G-type Lectin Research Articles

Tyrosine phosphorylation of the lectin receptor-like kinase LORE regulates plant immunity.

Plant pattern recognition receptors (PRRs) perceive pathogen-associated molecular patterns (PAMPs) to activate immune responses. Medium-chain 3-hydroxy fatty acids (mc-3-OH-FAs), which are widely present in Gram-negative bacteria, were recently shown to be novel PAMPs in Arabidopsis thaliana. The Arabidopsis PRR LIPOOLIGOSACCHARIDE-SPECIFIC REDUCED ELICITATION (LORE) is a G-type lectin receptor-like kinase that recognizes mc-3-OH-FAs and subsequently mounts an immune response; however, the mechanisms underlying LORE activation and downstream signaling are unexplored. Here, we report that one of the mc-3-OH-FAs, 3-OH-C10:0, induces phosphorylation of LORE at tyrosine residue 600 (Y600). Phosphorylated LORE subsequently trans-phosphorylates the receptor-like cytoplasmic kinase PBL34 and its close paralogs, PBL35 and PBL36, and therefore activates plant immunity. Phosphorylation of LORE Y600 is required for downstream phosphorylation of PBL34, PBL35, and PBL36. However, the Pseudomonas syringae effector HopAO1 targets LORE, dephosphorylating the tyrosine-phosphorylated Y600 and therefore suppressing the immune response. These observations uncover the mechanism by which LORE mediates signaling in response to 3-OH-C10:0 in Arabidopsis.

Proteomics and post-secretory content adjustment of Nicotiana tabacum nectar.

The tobacco nectar proteome mainly consists of pathogenesis-related proteins with two glycoproteins. Expression of nectarins was non-synchronous, and not nectary specific. After secretion, tobacco nectar changed from sucrose rich to hexose rich. Floral nectar proteins (nectarins) play important roles in inhibiting microbial growth in nectar, and probably also tailoring nectar chemistry before or after secretion; however, very few plant species have had their nectar proteomes thoroughly investigated. Nectarins from Nicotiana tabacum (NT) were separated using two-dimensional gel electrophoresis and then analysed using mass spectrometry. Seven nectarins were identified: acidic endochitinase, β-xylosidase, α-galactosidase, α-amylase, G-type lectin S-receptor-like serine/threonine-protein kinase, pathogenesis-related protein 5, and early nodulin-like protein 2. An eighth nectarin, a glycoprotein with unknown function, was identified following isolation from NT nectar using a Qproteome total glycoprotein kit, separation by SDS-PAGE, and identification by mass spectrometry. Expression of all identified nectarins, plus four invertase genes, was analysed by qRT PCR; none of these genes had nectary-specific expression, and none had synchronous expression. The total content of sucrose, hexoses, proteins, phenolics, and hydrogen peroxide were determined at different time intervals in secreted nectar, both within the nectar tube (in vivo) and following extraction from it during incubation at 30°C for up to 40h in plastic tubes (in vitro). After secretion, the ratio of hexose to sucrose substantially increased for in vivo nectar, but no sugar composition changes were detected in vitro. This implies that sucrose hydrolysis in vivo might be done by fixed apoplastic invertase. Both protein and hydrogen peroxide levels declined in vitro but not in vivo, implying that some factors other than nectarins act to maintain their levels in the flower, after secretion.

Identification and functional analysis of LecRLK genes in Taxodium 'Zhongshanshan'.

BackgroundLectin receptor-like protein kinases (LecRLKs) can transform external stimuli into intracellular signals and play important regulatory roles in plant development and response to environmental stressors. However, research on the LecRLK gene family of conifers has seldom been reported.MethodsPutative LecRLK genes were identified in the transcriptome of Taxodium ‘Zhongshanshan’. The classification, domain structures, subcellular localization prediction, and expression patterns of LecRLK genes, as well as co-expressed genes, were analyzed using bioinformatics methods. Fifteen representative genes were further selected for qRT-PCR analysis in six tissues and under five different environmental stressor conditions.ResultsIn total, 297 LecRLK genes were identified, including 155 G-type, 140 L-type, and 2 C-type. According to the classification, G-type and L-type LecRLK genes both can be organized into seven groups. The domain architecture of G-type proteins were more complex compared with that of L- and C-type proteins. Conservative motifs were found in G-type and L-type diverse lectin domains. Prediction and transient expression experiments to determine subcellular localization showed that LecRLKs were mainly concentrated in the cell membrane system, and some members were located at multiple sites at the same time. RNA-seq-based transcriptomics analysis suggested functional redundancy and divergence within each group. Unigenes co-expressed with LecRLKs in the transcriptome were found to be enriched in pathways related to signal transduction and environmental adaptation. Furthermore, qRT-PCR analysis of representative genes showed evidence of functional divergence between different groups.ConclusionsThis is the first study to conduct an identification and expression analysis of the LecRLK gene family in Taxodium. These results provide a basis for future studies on the evolution and function of this important gene family in Taxodium.

