Insight into the control of nodule immunity and senescence during Medicago truncatula symbiosis.

Medicago (Medicago truncatula) establishes a symbiosis with the rhizobia Sinorhizobium sp, resulting in the formation of nodules where the bacteria fix atmospheric nitrogen. The loss of immunity repression or early senescence activation compromises symbiont survival and leads to the formation of nonfunctional nodules (fix-). Despite many studies exploring an overlap between immunity and senescence responses outside the nodule context, the relationship between these processes in the nodule remains poorly understood. To investigate this phenomenon, we selected and characterized three Medicago mutants developing fix- nodules and showing senescence responses. Analysis of specific defense (PATHOGENESIS-RELATED PROTEIN) or senescence (CYSTEINE PROTEASE) marker expression demonstrated that senescence and immunity seem to be antagonistic in fix- nodules. The growth of senescence mutants on non-sterile (sand/perlite) substrate instead of sterile in vitro conditions decreased nodule senescence and enhanced defense, indicating that environment can affect the immunity/senescence balance. The application of wounding stress on wild-type (WT) fix+ nodules led to the death of intracellular rhizobia and associated with co-stimulation of defense and senescence markers, indicating that in fix+ nodules the relationship between the two processes switches from opposite to synergistic to control symbiont survival during response to the stress. Our data show that the immune response in stressed WT nodules is linked to the repression of DEFECTIVE IN NITROGEN FIXATION 2 (DNF2), Symbiotic CYSTEINE-RICH RECEPTOR-LIKE KINASE (SymCRK), and REGULATOR OF SYMBIOSOME DIFFERENTIATION (RSD), key genes involved in symbiotic immunity suppression. This study provides insight to understand the links between senescence and immunity in Medicago nodules.

Brachypodium distachyon UNICULME4 and LAXATUM-A are redundantly required for development.

In cultivated grasses, tillering, leaf, and inflorescence architecture, as well as abscission ability, are major agronomical traits. In barley (Hordeum vulgare), maize (Zea mays), rice (Oryza sativa), and brachypodium (Brachypodium distachyon), NOOT-BOP-COCH-LIKE (NBCL) genes are essential regulators of vegetative and reproductive development. Grass species usually possess 2-4 NBCL copies and until now a single study in O. sativa showed that the disruption of all NBCL genes strongly altered O. sativa leaf development. To improve our understanding of the role of NBCL genes in grasses, we extended the study of the two NBCL paralogs BdUNICULME4 (CUL4) and BdLAXATUM-A (LAXA) in the nondomesticated grass B. distachyon. For this, we applied reversed genetics and generated original B. distachyon single and double nbcl mutants by clustered regularly interspaced short palindromic repeats - CRISPR associated protein 9 (CRISPR-Cas9) approaches and genetic crossing between nbcl targeting induced local lesions in genomes (TILLING) mutants. Through the study of original single laxa CRISPR-Cas9 null alleles, we validated functions previously proposed for LAXA in tillering, leaf patterning, inflorescence, and flower development and also unveiled roles for these genes in seed yield. Furthermore, the characterization of cul4laxa double mutants revealed essential functions for nbcl genes in B. distachyon development, especially in the regulation of tillering, stem cell elongation and secondary cell wall composition as well as for the transition toward the reproductive phase. Our results also highlight recurrent antagonist interactions between NBCLs occurring in multiple aspects of B. distachyon development.

The transcriptional co-regulators NBCL1 and NBCL2 redundantly coordinate aerial organ development and root nodule identity in legumes.

