Lipo-chitooligosaccharides Research Articles

The Influence of Rhizobial Nod Factors on the Synthesis of Flavonoids in Common Buckwheat (Fagopyrum esculentum Moench)

Flavonoids constitute a class of polyphenolic secondary metabolites synthesised mainly by plants and possessing anticancer, antioxidant, anti-inflammatory, and antiviral properties. Common buckwheat (F. esculentum Moench) is a dicotyledonous plant rich in different classes of flavonoids (e.g., rutin) and other phenolic compounds. Lipochitooligosaccharides (LCOs), i.e., rhizobial Nod factors and important signalling molecules for the initiation of symbiosis with legumes, are very effective mitogens that stimulate cell division in plant meristems and the production of secondary metabolites. They can also act in this way in non-legume plants. It has been shown that rhizobial Nod factors noticeably improve plant growth. Rhizobial Nod factors influence the production of flavonoids in common buckwheat grown in greenhouse conditions. The amount of rutin and isoorientin in leaves and flowers has been shown to increase in a statistically significant way after application of Nod factors to buckwheat seeds. The presence of rhizobial Nod factors has no influence on the flavonoid content in stems and roots.

A soybean cyst nematode suppresses microbial plant symbionts using a lipochitooligosaccharide-hydrolysing enzyme.

Cyst nematodes are the most damaging species of plant-parasitic nematodes. They antagonize the colonization of beneficial microbial symbionts that are important for nutrient acquisition of plants. The molecular mechanism of the antagonism, however, remains elusive. Here, through biochemical combined with structural analysis, we reveal that Heterodera glycines, the most notorious soybean cyst nematode, suppresses symbiosis by secreting an enzyme named HgCht2 to hydrolyse the key symbiotic signalling molecules, lipochitooligosaccharides (LCOs). We solved the three-dimensional structures of apo HgCht2, as well as its chitooligosaccharide-bound and LCO-bound forms. These structures elucidated the substrate binding and hydrolysing mechanism of the enzyme. We designed an HgCht2 inhibitor, 1516b, which successfully suppresses the antagonism of cyst nematodes towards nitrogen-fixing rhizobia and phosphorus-absorbing arbuscular mycorrhizal symbioses. As HgCht2 is phylogenetically conserved across all cyst nematodes, our study revealed a molecular mechanism by which parasitic cyst nematodes antagonize the establishment of microbial symbiosis and provided a small-molecule solution.

A gap in the recognition of two mycorrhizal factors: new insights into two LysM-type mycorrhizal receptors.

Arbuscular mycorrhizal (AM) fungi are crucial components of the plant microbiota and can form symbioses with 72% of land plants. Researchers have long known that AM symbioses have dramatic effects on plant performance and also provide multiple ecological services in terrestrial environments. The successful establishment of AM symbioses relies on the host plant recognition of the diffusible mycorrhizal (Myc) factors, lipo-chitooligosaccharides (LCOs) and chitooligosaccharides (COs). Among them, the short-chain COs such as CO4/5 secreted by AM fungi are the major Myc factors in COs. In this review, we summarize current advances, develop the concept of mycorrhizal biceptor complex (double receptor complexes for Myc-LCOs and CO4/5 in the same plant), and provide a perspective on the future development of mycorrhizal receptors. First, we focus on the distinct perception of two Myc factors by different host plant species, highlighting the essential role of Lysin-Motif (LysM)-type mycorrhizal receptors in perceiving them. Second, we propose the underlying molecular mechanisms by which LysM-type mycorrhizal receptors in various plants recognize both the Myc-LCOs and -COs. Finally, we explore future prospects for studies on the biceptor complex (Myc-LCO and -CO receptors) in dicots to facilitate the utilization of them in cereal crops (particularly in modern cultivated rice). In conclusion, our understanding of the precise perception processes during host plant interacting with AM fungi, where LysM-type mycorrhizal receptors act as recruiters, provides the tools to design biotechnological applications addressing agricultural challenges.

