The role of Nod signal structures in the determination of host specificity in the Rhizobium-legume symbiosis

During the past five years the structure of nodulation signals from more than a dozen different Rhizobium species has been elucidated. In addition, the role of numerous nod genes in the biosynthesis of the lipooligosaccharides has been identified. This review discusses how Nod signal structure is determined by the specificity of the various biosynthetic steps and how this influences variation in host specificity. Until recently, it appeared that the decorations of a common lipochitooligosaccharide core determine the host-specific recognition of the signals, possibly via specific receptors in the host plant cell. A number of recent publications, however, suggest that beyond the interaction of Nod signals with a putative receptor, certain structural features of Nod factors are involved in controlling the concentration of the signals during their uptake by the root tissue.The authors are with the Institut des Sciences Végétales, Centre National de la Recherche Scientifique, Avenue de la Terrasse, F-91198 Gif-sur-Yvette, France; A. Kondorosi is also with the Institute of Genetics, Biological Research Center, Hungarian Academy of Sciences P.O Box 521, H-6701 Szeged, Hungary.     

Recent advances in Rhizobium-legume symbiosis

The research findings in the field of Rhizobium-legume symbiosis reported worldwide during the years 2002 and 2003 (up to September) have been summarized. The information is presented under the various topics, viz., isolation and characterization of rhizobial strains, physiological aspects of nitrogen fixation, rhizosphere interactions and root surface signals, genomics and proteomics, plant genes involved in nodule formation, bioremediation and biocontrol, and review articles and conference reports. The postal and e-mail addresses of the concerned scientists have also been included.     

Bacteroid formation in the Rhizobium-legume symbiosis

During the Rhizobium-legume symbiosis, bacteria enter the cells of host plants and differentiate into nitrogen-fixing bacteroids. Recent mutant screens and expression studies have revealed bacterial genes involved in the developmental pathway and demonstrate how the genetic requirements can vary from one host-microbe system to another. Whether bacteroids are terminally differentiated cells is an ongoing debate and new experimental systems are required to address this issue.     

Proteomics: a novel approach to explore signal exchanges in Rhizobium-legume symbiosis

Recent developments and future strategies on the proteomics approach to explore the signal exchanges in Rhizobium-legume symbiosis have been discussed. It is expected that this approach will provide new possibilities for investigating the complex interactions of rhizobia and legumes.     

Evolution of signal transduction in intracellular symbiosis

Plant roots form intracellular symbioses with fungi and bacteria resulting in arbuscular mycorrhiza and nitrogen-fixing root nodules, respectively. A novel receptor like-kinase has been discovered that is required for the transduction of both bacterial and fungal symbiotic signals. This kinase defines an ancient signalling pathway that probably evolved in the context of arbuscular mycorrhiza and has been recruited subsequently for endosymbiosis with bacteria. An ancestral symbiotic interaction of roots with intracellular bacteria might have emerged from such a recruitment, in the progenitor of the nodulating clade of plants. Analysis of symbiotic mutants of host plants and bacterial microsymbionts has revealed that present-day endosymbioses require the coordinated induction of more than one signalling pathway for development.     

Nodulation competitiveness in the Rhizobium-legume symbiosis

Successful nodulation of legumes by rhizobia is a complex process that, in the open field, depends on many different environmental factors. Generally, legume productivity in an agricultural field may be improved by inoculation with selected highly effective N₂-fixing root nodule bacteria. However, field legume inoculation with Rhizobium and Bradyrhizobium spp. has often been unsuccessful because of the presence in the soil of native strains that compete with the introduced strain in nodule formation on the host plants. This ability to dominate nodulation is termed competitiveness and is critical for the successful use of inoculants.The author is with the Departmentode Microbiologia del Suelo y Sistemas Simbioticos, Estation Experimental del Zaidin, Consejo Superior de Investigaciones Cientificas, C/Professor Albareda 1, 18008 Granada, Spain     