Mediation of plant-mycorrhizal interaction by a lectin receptor-like kinase.

The molecular mechanisms underlying mycorrhizal symbioses, the most ubiquitous and impactful mutualistic plant-microbial interaction in nature, are largely unknown. Through genetic mapping, resequencing and molecular validation, we demonstrate that a G-type lectin receptor-like kinase (lecRLK) mediates the symbiotic interaction between Populus and the ectomycorrhizal fungus Laccaria bicolor. This finding uncovers an important molecular step in the establishment of symbiotic plant-fungal associations and provides a molecular target for engineering beneficial mycorrhizal relationships.

MiR393 regulation of lectin receptor-like kinases associated with LPS perception in Arabidopsis thaliana

microRNAs regulate dynamic aspects of innate immunity in Arabidopsis thaliana in response to lipopolysaccharides. Lectin-domain receptor-like kinases function as surveillance proteins and miR393 targets transcripts of an L-type LecRK (LECRK-V.7, At3g59740). This study investigated miR393 regulation of LecRLKs associated with LPS perception. Following pre-treatment of wild type -, miR393 ab double mutant - and miR393 overexpressor plants with LPS, the expression of miR393 and two other LecRLK genes (G-type lectin S-receptor-like protein kinases, SD1-13 (At1g11330) and SD1-29 (At1g61380) were evaluated. Overexpression and repression of miR393 respectively suppressed and induced transcripts of the LecRLK genes. The results indicate that miR393 regulates the three LecRLKs following perception of bacterial LPS, in support of immunity and basal resistance.

Identification of Candidate Ergosterol-Responsive Proteins Associated with the Plasma Membrane of Arabidopsis thaliana.

The impact of fungal diseases on crop production negatively reflects on sustainable food production and overall economic health. Ergosterol is the major sterol component in fungal membranes and regarded as a general elicitor or microbe-associated molecular pattern (MAMP) molecule. Although plant responses to ergosterol have been reported, the perception mechanism is still unknown. Here, Arabidopsis thaliana protein fractions were used to identify those differentially regulated following ergosterol treatment; additionally, they were subjected to affinity-based chromatography enrichment strategies to capture and categorize ergosterol-interacting candidate proteins using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Mature plants were treated with 250 nM ergosterol over a 24 h period, and plasma membrane-associated fractions were isolated. In addition, ergosterol was immobilized on two different affinity-based systems to capture interacting proteins/complexes. This resulted in the identification of defense-related proteins such as chitin elicitor receptor kinase (CERK), non-race specific disease resistance/harpin-induced (NDR1/HIN1)-like protein, Ras-related proteins, aquaporins, remorin protein, leucine-rich repeat (LRR)- receptor like kinases (RLKs), G-type lectin S-receptor-like serine/threonine-protein kinase (GsSRK), and glycosylphosphatidylinositol (GPI)-anchored protein. Furthermore, the results elucidated unknown signaling responses to this MAMP, including endocytosis, and other similarities to those previously reported for bacterial flagellin, lipopolysaccharides, and fungal chitin.

On a Cold Night: Transcriptomics of Grapevine Flower Unveils Signal Transduction and Impacted Metabolism.