Studies on Influence of Lipo-chito Oligosaccharides, Nano and Water-Soluble Fertilizer on Growth and Growth Parameters of Soybean [Glycine max (L.) Merrill

A field experiment was conducted at University of Agricultural Sciences, GKVK, Bengaluru during kharif 2022 to studies on influence of lipo-chito oligosaccharides, nano and water-soluble fertilizer on growth and growth parameters of soybean [Glycine max (L.) Merrill]. The experiment was laid out in randomized complete block design with ten treatments and replicated thrice. The results revealed that application of 75% recommended dose of NP + 100 % K along with soil application of lipo-chito oligosaccharide (LCO) fortified bio-fertilizer @ 10 kg ha-1 + foliar application of 19:19:19 @ 2 % spray at flowering and pod filling stages recorded significantly higher leaf area, leaf area index at 60 DAS (1772 cm2 plant-1 and 4.91 respectively), leaf area duration (57.59 and 81.07 cm2 day-1 at 30-60 DAS and 60 DAS-harvest, respectively), no. of nodules plant at 35 DAS (31.12), total dry matter accumulation (12.74 and 20.05 g plant-1 at 60 DAS and harvest, respectively), absolute growth rate (0.24 and 0.30 g day-1 at 30-60 DAS and 60 DAS-harvest, respectively), crop growth rate (8.15 and 9.21 g cm-2 day-1 at 30-60 DAS and 60 DAS-harvest, respectively) and relative growth rate (0.031 and 0.040 g g day-1 at 30-60 DAS and 60 DAS­­-at harvest, respectively).

Evolution of lipochitooligosaccharide binding to a LysM-RLK for nodulation in Medicago truncatula.

Lysin motif receptor like kinases (LysM-RLKs) are involved in the perception of chitooligosaccharides (COs) and related lipochitooligosaccharides (LCOs) in plants. Expansion and divergence of the gene family during evolution have led to various roles in symbiosis and defence. By studying proteins of the LYR-IA subclass of LysM-RLKs of the Poaceae, we show here that they are high affinity LCO binding proteins with a lower affinity for COs, consistent with a role in LCO perception to establish arbuscular mycorrhiza (AM). In Papilionoid legumes whole genome duplication has resulted in two LYR-IA paralogs, MtLYR1 and MtNFP in Medicago truncatula, with MtNFP playing an essential role in the root nodule symbiosis with nitrogen-fixing rhizobia. We show that MtLYR1 has retained the ancestral LCO binding characteristic and is dispensable for AM. Domain swapping between the three Lysin motifs (LysMs) of MtNFP and MtLYR1 and mutagenesis in MtLYR1 suggest that the MtLYR1 LCO binding site is on the second LysM, and that divergence in MtNFP led to better nodulation, but surprisingly with decreased LCO binding. These results suggest that divergence of the LCO binding site has been important for the evolution of a role of MtNFP in nodulation with rhizobia.

LysM receptor-like kinases involved in immunity perceive lipo-chitooligosaccharides in mycotrophic plants.

Symbiotic microorganisms such as arbuscular mycorrhizal fungi (AMF) produce both conserved microbial molecules that activate plant defense and lipo-chitooligosaccharides (LCOs) that modulate plant defense. Beside a well-established role of LCOs in the activation of a signaling pathway required for AMF penetration in roots, LCO perception and defense modulation during arbuscular mycorrhiza is not well understood. Here we show that members of the LYRIIIA phylogenetic group from the multigenic Lysin Motif Receptor-Like Kinase family have a conserved role in dicotyledons as modulators of plant defense and regulate AMF colonization in the Solanaceae species Nicotiana benthamiana. Interestingly, these proteins have a high-affinity for LCOs in plant species able to form a symbiosis with AMF but have lost this property in species that have lost this ability. Our data support the hypothesis that LYRIIIA proteins modulate plant defense upon LCO perception to facilitate AMF colonization in mycotrophic plant species and that only their role in plant defense, but not their ability to be regulated by LCOs, has been conserved in non-mycotrophic plants.

Long-lasting impact of chito-oligosaccharide application on strigolactone biosynthesis and fungal accommodation promotes arbuscular mycorrhiza in Medicago truncatula.