Medicago truncatula NODULE ROOT1 (MtNOOT1) and Pisum sativum COCHLEATA1 (PsCOCH1) are orthologous genes belonging to the NOOT-BOP-COCH-LIKE (NBCL) gene family which encodes key transcriptional co-regulators of plant development. In Mtnoot1 and Pscoch1 mutants, the development of stipules, flowers, and symbiotic nodules is altered. MtNOOT2 and PsCOCH2 represent the single paralogues of MtNOOT1 and PsCOCH1, respectively. In M. truncatula, MtNOOT1 and MtNOOT2 are both required for the establishment and maintenance of symbiotic nodule identity. In legumes, the role of NBCL2 in above-ground development is not known. To better understand the roles of NBCL genes in legumes, we used M. truncatula and P. sativum nbcl mutants, isolated a knockout mutant for the PsCOCH2 locus and generated Pscoch1coch2 double mutants in P. sativum. Our work shows that single Mtnoot2 and Pscoch2 mutants develop wild-type stipules, flowers, and symbiotic nodules. However, the number of flowers was increased and the pods and seeds were smaller compared to the wild type. Furthermore, in comparison to the corresponding nbcl1 single mutants, both the M. truncatula and P. sativum nbcl double mutants show a drastic alteration in stipule, inflorescence, flower, and nodule development. Remarkably, in both M. truncatula and P. sativum nbcl double mutants, stipules are transformed into a range of aberrant leaf-like structures.

Arabidopsis thaliana SHOOT MERISTEMLESS Substitutes for Medicago truncatula SINGLE LEAFLET1 to Form Complex Leaves and Petals.

LEAFY plant-specific transcription factors, which are key regulators of flower meristem identity and floral patterning, also contribute to meristem activity. Notably, in some legumes, LFY orthologs such as Medicago truncatula SINGLE LEAFLET (SGL1) are essential in maintaining an undifferentiated and proliferating fate required for leaflet formation. This function contrasts with most other species, in which leaf dissection depends on the reactivation of KNOTTED-like class I homeobox genes (KNOXI). KNOXI and SGL1 genes appear to induce leaf complexity through conserved downstream genes such as the meristematic and boundary CUP-SHAPED COTYLEDON genes. Here, we compare in M. truncatula the function of SGL1 with that of the Arabidopsis thaliana KNOXI gene, SHOOT MERISTEMLESS (AtSTM). Our data show that AtSTM can substitute for SGL1 to form complex leaves when ectopically expressed in M. truncatula. The shared function between AtSTM and SGL1 extended to the major contribution of SGL1 during floral development as ectopic AtSTM expression could promote floral organ identity gene expression in sgl1 flowers and restore sepal shape and petal formation. Together, our work reveals a function for AtSTM in floral organ identity and a higher level of interchangeability between meristematic and floral identity functions for the AtSTM and SGL1 transcription factors than previously thought.

Effector-Dependent and -Independent Molecular Mechanisms of Soybean-Microbe Interaction.

Soybean is a pivotal staple crop worldwide, supplying the main food and feed plant proteins in some countries. In addition to interacting with mutualistic microbes, soybean also needs to protect itself against pathogens. However, to grow inside plant tissues, plant defense mechanisms ranging from passive barriers to induced defense reactions have to be overcome. Pathogenic but also symbiotic micro-organisms effectors can be delivered into the host cell by secretion systems and can interfere with the immunity system and disrupt cellular processes. This review summarizes the latest advances in our understanding of the interaction between secreted effectors and soybean feedback mechanism and uncovers the conserved and special signaling pathway induced by pathogenic soybean cyst nematode, Pseudomonas, Xanthomonas as well as by symbiotic rhizobium.

Roles of BdUNICULME4 and BdLAXATUM-A in the non-domesticated grass Brachypodium distachyon.