Low temperature is a critical environmental factor limiting plant productivity, especially in northern vineyards. To clarify the impact of this stress on grapevine flower, we used the Vitis array based on Roche-NimbleGen technology to investigate the gene expression of flowers submitted to a cold night. Our objectives were to identify modifications in the transcript levels after stress and during recovery. Consequently, our results confirmed some mechanisms known in grapes or other plants in response to cold stress, notably, (1) the pivotal role of calcium/calmodulin-mediated signaling; (2) the over-expression of sugar transporters and some genes involved in plant defense (especially in carbon metabolism), and (3) the down-regulation of genes encoding galactinol synthase (GOLS), pectate lyases, or polygalacturonases. We also identified some mechanisms not yet known to be involved in the response to cold stress, i.e., (1) the up-regulation of genes encoding G-type lectin S-receptor-like serine threonine-protein kinase, pathogen recognition receptor (PRR5), or heat-shock factors among others; (2) the down-regulation of Myeloblastosis (MYB)-related transcription factors and the Constans-like zinc finger family; and (3) the down-regulation of some genes encoding Pathogen-Related (PR)-proteins. Taken together, our results revealed interesting features and potentially valuable traits associated with stress responses in the grapevine flower. From a long-term perspective, our study provides useful starting points for future investigation.

Isolation and characterization of fusarium wilt resistance gene analogs in radish.

The resistance gene analog (RGA)-based marker strategy is an effective supplement for current marker reservoir of radish disease-resistance breeding. In this study, we identified RGAs based on the conserved nucleotide-binding site (NBS) and S-receptor-like kinase (SRLK) domains. A total of 68 NBS-RGAs and 46 SRLK-RGAs were isolated from two FW-resistant radish inbred lines, B2 and YR31, and one susceptible line, YR15. A BLASTx search revealed that the NBS-RGAs contained six conserved motifs (i.e., P loop, RNBS-A, Kinase-2, RNBS-B, RNBS-C, and GLPL) and the SRLK-RGAs, contained two conserved motifs (i.e., G-type lectin and PAN-AP). A phylogenetic analysis indicated that the NBS-RGAs could be separated into two classes (i.e., toll/interleukin receptor and coiled-coil types), with six subgroups, and the SRLK-RGAs were divided into three subgroups. Moreover, we designed RGA-specific markers from data-mining approach in radish databases. Based on marker analysis, 24 radish inbred lines were clustered into five main groups with a similarity index of 0.44 and showing genetic diversity with resistance variation in those radish inbred lines. The development of RGA-specific primers would be valuable for marker-assisted selection during the breeding of disease-resistant radish cultivars.

Transcriptomic analysis between self- and cross-pollinated pistils of tea plants (Camellia sinensis)

BackgroundSelf-incompatibility (SI) is a major barrier that obstructs the breeding process in most horticultural plants including tea plants (Camellia sinensis). The aim of this study was to elucidate the molecular mechanism of SI in tea plants through a high throughput transcriptome analysis.ResultsIn this study, the transcriptomes of self- and cross-pollinated pistils of two tea cultivars ‘Fudingdabai’ and ‘Yulv’ were compared to elucidate the SI mechanism of tea plants. In addition, the ion components and pollen tube growth in self- and cross-pollinated pistils were investigated. Our results revealed that both cultivars had similar pollen activities and cross-pollination could promote the pollen tube growth. In tea pistils, the highest ion content was potassium (K+), followed by calcium (Ca2+), magnesium (Mg2+) and phosphorus (P5+). Ca2+ content increased after self-pollination but decreased after cross-pollination, while K+ showed reverse trend with Ca2+. A total of 990 and 3 common differentially expressed genes (DEGs) were identified in un-pollinated vs. pollinated pistils and self- vs. cross-pollinated groups after 48 h, respectively. Function annotation indicated that three genes encoding UDP-glycosyltransferase 74B1 (UGT74B1), Mitochondrial calcium uniporter protein 2 (MCU2) and G-type lectin S-receptor-like serine/threonine-protein kinase (G-type RLK) might play important roles during SI process in tea plants.ConclusionCa2+ and K+ are important signal for SI in tea plants, and three genes including UGT74B1, MCU2 and G-type RLK play essential roles during SI signal transduction.

