Leguminous plants (such as peas and soybeans) and rhizobial soil bacteria are symbiotic partners that communicate through molecular signaling pathways, resulting in the formation of nodules on legume roots and occasionally stems that house nitrogen-fixing bacteria. Nodule formation has been assumed to be exclusively initiated by the binding of bacterial, host-specific lipochito-oligosaccharidic Nod factors, encoded by the nodABC genes, to kinase-like receptors of the plant. Here we show by complete genome sequencing of two symbiotic, photosynthetic, Bradyrhizobium strains, BTAi1 and ORS278, that canonical nodABC genes and typical lipochito-oligosaccharidic Nod factors are not required for symbiosis in some legumes. Mutational analyses indicated that these unique rhizobia use an alternative pathway to initiate symbioses, where a purine derivative may play a key role in triggering nodule formation.

The mechanisms involved in the posttranslational targeting of membrane proteins are not well understood. The light-harvesting chlorophyll proteins (LHCP) of the thylakoid membrane are a large family of hydrophobic proteins that are targeted in this manner. They are synthesized in the cytoplasm, translocated across the chloroplast envelope membranes into the stroma, bound by a stromal factor to form a soluble intermediate, ''transit complex'', and then integrated into the thylakoid membrane by a GTP dependent reaction. Signal recognition particle (SRP), a cytoplasmic ribonucleoprotein, is known to mediate the GTP dependent cotranslational targeting of proteins to the endoplasmic reticulum. We show that chloroplasts contain an SRP consisting of, cpSRP54, a homologue of SRP54 and a previously undescribed 43-kDa polypeptide (cpSRP43) instead of an RNA. We demonstrate that both subunits of cpSRP are required for the formation of the transit complex with LHCP. Furthermore, cpSRP54, cpSRP43, and LHCP are sufficient to form a complex that appears to be identical to authentic transit complex. We also show that the complex formed between LHCP and cpSRP, together with an additional soluble factor(s) are required for the proper integration of LHCP into the thylakoid membrane. It appears that the expanded role of cpSRP in posttranslational targeting of LHCP has arisen through the evolution of the 43-kDa protein.The insertion of proteins into membranes is a fundamental process essential for the vitality of all organisms. The paradigm for this process is the targeting mediated by signal recognition particle (SRP), a cytoplasmic ribonucleoprotein. In prokaryotes, the SRP-RNA binds a single 54-kDa-polypeptide subunit, while in eukaryotes, five additional subunits are bound. One of the distinctive features of this targeting mechanism is cotranslational protein insertion. The synthesis of hydrophobic protein domains at the membrane circumvents potential protein folding problems that might otherwise occur in an aqueous environment. However not all hydrophobic proteins are targeted cotranslationally. For example, the major proteins of the thylakoid membrane, the light harvesting chlorophyll proteins (LHCP), are targeted by a posttranslational mechanism. LHCP form a large family of related proteins that have three to four transmembrane domains. They are synthesized in the cytoplasm, and are targeted to the thylakoid membrane through three aqueous compartments: the cytoplasm, the inter-envelope space, and the stroma (1, 2). The factors that mediate the posttranslational targeting of members of the LHCP family have not been definitively identified.The LHCP are inserted into the thylakoid membrane in a reaction requiring GTP and stroma (3-5). It was found previously that a stromal factor binds LHCP to form a soluble intermediate, designated transit complex, that maintains the solubility of these proteins as they are transported through the stroma (6, 7). The transit complex is the only soluble form of LHCP that accumulates when ...

Colonization of the rhizosphere by micro-organisms results in modifications in plant growth and development. This review examines the mechanisms involved in growth promotion by plant growth-promoting rhizobacteria which are divided into indirect and direct effects. Direct effects include enhanced provision of nutrients and the production of phytohormones. Indirect effects involve aspects of biological control: the production of antibiotics and ironchelating siderophores and the induction of plant resistance mechanisms. The study of the molecular basis of growth promotion demonstrated the important role of bacterial traits (motility, adhesion and growth rate) for colonization. New research areas emerge from the discovery that molecular signalling occurs through plant perception of eubacterial flagellins. Recent perspectives in the molecular genetics of cross-talking mechanisms governing plant-rhizobacteria interactions are also discussed.

Nutritional symbiotic interactions require the housing of large numbers of microbial symbionts, which produce essential compounds for the growth of the host. In the legume-rhizobium nitrogen-fixing symbiosis, thousands of rhizobium microsymbionts, called bacteroids, are confined intracellularly within highly specialized symbiotic host cells. In Inverted Repeat-Lacking Clade (IRLC) legumes such as Medicago spp., the bacteroids are kept under control by an arsenal of nodulespecific cysteine-rich (NCR) peptides, which induce the bacteria in an irreversible, strongly elongated, and polyploid state. Here, we show that in Aeschynomene spp. legumes belonging to the more ancient Dalbergioid lineage, bacteroids are elongated or spherical depending on the Aeschynomene spp. and that these bacteroids are terminally differentiated and polyploid, similar to bacteroids in IRLC legumes. Transcriptome, in situ hybridization, and proteome analyses demonstrated that the symbiotic cells in the Aeschynomene spp. nodules produce a large diversity of NCR-like peptides, which are transported to the bacteroids. Blocking NCR transport by RNA interference-mediated inactivation of the secretory pathway inhibits bacteroid differentiation. Together, our results support the view that bacteroid differentiation in the Dalbergioid clade, which likely evolved independently from the bacteroid differentiation in the IRLC clade, is based on very similar mechanisms used by IRLC legumes.