In cultivated grasses, tillering, spike architecture and seed shattering represent major agronomical traits. In barley, maize and rice, the NOOT-BOP-COCH-LIKE (NBCL) genes play important roles in development, especially in ligule development, tillering and flower identity. However, compared with dicots, the role of grass NBCL genes is underinvestigated. To better understand the role of grass NBCLs and to overcome any effects of domestication that might conceal their original functions, we studied TILLING nbcl mutants in the non-domesticated grass Brachypodium distachyon. In B. distachyon, the NBCL genes BdUNICULME4 (CUL4) and BdLAXATUM-A (LAXA) are orthologous, respectively, to the barley HvUniculme4 and HvLaxatum-a, to the maize Zmtassels replace upper ears1 and Zmtassels replace upper ears2 and to the rice OsBLADE-ON-PETIOLE1 and OsBLADE-ON-PETIOLE2/3. In B. distachyon, our reverse genetics study shows that CUL4 is not essential for the establishment of the blade-sheath boundary but is necessary for the development of the ligule and auricles. We report that CUL4 also exerts a positive role in tillering and a negative role in spikelet meristem activity. On the other hand, we demonstrate that LAXA plays a negative role in tillering, positively participates in spikelet development and contributes to the control of floral organ number and identity. In this work, we functionally characterized two new NBCL genes in a context of non-domesticated grass and highlighted original roles for grass NBCL genes that are related to important agronomical traits.

Sinorhizobium meliloti succinylated high-molecular-weight succinoglycan and the Medicago truncatula LysM receptor-like kinase MtLYK10 participate independently in symbiotic infection.

The formation of nitrogen-fixing nodules on legume hosts is a finely tuned process involving many components of both symbiotic partners. Production of the exopolysaccharide succinoglycan by the nitrogen-fixing bacterium Sinorhizobium meliloti 1021 is needed for an effective symbiosis with Medicago spp., and the succinyl modification to this polysaccharide is critical. However, it is not known when succinoglycan intervenes in the symbiotic process, and it is not known whether the plant lysin-motif receptor-like kinase MtLYK10 intervenes in recognition of succinoglycan, as might be inferred from work on the Lotus japonicus MtLYK10 ortholog, LjEPR3. We studied the symbiotic infection phenotypes of S. meliloti mutants deficient in succinoglycan production or producing modified succinoglycan, in wild-type Medicago truncatula plants and in Mtlyk10 mutant plants. On wild-type plants, S. meliloti strains producing no succinoglycan or only unsuccinylated succinoglycan still induced nodule primordia and epidermal infections, but further progression of the symbiotic process was blocked. These S. meliloti mutants induced a more severe infection phenotype on Mtlyk10 mutant plants. Nodulation by succinoglycan-defective strains was achieved by in trans rescue with a Nod factor-deficient S. meliloti mutant. While the Nod factor-deficient strain was always more abundant inside nodules, the succinoglycan-deficient strain was more efficient than the strain producing only unsuccinylated succinoglycan. Together, these data show that succinylated succinoglycan is essential for infection thread formation in M. truncatula, and that MtLYK10 plays an important, but different role in this symbiotic process. These data also suggest that succinoglycan is more important than Nod factors for bacterial survival inside nodules.

Lotus japonicus NOOT-BOP-COCH-LIKE1 is essential for nodule, nectary, leaf and flower development.

The NOOT-BOP-COCH-LIKE (NBCL) genes are orthologs of Arabidopsis thaliana BLADE-ON-PETIOLE1/2. The NBCLs are developmental regulators essential for plant shaping, mainly through the regulation of organ boundaries, the promotion of lateral organ differentiation and the acquisition of organ identity. In addition to their roles in leaf, stipule and flower development, NBCLs are required for maintaining the identity of indeterminate nitrogen-fixing nodules with persistent meristems in legumes. In legumes forming determinate nodules, without persistent meristem, the roles of NBCL genes are not known. We thus investigated the role of Lotus japonicus NOOT-BOP-COCH-LIKE1 (LjNBCL1) in determinate nodule identity and studied its functions in aerial organ development using LORE1 insertional mutants and RNA interference-mediated silencing approaches. In Lotus, LjNBCL1 is involved in leaf patterning and participates in the regulation of axillary outgrowth. Wild-type Lotus leaves are composed of five leaflets and possess a pair of nectaries at the leaf axil. Legumes such as pea and Medicago have a pair of stipules, rather than nectaries, at the base of their leaves. In Ljnbcl1, nectary development is abolished, demonstrating that nectaries and stipules share a common evolutionary origin. In addition, ectopic roots arising from nodule vascular meristems and reorganization of the nodule vascular bundle vessels were observed on Ljnbcl1 nodules. This demonstrates that NBCL functions are conserved in both indeterminate and determinate nodules through the maintenance of nodule vascular bundle identity. In contrast to its role in floral patterning described in other plants, LjNBCL1 appears essential for the development of both secondary inflorescence meristem and floral meristem.