• The establishment of arbuscular mycorrhiza (AM) between plants and Glomeromycotina fungi is preceded by the exchange of chemical signals: fungal released Myc-factors, including chitoligosaccharides (CO) and lipo-chitooligosaccharides (LCO), activate plant symbiotic responses, while root exuded strigolactones stimulate hyphal branching and boost CO release. Furthermore, fungal signaling reinforcement through CO application was shown to promote AM development in Medicago truncatula, but the cellular and molecular bases of this effect remained unclear. • Here we focused on long-term M. truncatula responses to CO treatment, demonstrating its impact on the transcriptome of both mycorrhizal and non-mycorrhizal roots over several weeks and providing an insight into the mechanistic bases of the CO-dependent promotion of AM colonization. • CO treatment caused the long-lasting regulation of strigolactone biosynthesis and fungal accommodation related genes. This was mirrored by an increase in root didehydro-orobanchol content, and the promotion of accommodation responses to AM fungi in root epidermal cells. Lastly, an advanced down-regulation of AM symbiosis marker genes was observed at the latest time point in CO-treated plants, in line with an increased number of senescent arbuscules. • Overall, CO treatment triggered molecular, metabolic and cellular responses underpinning a protracted acceleration of AM development.

Exploring the role of plant lysin motif receptor-like kinases in regulating plant-microbe interactions in the bioenergy crop Populus

For plants, distinguishing between mutualistic and pathogenic microbes is a matter of survival. All microbes contain microbe-associated molecular patterns (MAMPs) that are perceived by plant pattern recognition receptors (PRRs). Lysin motif receptor-like kinases (LysM-RLKs) are PRRs attuned for binding and triggering a response to specific MAMPs, including chitin oligomers (COs) in fungi, lipo-chitooligosaccharides (LCOs), which are produced by mycorrhizal fungi and nitrogen-fixing rhizobial bacteria, and peptidoglycan in bacteria. The identification and characterization of LysM-RLKs in candidate bioenergy crops including Populus are limited compared to other model plant species, thus inhibiting our ability to both understand and engineer microbe-mediated gains in plant productivity. As such, we performed a sequence analysis of LysM-RLKs in the Populus genome and predicted their function based on phylogenetic analysis with known LysM-RLKs. Then, using predictive models, molecular dynamics simulations, and comparative structural analysis with previously characterized CO and LCO plant receptors, we identified probable ligand-binding sites in Populus LysM-RLKs. Using several machine learning models, we predicted remarkably consistent binding affinity rankings of Populus proteins to CO. In addition, we used a modified Random Walk with Restart network-topology based approach to identify a subset of Populus LysM-RLKs that are functionally related and propose a corresponding signal transduction cascade. Our findings provide the first look into the role of LysM-RLKs in Populus-microbe interactions and establish a crucial jumping-off point for future research efforts to understand specificity and redundancy in microbial perception mechanisms.

Lipo-Chitooligosaccharides Induce Specialized Fungal Metabolite Profiles That Modulate Bacterial Growth.

Lipo-chitooligosaccharides (LCOs) are historically known for their role as microbial-derived signaling molecules that shape plant symbiosis with beneficial rhizobia or mycorrhizal fungi. Recent studies showing that LCOs are widespread across the fungal kingdom have raised questions about the ecological function of these compounds in organisms that do not form symbiotic relationships with plants. To elucidate the ecological function of these compounds, we investigate the metabolomic response of the ubiquitous human pathogen Aspergillus fumigatus to LCOs. Our metabolomics data revealed that exogenous application of various types of LCOs to A. fumigatus resulted in significant shifts in the fungal metabolic profile, with marked changes in the production of specialized metabolites known to mediate ecological interactions. Using network analyses, we identify specific types of LCOs with the most significant effect on the abundance of known metabolites. Extracts of several LCO-induced metabolic profiles significantly impact the growth rates of diverse bacterial species. These findings suggest that LCOs may play an important role in the competitive dynamics of non-plant-symbiotic fungi and bacteria. This study identifies specific metabolomic profiles induced by these ubiquitously produced chemicals and creates a foundation for future studies into the potential roles of LCOs as modulators of interkingdom competition. IMPORTANCE The activation of silent biosynthetic gene clusters (BGC) for the identification and characterization of novel fungal secondary metabolites is a perpetual motion in natural product discoveries. Here, we demonstrated that one of the best-studied symbiosis signaling compounds, lipo-chitooligosaccharides (LCOs), play a role in activating some of these BGCs, resulting in the production of known, putative, and unknown metabolites with biological activities. This collection of metabolites induced by LCOs differentially modulate bacterial growth, while the LCO standards do not convey the same effect. These findings create a paradigm shift showing that LCOs have a more prominent role outside of host recognition of symbiotic microbes. Importantly, our work demonstrates that fungi use LCOs to produce a variety of metabolites with biological activity, which can be a potential source of bio-stimulants, pesticides, or pharmaceuticals.

