same plasmid carrying the vector did.     

中国沙棘营养器官结构特征及其与生境关系的研究

对中国沙棘初生结构的分化和次生生长过程进行了观察。发现沙棘根系角质层发达，外被粘质物，外皮层细胞的细胞壁大量栓质化。根瘤的形成与木质部和皮层有密切关系，弗兰克氏菌的侵入刺激皮层形成大量的薄壁细胞，向外突出形成根瘤。茎的皮层外部细胞较小，排列整齐，细胞壁厚，表皮外有很厚的角质层和一定量的蜡质。叶的结构更加体现了抗旱、耐寒植物的特点，栅栏组织发达，排列紧密，海绵组织不发达，细胞间隙较大，气孔深陷，且具有角质层和密而叠置的星状毛及鳞片等。这些均为生活在旱、寒环境里表现出的适应性。