Legume Nodules: Massive Infection in the Absence of Defense Induction.

Plants of the legume family host massive intracellular bacterial populations in the tissues of specialized organs, the nodules. In these organs, the bacteria, named rhizobia, can fix atmospheric nitrogen and transfer it to the plant. This special metabolic skill provides to the legumes an advantage when they grow on nitrogen-scarce substrates. While packed with rhizobia, the nodule cells remain alive, metabolically active, and do not develop defense reactions. Here, we review our knowledge on the control of plant immunity during the rhizobia-legume symbiosis. We present the results of an evolutionary process that selected both divergence of microbial-associated molecular motifs and active suppressors of immunity on the rhizobial side and, on the legume side, active mechanisms that contribute to suppression of immunity.

MtNODULE ROOT1 and MtNODULE ROOT2 Are Essential for Indeterminate Nodule Identity.

Symbiotic interactions between legume plants and rhizobia result in the formation of nitrogen-fixing nodules, but the molecular actors and the mechanisms allowing for the maintenance of nodule identity are poorly understood. Medicago truncatula NODULE ROOT1 (MtNOOT1), Pisum sativum COCHLEATA1 (PsCOCH1), and Lotus japonicus NOOT-BOP-COCH-LIKE1 (LjNBCL1) are orthologs of Arabidopsis (Arabidopsis thaliana) AtBLADE-ON-PETIOLE1/2 and are members of the NBCL gene family, which has conserved roles in plant development and is essential for indeterminate and determinate nodule identity in legumes. The loss of function of MtNOOT1, PsCOCH1, and LjNBCL1 triggers a partial loss of nodule identity characterized by the development of ectopic roots arising from nodule vascular meristems. Here, we report the identification and characterization of a second gene involved in regulating indeterminate nodule identity in M. truncatula, MtNOOT2MtNOOT2 is the paralog of MtNOOT1 and belongs to a second legume-specific NBCL subclade, the NBCL2 clade. MtNOOT2 expression was induced during early nodule formation, and it was expressed primarily in the nodule central meristem. Mtnoot2 mutants did not present any particular symbiotic phenotype; however, the loss of function of both MtNOOT1 and MtNOOT2 resulted in the complete loss of nodule identity and was accompanied by drastic changes in the expression of symbiotic, defense, and root apical meristem marker genes. Mtnoot1 noot2 double mutants developed only nonfixing root-like structures that were no longer able to host symbiotic rhizobia. This study provides original insights into the molecular basis underlying nodule identity in legumes forming indeterminate nodules.

Isoflavone production in hairy root cultures and plantlets of Trifolium pratense.

OBJECTIVES: The aim of this study was to develop a Trifolium pratense hairy root (HR) production protocol and select HR lines with high isoflavone yield following elicitor treatments. RESULTS: We obtained 13 independent HR lines, producing approximately three times more isoflavonoids than seedlings (3.3 mg/g dry weight) and in which 27 isoflavonoids were detected. Each HR line had its own isoflavonoid profile. These lines produced as major components daidzein, genistein, formononetin and biochanin A. Sucrose, salicylic acid (SA), yeast extract (YE) and flagellin 22 (flg22) were tested as elicitors. Using SA 140 mg/L, allowed the maximum isoflavonoid production in plantlets (11.9 mg/g dry weight) but reduced root growth, possibly as a result of its toxicity. The highest isoflavone production in HR (27.9 mg/g dry weight) was obtained using sucrose 60 g/L, for 3.5 days. CONCLUSION: This work reports the high production of various isoflavonoids with T. pratense elicited HR cultures.