Nutrient regulation of lipochitooligosaccharide recognition in plants via NSP1 and NSP2

Many plants associate with arbuscular mycorrhizal fungi for nutrient acquisition, while legumes also associate with nitrogen-fixing rhizobial bacteria. Both associations rely on symbiosis signaling and here we show that cereals can perceive lipochitooligosaccharides (LCOs) for activation of symbiosis signaling, surprisingly including Nod factors produced by nitrogen-fixing bacteria. However, legumes show stringent perception of specifically decorated LCOs, that is absent in cereals. LCO perception in plants is activated by nutrient starvation, through transcriptional regulation of Nodulation Signaling Pathway (NSP)1 and NSP2. These transcription factors induce expression of an LCO receptor and act through the control of strigolactone biosynthesis and the karrikin-like receptor DWARF14-LIKE. We conclude that LCO production and perception is coordinately regulated by nutrient starvation to promote engagement with mycorrhizal fungi. Our work has implications for the use of both mycorrhizal and rhizobial associations for sustainable productivity in cereals.

Evolution of LysM-RLK Gene Family in Wild and Cultivated Peanut Species

In legumes, a LysM-RLK perception of rhizobial lipo-chitooligosaccharides (LCOs) known as Nod factors (NFs), triggers a signaling pathway related to the onset of symbiosis development. On the other hand, activation of LysM-RLKs upon recognition of chitin-derived short-chitooligosaccharides initiates defense responses. In this work, we identified the members of the LysM-RLK family in cultivated (Arachis hypogaea L.) and wild (A. duranensis and A. ipaensis) peanut genomes, and reconstructed the evolutionary history of the family. Phylogenetic analyses allowed the building of a framework to reinterpret the functional data reported on peanut LysM-RLKs. In addition, the potential involvement of two identified proteins in NF perception and immunity was assessed by gene expression analyses. Results indicated that peanut LysM-RLK is a highly diverse family. Digital expression analyses indicated that some A. hypogaea LysM-RLK receptors were upregulated during the early and late stages of symbiosis. In addition, expression profiles of selected LysM-RLKs proteins suggest participation in the receptor network mediating NF and/or chitosan perception. The analyses of LysM-RLK in the non-model legume peanut can contribute to gaining insight into the molecular basis of legume–microbe interactions and to the understanding of the evolutionary history of this gene family within the Fabaceae.

An LCO-responsive homolog of NODULE INCEPTION positively regulates lateral root formation in Populus sp.

The transcription factor NODULE INCEPTION (NIN) has been studied extensively for its multiple roles in root nodule symbiosis within plants of the nitrogen-fixing clade (NFC) that associate with soil bacteria, such as rhizobia and Frankia. However, NIN homologs are present in plants outside the NFC, suggesting a role in other developmental processes. Here, we show that the biofuel crop Populus sp., which is not part of the NFC, contains eight copies of NIN with diversified protein sequence and expression patterns. Lipo-chitooligosaccharides (LCOs) are produced by rhizobia and a wide range of fungi, including mycorrhizal ones, and act as symbiotic signals that promote lateral root formation. RNAseq analysis of Populus sp. treated with purified LCO showed induction of the PtNIN2 subfamily. Moreover, the expression of PtNIN2b correlated with the formation of lateral roots and was suppressed by cytokinin treatment. Constitutive expression of PtNIN2b overcame the inhibition of lateral root development by cytokinin under high nitrate conditions. Lateral root induction in response to LCOs likely represents an ancestral function of NIN retained and repurposed in nodulating plants, as we demonstrate that the role of NIN in LCO-induced root branching is conserved in both Populus sp. and legumes. We further established a visual marker of LCO perception in Populus sp. roots, the putative sulfotransferase PtSS1 that can be used to study symbiotic interactions with the bacterial and fungal symbionts of Populus sp.