Calcium and signal transduction in plants

Environmental and hormonal signals control diverse physiological processes in plants. The mechanisms by which plant cells perceive and transduce these signals are poorly understood. Understanding biochemical and molecular events involved in signal transduction pathways has become one of the most active areas of plant research. Research during the last 15 years has established that Ca2+ acts as a messenger in transducing external signals. The evidence in support of Ca2+ as a messenger is unequivocal and fulfills all the requirements of a messenger. The role of Ca2+ becomes even more important because it is the only messenger known so far in plants. Since our last review on the Ca2+ messenger system in 1987, there has been tremendous progress in elucidating various aspects of Ca(2+) -signaling pathways in plants. These include demonstration of signal-induced changes in cytosolic Ca2+, calmodulin and calmodulin-like proteins, identification of different Ca2+ channels, characterization of Ca(2+) -dependent protein kinases (CDPKs) both at the biochemical and molecular levels, evidence for the presence of calmodulin-dependent protein kinases, and increased evidence in support of the role of inositol phospholipids in the Ca(2+) -signaling system. Despite the progress in Ca2+ research in plants, it is still in its infancy and much more needs to be done to understand the precise mechanisms by which Ca2+ regulates a wide variety of physiological processes. The purpose of this review is to summarize some of these recent developments in Ca2+ research as it relates to signal transduction in plants.     

Plant genes induced in the Rhizobium-legume symbiosis

Rhizobium, Bradyrhizobium and Azorhizobium can elicit the formation of N₂-fixing nodules on the roots or stems of their leguminous host plants. The nodule formation involves several developmental steps determined by different sets of genes from both partners, the gene expression being temporally and spatially coordinated. The plant proteins that are specifically synthesised during the formation and function of the nodule are called nodulins. The nodulins that are expressed before the onset of N₂ fixation are termed early nodulins. These proteins are probably involved in the infection process as well as in nodule morphogenesis rather than in nodule function. The nodulins expressed just before or during N₂ fixation are termed late nodulins and they participate in the function of the nodule by creating the physiological conditions required for nitrogen fixation, ammonium assimilation and transport. In this review we will describe nodulins, nodulin genes and the relationship between nodulin gene expression and nodule development. The study of nodulin gene expression may provide insight into root-nodule development and the mechanism of communication between bacteria and host plant.J.A. Muñoz and A.J. Palomares are with the Departamento de Microbiologia y Parasitología, Facultad de Farmacia, Universidad de Sevilla, 41012 Sevilla, Spain. P. Ratet is with the Institut des Sciences Végétales, CNRS, Avenue de la Terrasse, F-91198 Gif-sur-Yvette, Fance     

Calcium signal transduction from caveolae

Caveolae are specialized membrane microdomains that are found on the plasma membrane of most cells. Recent studies indicate that a variety of signaling molecules are highly organized in caveolae, where their interactions initiate specific signaling cascades. Molecules enriched in this membrane include G protein-coupled receptors, heterotrimeric GTP binding proteins, IP3 receptor-like protein, Ca2+ ATPase, eNOS, and several PKC isoforms. Direct measurements of calcium changes in endothelial cells suggest that caveolae may be sites that regulate intracellular Ca2+ concentration and Ca2+ dependent signal transduction. This review will focus on the role of caveolae in controlling the spatial and temporal pattern of intracellular Ca2+ signaling.     

Simulation of marked root hair curling in Rhizobium-legume symbiosis

The soil bacterium Rhizobium infects its leguminous host plants in temperate regions of the world mostly by way of the growing root hairs. Root hair curling is a prerequisite for root hair infection, although sidelong root hair infections occasionally have been observed. The processes underlying Rhizobium -induced root hair curling are unknown.
Computer simulation of root hair growth indicates that one-sided tip growth inhibition by Rhizobium can result in root hair curling when three conditions are simultaneously fulfilled: 1) rhizobial growth inhibition is strong enough to prevent removal out of the tip growth range: 2) root hair surface growth between the attached Rhizobium and the root hair top is inhibited; 3) rhizobial growth inhibition is limited to one side of the root hair.
The results predict that root hair curling by stimulation of tip growth is improbable. This study accentuates the need for information about the growth processes contributing to tip growth in leguminous root hairs.     

Calcium signals in growth factor signal transduction

There is a substantial amount of information which has been obtained concerning the effects of growth factors on [Ca²⁺]_i in proliferating cells. A number of different mitogens are known to induce elevations in [Ca²⁺]_i and some characterization of the Ca²⁺ response to different classes of mitogens has been obtained. In addition, much is known about whether the Ca²⁺ response to a particular growth factor occurs as the result of an influx of external Ca²⁺ or a mobilization of internal Ca²⁺ stores. In addition, a considerable amount of information is available on the mechanism by which the Ins(1,4,5)P₃-sensitive internal Ca²⁺ store takes up and releases Ca²⁺. However, there is still a large deficiency in our information concerning other Ca²⁺ stores in proliferating cells as well as in our knowledge of the mechanisms for regulating Ca²⁺ entry pathways. Much more data addressing these issues exists for other types of agonist-stimulated cells, and we have discussed much of it in this review article. While the wealth of data in nonproliferating cells provides some indications of what mechanisms might be involved in the growth factor-induced changes in [Ca²⁺]_i, it is clear that much work must be done in proliferating cells to fully understand how external factors such as growth factors control [Ca²⁺]_i. In addition, much work remains to be done in identifying the mechanisms for the internal control of [Ca²⁺]_i as cells move through the cell cycle and in identifying the role that these changes in [Ca²⁺]_i may play throughout the cell cycle.     