Control of the ethylene signaling pathway prevents plant defenses during intracellular accommodation of the rhizobia.

Massive intracellular populations of symbiotic bacteria, referred to as rhizobia, are housed in legume root nodules. Little is known about the mechanisms preventing the development of defense in these organs although genes such as SymCRK and DNF2 of the model legume Medicago truncatula are required for this control after rhizobial internalization in host nodule cells. Here we investigated the molecular basis of the symbiotic control of immunity. Proteomic analysis was performed to compare functional (wild-type) and defending nodules (symCRK). Based on the results, the control of plant immunity during the functional step of the symbiosis was further investigated by biochemical and pharmacological approaches as well as by transcript and histology analysis. Ethylene was identified as a potential signal inducing plant defenses in symCRK nodules. Involvement of this phytohormone in symCRK and dnf2-developed defenses and in the death of intracellular rhizobia was confirmed. This negative effect of ethylene depended on the M. truncatula sickle gene and was also observed in the legume Lotus japonicus. Together, these data indicate that prevention of ethylene-triggered defenses is crucial for the persistence of endosymbiosis and that the DNF2 and SymCRK genes are required for this process.

A H+-ATPase That Energizes Nutrient Uptake during Mycorrhizal Symbioses in Rice and Medicago truncatula.

Most plant species form symbioses with arbuscular mycorrhizal (AM) fungi, which facilitate the uptake of mineral nutrients such as phosphate from the soil. Several transporters, particularly proton-coupled phosphate transporters, have been identified on both the plant and fungal membranes and contribute to delivering phosphate from fungi to plants. The mechanism of nutrient exchange has been studied in plants during mycorrhizal colonization, but the source of the electrochemical proton gradient that drives nutrient exchange is not known. Here, we show that plasma membrane H+-ATPases that are specifically induced in arbuscule-containing cells are required for enhanced proton pumping activity in membrane vesicles from AM-colonized roots of rice (Oryza sativa) and Medicago truncatula. Mutation of the H+-ATPases reduced arbuscule size and impaired nutrient uptake by the host plant through the mycorrhizal symbiosis. Overexpression of the H+-ATPase Os-HA1 increased both phosphate uptake and the plasma membrane potential, suggesting that this H+-ATPase plays a key role in energizing the periarbuscular membrane, thereby facilitating nutrient exchange in arbusculated plant cells.

Local and systemic regulation of plant root system architecture and symbiotic nodulation by a receptor-like kinase.

In plants, root system architecture is determined by the activity of root apical meristems, which control the root growth rate, and by the formation of lateral roots. In legumes, an additional root lateral organ can develop: the symbiotic nitrogen-fixing nodule. We identified in Medicago truncatula ten allelic mutants showing a compact root architecture phenotype (cra2) independent of any major shoot phenotype, and that consisted of shorter roots, an increased number of lateral roots, and a reduced number of nodules. The CRA2 gene encodes a Leucine-Rich Repeat Receptor-Like Kinase (LRR-RLK) that primarily negatively regulates lateral root formation and positively regulates symbiotic nodulation. Grafting experiments revealed that CRA2 acts through different pathways to regulate these lateral organs originating from the roots, locally controlling the lateral root development and nodule formation systemically from the shoots. The CRA2 LRR-RLK therefore integrates short- and long-distance regulations to control root system architecture under non-symbiotic and symbiotic conditions.

LOOSE FLOWER, a WUSCHEL-like Homeobox gene, is required for lateral fusion of floral organs in Medicago truncatula.