An integrated approach reveals how lipo-chitooligosaccharides interact with the lysin motif receptor-like kinase MtLYR3.

N‐acetylglucosamine containing compounds acting as pathogenic or symbiotic signals are perceived by plant‐specific Lysin Motif Receptor‐Like Kinases (LysM‐RLKs). The molecular mechanisms of this perception are not fully understood, notably those of lipo‐chitooligosaccharides (LCOs) produced during root endosymbioses with nitrogen‐fixing bacteria or arbuscular mycorrhizal fungi. In Medicago truncatula, we previously identified the LysM‐RLK LYR3 (MtLYR3) as a specific LCO‐binding protein. We also showed that the absence of LCO binding to LYR3 of the non‐mycorrhizal Lupinus angustifolius, (LanLYR3), was related to LysM3, which differs from that of MtLYR3 by several amino acids and, particularly, by a critical tyrosine residue absent in LanLYR3. Here, we aimed to define the LCO binding site of MtLYR3 by using molecular modelling and simulation approaches, combined with site‐directed mutagenesis and LCO binding experiments. 3D models of MtLYR3 and LanLYR3 ectodomains were built, and homology modelling and molecular dynamics (MD) simulations were performed. Molecular docking and MD simulation on the LysM3 identified potential key residues for LCO binding. We highlighted by steered MD simulations that in addition to the critical tyrosine, two other residues were important for LCO binding in MtLYR3. Substitution of these residues in LanLYR3‐LysM3 by those of MtLYR3‐LysM3 allowed the recovery of high‐affinity LCO binding in experimental radioligand‐binding assays. An analysis of selective constraints revealed that the critical tyrosine has experienced positive selection pressure and is absent in some LYR3 proteins. These findings now pave the way to uncover the functional significance of this specific evolutionary pattern.

Lipo-Chitooligosaccharides (LCOs) as Elicitors of the Enzymatic Activities Related to ROS Scavenging to Alleviate Oxidative Stress Generated in Tomato Plants under Stress by UV-B Radiation.

Exposure to ultraviolet-B (UV-B) radiation can lead to oxidative damage in plants, increasing reactive oxygen species (ROS) production. To overcome ROS burst, plants have antioxidant mechanisms related to ROS scavenging which can be improved by elicitation with biological agents or derived molecules (elicitors), as they can trigger a physiological alert state called “priming”. This work describes the effects of lipo-chitooligosaccharides (LCOs) treatment applied to tomato plants under UV-B stress. The LCOs used in the study are produced by three species of the genus Ensifer (formerly Sinorhizobium) (SinCEU-1, SinCEU-2, and SinCEU-3) were assayed on tomato plants under UV-B stress. LCOs were able to significantly increase most of the enzymatic activities related to ROS scavenging while non-enzymatic antioxidants were not modified. This response was associated with a lower oxidative stress, according to malondialdehyde (MDA) levels and the higher antioxidant capacity of the plants. Furthermore, the photosynthetic efficiency of LCOs-treated plants indicated a better physiological state than the control plants. Therefore, although more studies and deepening of certain aspects are necessary, LCOs have shown great potential to protect plants from high UV-B radiation conditions.