Calcium channels and signal transduction in plant cells

An increasing number of studies indicate that changes in cytosolic free Ca²⁺ ([Ca²⁺]_c) mediate specific types of signal transduction in plant cells. Modulation of [Ca²⁺]_c is likely to be achieved through changes in the activity of Ca²⁺ channels, which catalyse passive influx of Ca²⁺ to the cytosol from extracellular and intracellular compartments. Voltage-sensitive Ca²⁺ channels have been detected in the plasma membranes of algae, where they control membrane electrical properties and cell turgor. These channels are sensitive to 1,4-dihydropyridines, which in animal cells specifically affect one class of voltage-regulated plasma membrane Ca²⁺ channel. Ca²⁺-permeable channels with different pharmacological properties have been found in the plasma membrane of higher plants. Recent evidence suggests the existence of two discrete classes of Ca²⁺ channel co-resident in the vacuolar membrane (tonoplast) of higher plants. The first is gated by inositol 1,4,5-trisphosphate, and bears a number of similarities to its animal counterpart which is located in the endoplasmic reticulum (ER). The second tonoplast Ca²⁺ channel is voltage-operated. However, the specific roles of these tonoplast channels in signal transduction have yet to be elucidated.     

GPCR和CaCC信号传导中的钙信号网络

G蛋白偶联受体(G protein-coupled receptor,GPCR)作为最大的一类人膜蛋白受体家族和最重要的药物靶标而倍受关注,其中钙离子在细胞内信号传导级联放大中起了关键的作用。阐述了GPCR和钙激活的氯离子通道蛋白(calcium-activated chloride channel,CaCC)中的钙信号网络与生理功能以及如何干扰阻断该网络,为药物设计和很多疾病的治疗提供了依据。     

Ethylene inhibits the Nod factor signal transduction pathway of Medicago truncatula

Legumes form a mutualistic symbiosis with bacteria collectively referred to as rhizobia. The bacteria induce the formation of nodules on the roots of the appropriate host plant, and this process requires the bacterial signaling molecule Nod factor. Although the interaction is beneficial to the plant, the number of nodules is tightly regulated. The gaseous plant hormone ethylene has been shown to be involved in the regulation of nodule number. The mechanism of the ethylene inhibition on nodulation is unclear, and the position at which ethylene acts in this complex developmental process is unknown. Here, we used direct and indirect ethylene application and inhibition of ethylene biosynthesis, together with comparison of wild-type plants and an ethylene-insensitive supernodulating mutant, to assess the effect of ethylene at multiple stages of this interaction in the model legume Medicago truncatula. We show that ethylene inhibited all of the early plant responses tested, including the initiation of calcium spiking. This finding suggests that ethylene acts upstream or at the point of calcium spiking in the Nod factor signal transduction pathway, either directly or through feedback from ethylene effects on downstream events. Furthermore, ethylene appears to regulate the frequency of calcium spiking, suggesting that it can modulate both the degree and the nature of Nod factor pathway activation.     

Regulation of signal transduction and bacterial infection during root nodule symbiosis

Among plant-microbe interactions, root nodule symbiosis is one of the most important beneficial interactions providing legume plants with nitrogenous compounds. Over the past years a number of genes required for root nodule symbiosis has been identified but most recently great advances have been made to dissect signalling pathways and molecular interactions triggered by a set of receptor-like kinases. Genetic and biochemical approaches have not only provided evidence for the cross talk between bacterial infection of the host plant and organogenesis of a root nodule but also gained insights into dynamic regulation processes underlying successful infection events. Here, we summarise recent progress in the understanding of molecular mechanisms that regulate and trigger cellular signalling cascades during this mutualistic interaction.     