A recessive mutation in Arabidopsis, named chaos (for chlorophyll a/b binding protein harvesting-organelle specific; designated gene symbol CAO ), was isolated by using transposon tagging. Characterization of the phenotype of the chaos mutant revealed a specific reduction of pigment binding antenna proteins in the thylakoid membrane. These nuclear-encoded proteins utilize a chloroplast signal recognition particle (cpSRP) system to reach the thylakoid membrane. Both prokaryotes and eukaryotes possess a cytoplasmic SRP containing a 54-kD protein (SRP54) and an RNA. In chloroplasts, the homolog of SRP54 was found to bind a 43-kD protein (cpSRP43) rather than to an RNA. We cloned the CAO gene, which encodes a protein identified as Arabidopsis cpSRP43. The product of the CAO gene does not resemble any protein in the databases, although it contains motifs that are known to mediate protein-protein interactions. These motifs include ankyrin repeats and chromodomains. Therefore, CAO encodes an SRP component that is unique to plants. Surprisingly, the phenotype of the cpSRP43 mutant (i.e., chaos ) differs from that of the Arabidopsis cpSRP54 mutant, suggesting that the functions of the two proteins do not strictly overlap. This difference also suggests that the function of cpSRP43 is most likely restricted to protein targeting into the thylakoid membrane, whereas cpSRP54 may be involved in an additional process(es), such as chloroplast biogenesis, perhaps through chloroplast-ribosomal association with chloroplast ribosomes. INTRODUCTIONHigher plants require chloroplasts for essential functions ranging from photosynthesis to sugar, lipid, and amino acid biosyntheses. Many of the proteins required for the light reactions of photosynthesis, including the chlorophyll-containing antenna proteins, are localized in the thylakoid membrane. Thylakoid proteins are encoded by both the nuclear and chloroplast genomes. Nuclear-encoded proteins destined for the thylakoid membrane are synthesized with a cleavable N-terminal extension (transit peptide) that targets the protein across the envelope membranes into the stroma (reviewed in Cline and Henry, 1996;Kermode, 1996;Lübeck et al., 1997). Multiple mechanisms exist for targeting proteins from the stroma to the thylakoid membrane (reviewed in Cline and Henry, 1996;Klösgen, 1997). These include spontaneous insertion, a chloroplast Sec r -dependent pathway, a chloroplast signal recognition particle (cpSRP) pathway, and a pathway requiring only an electrochemical potential of ⌬ pH across the thylakoid membrane. These pathways can be distinguished by specific requirements for stromal proteins and/or by energetic requirements when in 1 These authors contributed equally to this work. 2 To whom correspondence should be addressed: E-mail lnussaume @cea.fr; fax 33-442-25-4656. 88The Plant Cell vitro reconstitution assays are performed. In vitro, precursor proteins are targeted exclusively via one of the aforementioned pathways.LHCIIb proteins are the most abundant of the thylakoid membrane ...

SignificanceLegumes have a tremendous ecological and agronomic importance due to their ability to interact symbiotically with nitrogen-fixing rhizobia. In most of the rhizobial–legume symbioses, the establishment of the interaction requires the plant perception of the bacterial lipochitooligosaccharide Nod factor signal. However, some bradyrhizobia can activate the symbiosis differently, thanks to their type III secretion system, which delivers effector proteins into the host cell. Here, we demonstrate that this symbiotic process relies on a small set of effectors playing distinct and complementary roles. Most remarkably, a nuclear-targeted effector named ErnA conferred the ability to form nodules. The understanding of this alternative pathway toward nitrogen-fixing symbiosis could pave the way for designing new strategies to transfer nodulation into cereals.

Research on the nitrogen-fixing symbiosis has been focused, thus far, on two model legumes, Medicago truncatula and Lotus japonicus, which use a sophisticated infection process involving infection thread formation. However, in 25% of the legumes, the bacterial entry occurs more simply in an intercellular fashion. Among them, some Aeschynomene spp. are nodulated by photosynthetic Bradyrhizobium spp. that do not produce Nod factors. This interaction is believed to represent a living testimony of the ancestral state of the rhizobium-legume symbiosis. To decipher the mechanisms of this Nod-independent process, we propose Aeschynomene evenia as a model legume because it presents all the characteristics required for genetic and molecular analysis. It is a short-perennial and autogamous species, with a diploid and relatively small genome (2n=20; 460 Mb/1C). A. evenia 'IRFL6945' is nodulated by the well-characterized photosynthetic Bradyrhizobium sp. strain ORS278 and is efficiently transformed by Agrobacterium rhizogenes. Aeschynomene evenia is genetically homozygous but polymorphic accessions were found. A manual hybridization procedure has been set up, allowing directed crosses. Therefore, it should be relatively straightforward to unravel the molecular determinants of the Nod-independent process in A. evenia. This should shed new light on the evolution of rhizobium-legume symbiosis and could have important agronomic implications.

Among legumes (Fabaceae) capable of nitrogen-fixing nodulation, several Aeschynomene spp. use a unique symbiotic process that is independent of Nod factors and infection threads. They are also distinctive in developing root and stem nodules with photosynthetic bradyrhizobia. Despite the significance of these symbiotic features, their understanding remains limited. To overcome such limitations, we conduct genetic studies of nodulation in Aeschynomene evenia, supported by the development of a genome sequence for A. evenia and transcriptomic resources for 10 additional Aeschynomene spp. Comparative analysis of symbiotic genes substantiates singular mechanisms in the early and late nodulation steps. A forward genetic screen also shows that AeCRK, coding a receptor-like kinase, and the symbiotic signaling genes AePOLLUX, AeCCamK, AeCYCLOPS, AeNSP2, and AeNIN are required to trigger both root and stem nodulation. This work demonstrates the utility of the A. evenia model and provides a cornerstone to unravel mechanisms underlying the rhizobium–legume symbiosis.