Nod-Factor structure and functional redundancy of nod genes contribute the broad host range Bradyrhizobium sp. DOA9

Bradyrhizobium sp. DOA9 has a broad host range and a highly divergent nodulation (nod) gene on the megaplasmid (pDOA9). Duplicated copies of the general regulator nodD (nodD1 and nodD2) and nodA (nodA1 and nodA2) genes were found on pDOA9. The nodA gene is encoding an acyltransferase that controls the transfer of an acyl chain to the chitooligosaccharides (COs) structure and contribute to the determination of host specify. In general, compatible recognition between transcriptional regulators NodD and plant flavonoid inducer able to activate the function of other nodulation (nod) genes which responsible to produce lipo-chitooligosaccharides (LCOs), calls Nod-Factor, a major determinants of host range, infection, and nodulation. In this study, the function of the duplicated nodD (nodD1 and nodD2) and nodA (nodA1 and nodA2) genes were disrupted to determine their role on the symbiotic nodulation. The results showed that the mutation of nodD1 or nodD2 did not affected to the symbiotic nodulation, indicating the functional redundancy of the regulatory nodD1 and nodD2 genes in strain DOA9. Whereas nodA2 and nodB genes plays as a central role in the Nod-Factor biosynthesis and the symbiotic nodulation in all plant tests. Moreover, Nod-Factor produced from DOA9 under genistein induction shared a high similarity with the major Nod-Factor of Rhizobium sp. NGR234. Interestingly, mutation of nodA1 (ΔnodA1) did not affect to the nodulation but lead to induce the production of Nod-Factor belonged to the wild type, with the absence of Nod-Factor bearing the C18:1 acyl group. All these results suggest that the host range of the strain DOA9 might acquire from the functional redundancy of regulatory nodD genes and the Nod-Factor structure.

Expanding the Biological Role of Lipo-Chitooligosaccharides and Chitooligosaccharides in Laccaria bicolor Growth and Development.

The role of lipo-chitooligosaccharides (LCOs) as signaling molecules that mediate the establishment of symbiotic relationships between fungi and plants is being redefined. New evidence suggests that the production of these molecular signals may be more of a common trait in fungi than what was previously thought. LCOs affect different aspects of growth and development in fungi. For the ectomycorrhizal forming fungi, Laccaria bicolor, the production and effects of LCOs have always been studied with a symbiotic plant partner; however, there is still no scientific evidence describing the effects that these molecules have on this organism. Here, we explored the physiological, molecular, and metabolomic changes in L. bicolor when grown in the presence of exogenous sulfated and non-sulfated LCOs, as well as the chitooligomers, chitotetraose (CO4), and chitooctaose (CO8). Physiological data from 21 days post-induction showed reduced fungal growth in response to CO and LCO treatments compared to solvent controls. The underlying molecular changes were interrogated by proteomics, which revealed substantial alterations to biological processes related to growth and development. Moreover, metabolite data showed that LCOs and COs caused a downregulation of organic acids, sugars, and fatty acids. At the same time, exposure to LCOs resulted in the overproduction of lactic acid in L. bicolor. Altogether, these results suggest that these signals might be fungistatic compounds and contribute to current research efforts investigating the emerging impacts of these molecules on fungal growth and development.

Phenolic Acids Induce Nod Factor Production in Lotus japonicus-Mesorhizobium Symbiosis.

In legume–rhizobia symbiosis, partner recognition and the initiation of symbiosis processes require the mutual exchange of chemical signals. Chemicals, generally (iso)flavonoids, in the root exudates of the host plant induce the expression of nod genes in rhizobia, and, thus, are called nod gene inducers. The expression of nod genes leads to the production of lipochitooligosaccharides (LCOs) called Nod factors. Natural nod gene inducer(s) in Lotus japonicus–Mesorhizobium symbiosis remain unknown. Therefore, we developed an LCO detection method based on ultra-high-performance liquid chromatography–tandem-quadrupole mass spectrometry (UPLC-TQMS) to identify these inducers and used it herein to screen 40 phenolic compounds and aldonic acids for their ability to induce LCOs in Mesorhizobium japonicum MAFF303099. We identified five phenolic acids with LCO-inducing activities, including p-coumaric, caffeic, and ferulic acids. The induced LCOs caused root hair deformation, and nodule numbers in L. japonicus inoculated with M. japonicum were increased by these phenolic acids. The three phenolic acids listed above induced the expression of the nodA, nodB, and ttsI genes in a strain harboring a multicopy plasmid encoding NodD1, but not that encoding NodD2. The presence of p-coumaric and ferulic acids in the root exudates of L. japonicus was confirmed by UPLC-TQMS, and the induction of ttsI::lacZ in the strain harboring the nodD1 plasmid was detected in the rhizosphere of L. japonicus. Based on these results, we propose that phenolic acids are a novel type of nod gene inducer in L. japonicus–Mesorhizobium symbiosis.