Evidence for structurally specific negative feedback in the Nod factor signal transduction pathway

Nod factor is a critical signalling molecule in the establishment of the legume/rhizobial symbiosis. The Nod factor of Sinorhizobium meliloti carries O-sulphate, O-acetate and C16:2 N-acyl attachments that define its activity and host specificity. Here we assess the relative importance of these modifications for the induction of calcium spiking in Medicago truncatula. We find that Nod factor structures lacking the O-sulphate, structures lacking the O-acetate and N-acyl groups, and structures lacking the O-acetate combined with a C18:1 N-acyl group all show calcium spiking when applied at high concentrations. These calcium responses are blocked in dmi1 and dmi2 mutants, suggesting that they function through the Nod factor signal transduction pathway. The dmi3 mutant, which is proposed to function in the Nod factor signal transduction pathway downstream of calcium spiking, shows increased sensitivity to Nod factor. This increased sensitivity is only active with wild-type Nod factor and was not present when the plants were treated with mutant Nod factor structures. We propose that the Nod factor signal transduction pathway is under negative feedback regulation that is activated at or downstream of DMI3 and requires structural components of the Nod factor molecule for activity.     

Nod genes and Nod signals and the evolution of the Rhizobium legume symbiosis.

The establishment of the nitrogen-fixing symbiosis between rhizobia and legumes requires an exchange of signals between the two partners. In response to flavonoids excreted by the host plant, rhizobia synthesize Nod factors (NFs) which elicit, at very low concentrations and in a specific manner, various symbiotic responses on the roots of the legume hosts. NFs from several rhizobial species have been characterized. They all are lipo-chitooligosaccharides, consisting of a backbone of generally four or five glucosamine residues N-acylated at the non-reducing end, and carrying various O-substituents. The N-acyl chain and the other substituents are important determinants of the rhizobial host specificity. A number of nodulation genes which specify the synthesis of NFs have been identified. All rhizobia, in spite of their diversity, possess conserved nodABC genes responsible for the synthesis of the N-acylated oligosaccharide core of NFs, which suggests that these genes are of a monophyletic origin. Other genes, the host specific nod genes, specify the substitutions of NFs. The central role of NFs and nod genes in the Rhizobium-legume symbiosis suggests that these factors could be used as molecular markers to study the evolution of this symbiosis. We have studied a number of NFs which are N-acylated by alpha,beta-unsaturated fatty acids. We found that the ability to synthesize such NFs does not correlate with taxonomic position of the rhizobia. However, all rhizobia that produce NFs such nodulate plants belonging to related tribes of legumes, the Trifolieae, Vicieae, and Galegeae, all of them being members of the so-called galegoid group. This suggests that the ability to recognize the NFs with alpha-beta-unsaturated fatty acids is limited to this group of legumes, and thus might have appeared only once in the course of legume evolution, in the galegoid phylum.     

Structure-function analysis of nod factor-induced root hair calcium spiking in Rhizobium-legume symbiosis

In the Rhizobium-legume symbiosis, compatible bacteria and host plants interact through an exchange of signals: Host compounds promote the expression of bacterial biosynthetic nod (nodulation) genes leading to the production of a lipochito-oligosaccharide signal, the Nod factor (NF). The particular array of nod genes carried by a given species of Rhizobium determines the NF structure synthesized and defines the range of legume hosts by which the bacterium is recognized. Purified NF can induce early host responses even in the absence of live Rhizobium One of the earliest known host responses to NF is an oscillatory behavior of cytoplasmic calcium, or calcium spiking, in root hair cells, initially observed in Medicago spp. and subsequently characterized in four other genera (D.W. Ehrhardt, R. Wais, S.R. Long [1996] Cell 85: 673-681; S.A. Walker, V. Viprey, J.A. Downie [2000] Proc Natl Acad Sci USA 97: 13413-13418; D.W. Ehrhardt, J.A. Downie, J. Harris, R.J. Wais, and S.R. Long, unpublished data). We sought to determine whether live Rhizobium trigger a rapid calcium spiking response and whether this response is NF dependent. We show that, in the Sinorhizobium meliloti-Medicago truncatula interaction, bacteria elicit a calcium spiking response that is indistinguishable from the response to purified NF. We determine that calcium spiking is a nod gene-dependent host response. Studies of calcium spiking in M. truncatula and alfalfa (Medicago sativa) also uncovered the possibility of differences in early NF signal transduction. We further demonstrate the sufficiency of the nod genes for inducing calcium spiking by using Escherichia coli BL21 (DE3) engineered to express 11 S. meliloti nod genes.