The Medicago truncatula WOX gene, STENOFOLIA (STF), and its orthologs in Petunia, pea, and Nicotiana sylvestris are required for leaf blade outgrowth and floral organ development as demonstrated by severe phenotypes in single mutants. But the Arabidopsis wox1 mutant displays a narrow leaf phenotype only when combined with the prs/wox3 mutant. In maize and rice, WOX3 homologs are major regulators of leaf blade development. Here we investigated the role of WOX3 in M. truncatula development by isolating the lfl/wox3 loss-of-function mutant and performing genetic crosses with the stf mutant. Lack of WOX3 function in M. truncatula leads to a loose-flower (lfl) phenotype, where defects are observed in sepal and petal development, but leaf blades are apparently normal. The stf lfl double mutant analysis revealed that STF and LFL act mainly independently with minor redundant functions in flower development, but LFL has no obvious role in leaf blade outgrowth in M. truncatula on its own or in combination with STF. Interestingly, LFL acts as a transcriptional repressor by recruiting TOPLESS in the same manner as STF does, and can substitute for STF function in leaf blade and flower development if expressed under the STF promoter. STF also complements the lfl mutant phenotype in the flower if expressed under the LFL promoter. Our data suggest that the STF/WOX1 and LFL/WOX3 genes of M. truncatula employ a similar mechanism of action in organizing cell proliferation for lateral outgrowth but may have evolved different cis elements to acquire distinct functions.

The legume NOOT-BOP-COCH-LIKE genes are conserved regulators of abscission, a major agronomical trait in cultivated crops.

Plants are able to lose organs selectively through a process called abscission. This process relies on the differentiation of specialized territories at the junction between organs and the plant body that are called abscission zones (AZ). Several genes control the formation or functioning of these AZ. We have characterized BLADE-ON-PETIOLE (BOP) orthologues from several legume plants and studied their roles in the abscission process using a mutant approach. Here, we show that the Medicago truncatula NODULE ROOT (NOOT), the Pisum sativum COCHLEATA (COCH) and their orthologue in Lotus japonicus are strictly necessary for the abscission of not only petals, but also leaflets, leaves and fruits. We also showed that the expression pattern of the M. truncatula pNOOT::GUS fusion is associated with functional and vestigial AZs when expressed in Arabidopsis. In addition, we show that the stip mutant from Lupinus angustifolius, defective in stipule formation and leaf abscission, is mutated in a BOP orthologue. In conclusion, this study shows that this clade of proteins plays an important conserved role in promoting abscission of all aerial organs studied so far.

Multiple steps control immunity during the intracellular accommodation of rhizobia.

Medicago truncatula belongs to the legume family and forms symbiotic associations with nitrogen fixing bacteria, the rhizobia. During these interactions, the plants develop root nodules in which bacteria invade the plant cells and fix nitrogen for the benefit of the plant. Despite massive infection, legume nodules do not develop visible defence reactions, suggesting a special immune status of these organs. Some factors influencing rhizobium maintenance within the plant cells have been previously identified, such as the M. truncatula NCR peptides whose toxic effects are reduced by the bacterial protein BacA. In addition, DNF2, SymCRK, and RSD are M. truncatula genes required to avoid rhizobial death within the symbiotic cells. DNF2 and SymCRK are essential to prevent defence-like reactions in nodules after bacteria internalization into the symbiotic cells. Herein, we used a combination of genetics, histology and molecular biology approaches to investigate the relationship between the factors preventing bacterial death in the nodule cells. We show that the RSD gene is also required to repress plant defences in nodules. Upon inoculation with the bacA mutant, defence responses are observed only in the dnf2 mutant and not in the symCRK and rsd mutants. In addition, our data suggest that lack of nitrogen fixation by the bacterial partner triggers bacterial death in nodule cells after bacteroid differentiation. Together our data indicate that, after internalization, at least four independent mechanisms prevent bacterial death in the plant cell. These mechanisms involve successively: DNF2, BacA, SymCRK/RSD and bacterial ability to fix nitrogen.