Kinetic proofreading of lipochitooligosaccharides determines signal activation of symbiotic plant receptors

Plants and animals use cell surface receptors to sense and interpret environmental signals. In legume symbiosis with nitrogen-fixing bacteria, the specific recognition of bacterial lipochitooligosaccharide (LCO) signals by single-pass transmembrane receptor kinases determines compatibility. Here, we determine the structural basis for LCO perception from the crystal structures of two lysin motif receptor ectodomains and identify a hydrophobic patch in the binding site essential for LCO recognition and symbiotic function. We show that the receptor monitors the composition of the amphiphilic LCO molecules and uses kinetic proofreading to control receptor activation and signaling specificity. We demonstrate engineering of the LCO binding site to fine-tune ligand selectivity and correct binding kinetics required for activation of symbiotic signaling in plants. Finally, the hydrophobic patch is found to be a conserved structural signature in this class of LCO receptors across legumes that can be used for in silico predictions. Our results provide insights into the mechanism of cell-surface receptor activation by kinetic proofreading of ligands and highlight the potential in receptor engineering to capture benefits in plant-microbe interactions.

Deciphering the Chitin Code in Plant Symbiosis, Defense, and Microbial Networks.

Chitin is a structural polymer in many eukaryotes. Many organisms can degrade chitin to defend against chitinous pathogens or use chitin oligomers as food. Beneficial microorganisms like nitrogen-fixing symbiotic rhizobia and mycorrhizal fungi produce chitin-based signal molecules called lipo-chitooligosaccharides (LCOs) and short chitin oligomers to initiate a symbiotic relationship with their compatible hosts and exchange nutrients. A recent study revealed that a broad range of fungi produce LCOs and chitooligosaccharides (COs), suggesting that these signaling molecules are not limited to beneficial microbes. The fungal LCOs also affect fungal growth and development, indicating that the roles of LCOs beyond symbiosis and LCO production may predate mycorrhizal symbiosis. This review describes the diverse structures of chitin; their perception by eukaryotes and prokaryotes; and their roles in symbiotic interactions, defense, and microbe-microbe interactions. We also discuss potential strategies of fungi to synthesize LCOs and their roles in fungi with different lifestyles.

Spatiotemporal cytokinin response imaging and ISOPENTENYLTRANSFERASE 3 function in Medicago nodule development.

Most legumes can establish a symbiotic association with soil rhizobia that trigger the development of root nodules. These nodules host the rhizobia and allow them to fix nitrogen efficiently. The perception of bacterial lipo-chitooligosaccharides (LCOs) in the epidermis initiates a signaling cascade that allows rhizobial intracellular infection in the root and de-differentiation and activation of cell division that gives rise to the nodule. Thus, nodule organogenesis and rhizobial infection need to be coupled in space and time for successful nodulation. The plant hormone cytokinin (CK) contributes to the coordination of this process, acting as an essential positive regulator of nodule organogenesis. However, the temporal regulation of tissue-specific CK signaling and biosynthesis in response to LCOs or Sinorhizobium meliloti inoculation in Medicago truncatula remains poorly understood. In this study, using a fluorescence-based CK sensor (pTCSn::nls:tGFP), we performed a high-resolution tissue-specific temporal characterization of the sequential activation of CK response during root infection and nodule development in M. truncatula after inoculation with S. meliloti. Loss-of-function mutants of the CK-biosynthetic gene ISOPENTENYLTRANSFERASE 3 (IPT3) showed impairment of nodulation, suggesting that IPT3 is required for nodule development in M. truncatula. Simultaneous live imaging of pIPT3::nls:tdTOMATO and the CK sensor showed that IPT3 induction in the pericycle at the base of nodule primordium contributes to CK biosynthesis, which in turn promotes expression of positive regulators of nodule organogenesis in M. truncatula.