Strigolactones contribute to shoot elongation and to the formation of leaf margin serrations in Medicago truncatula R108.

Strigolactones were recently identified as a new class of plant hormones involved in the control of shoot branching. The characterization of strigolactone mutants in several species has progressively revealed their contribution to several other aspects of development in roots and shoots. In this article, we characterize strigolactone-deficient and strigolactone-insensitive mutants of the model legume Medicago truncatula for aerial developmental traits. The most striking mutant phenotype observed was compact shoot architecture. In contrast with what was reported in other species, this could not be attributed to enhanced shoot branching, but was instead due to reduced shoot elongation. Another notable feature was the modified leaf shape in strigolactone mutants: serrations at the leaf margin were smaller in the mutants than in wild-type plants. This phenotype could be rescued in a dose-dependent manner by exogenous strigolactone treatments of strigolactone-deficient mutants, but not of strigolactone-insensitive mutants. Treatment with the auxin transport inhibitor N-1-naphthylphtalamic acid resulted in smooth leaf margins, opposite to the effect of strigolactone treatment. The contribution of strigolactones to the formation of leaf serrations in M. truncatula R108 line represents a novel function of these hormones, which has not been revealed by the analysis of strigolactone mutants in other species.

Loss of abaxial leaf epicuticular wax in Medicago truncatula irg1/palm1 mutants results in reduced spore differentiation of anthracnose and nonhost rust pathogens.

To identify genes that confer nonhost resistance to biotrophic fungal pathogens, we did a forward-genetics screen using Medicago truncatula Tnt1 retrotransposon insertion lines. From this screen, we identified an inhibitor of rust germ tube differentation1 (irg1) mutant that failed to promote preinfection structure differentiation of two rust pathogens, Phakopsora pachyrhizi and Puccinia emaculata, and one anthracnose pathogen, Colletotrichum trifolii, on the abaxial leaf surface. Cytological and chemical analyses revealed that the inhibition of rust preinfection structures in irg1 mutants is due to complete loss of the abaxial epicuticular wax crystals and reduced surface hydrophobicity. The composition of waxes on abaxial leaf surface of irg1 mutants had >90% reduction of C30 primary alcohols and a preferential increase of C29 and C31 alkanes compared with the wild type. IRG1 encodes a Cys(2)His(2) zinc finger transcription factor, PALM1, which also controls dissected leaf morphology in M. truncatula. Transcriptome analysis of irg1/palm1 mutants revealed downregulation of eceriferum4, an enzyme implicated in primary alcohol biosynthesis, and MYB96, a major transcription factor that regulates wax biosynthesis. Our results demonstrate that PALM1 plays a role in regulating epicuticular wax metabolism and transport and that epicuticular wax influences spore differentiation of host and nonhost fungal pathogens.

NODULE ROOT and COCHLEATA maintain nodule development and are legume orthologs of Arabidopsis BLADE-ON-PETIOLE genes.

During their symbiotic interaction with rhizobia, legume plants develop symbiosis-specific organs on their roots, called nodules, that house nitrogen-fixing bacteria. The molecular mechanisms governing the identity and maintenance of these organs are unknown. Using Medicago truncatula nodule root (noot) mutants and pea (Pisum sativum) cochleata (coch) mutants, which are characterized by the abnormal development of roots from the nodule, we identified the NOOT and COCH genes as being necessary for the robust maintenance of nodule identity throughout the nodule developmental program. NOOT and COCH are Arabidopsis thaliana BLADE-ON-PETIOLE orthologs, and we have shown that their functions in leaf and flower development are conserved in M. truncatula and pea. The identification of these two genes defines a clade in the BTB/POZ-ankyrin domain proteins that shares conserved functions in eudicot organ development and suggests that NOOT and COCH were recruited to repress root identity in the legume symbiotic organ.