Classification and phylogenetic analyses of the Arabidopsis and tomato G-type lectin receptor kinases

BackgroundPathogen perception by plants is mediated by plasma membrane-localized immune receptors that have varied extracellular domains. Lectin receptor kinases (LecRKs) are among these receptors and are subdivided into 3 classes, C-type LecRKs (C-LecRKs), L-type LecRKs (L-LecRKs) and G-type LecRKs (G-LecRKs). While C-LecRKs are represented by one or two members in all plant species investigated and have unknown functions, L-LecRKs have been characterized in a few plant species and have been shown to play roles in plant defense against pathogens. Whereas Arabidopsis G-LecRKs have been characterized, this family of LecRKs has not been studied in tomato.ResultsThis investigation updates the current characterization of Arabidopsis G-LecRKs and characterizes the tomato G-LecRKs, using LecRKs from the monocot rice and the basal eudicot columbine to establish a basis for comparisons between the two core eudicots. Additionally, revisiting parameters established for Arabidopsis nomenclature for LecRKs is suggested for both Arabidopsis and tomato. Moreover, using phylogenetic analysis, we show the relationship among and between members of G-LecRKs from all three eudicot plant species. Furthermore, investigating presence of motifs in G-LecRKs we identified conserved motifs among members of G-LecRKs in tomato and Arabidopsis, with five present in at least 30 of the 38 Arabidopsis members and in at least 45 of the 73 tomato members.ConclusionsThis work characterized tomato G-LecRKs and added members to the currently characterized Arabidopsis G-LecRKs. Additionally, protein sequence analysis showed an expansion of this family in tomato as compared to Arabidopsis, and the existence of conserved common motifs in the two plant species as well as conserved species-specific motifs.

Ectopic Expression of GsSRK in Medicago sativa Reveals Its Involvement in Plant Architecture and Salt Stress Responses.

Receptor-like kinases (RLK) play fundamental roles in plant growth and stress responses. Compared with other RLKs, little information is provided concerning the S-locus LecRLK subfamily, which is characterized by an extracellular G-type lectin domain and an S-locus-glycop domain. Until now, the function of the G-type lectin domain is still unknown. In a previous research, we identified a Glycine soja S-locus LecRLK gene GsSRK, which conferred increased salt stress tolerance in transgenic Arabidopsis. In this study, to investigate the role of the G-type lectin domain and to breed transgenic alfalfa with superior salt stress tolerance, we transformed the full-length GsSRK (GsSRK-f) and a truncated version of GsSRK (GsSRK-t) deleting the G-type lectin domain into alfalfa. Our results showed that overexpression of GsSRK-t, but not GsSRK-f, resulted in changes of plant architecture, as evidenced by more branches but shorter shoots of GsSRK-t transgenic alfalfa, indicating a potential role of the extracellular G-type lectin domain in regulating plant architecture. Furthermore, we also found that transgenic alfalfa overexpressing either GsSRK-f or GsSRK-t showed increased salt stress tolerance, and GsSRK-t transgenic alfalfa displayed better growth (more branches and higher fresh weight) than GsSRK-f lines under salt stress. In addition, our results suggested that both GsSRK-f and GsSRK-t were involved in ion homeostasis, ROS scavenging, and osmotic regulation. Under salt stress, the Na+ content in the transgenic lines was significantly lower, while the K+ content was slightly higher than that in WT. Moreover, the transgenic lines displayed reduced ion leakage and MDA content, but increased SOD activity and proline content than WT. Notably, no obvious difference in these physiological indices was observed between GsSRK-f and GsSRK-t transgenic lines, implying that deletion of the GsSRK G-type lectin domain does not affect its physiological function in salt stress responses. In conclusion, results in this research reveal the dual role of GsSRK in regulating both plant architecture and salt stress responses.

Identification of chitosan oligosaccharides binding proteins from the plasma membrane of wheat leaf cell

Chitosan oligosaccharides (COS) have the ability to improve plant resistance to pests and diseases through activating plant immune system. However, it remains unclear whether stimulating reason of COS was associated with the plasma membrane proteins. Here, the interaction of COS with wheat leaf cell demonstrated that fluorescence-labeled COS were enriched on the cell surface and the interaction of COS with plasma membrane proteins was confirmed by quartz crystal microbalance (QCM) biosensor. What's more, HPLC and SDS-PAGE analysis showed that COS binding proteins exhibited more than three peaks and the molecular weight were 66 kDa to 97 kDa, where the COS binding proteins were fished out from wheat plasma membrane proteins by the COS affinity column. More importantly, LC-MS/MS analysis demonstrated that several candidates, including W5G2U8_WHEAT (a potential wall-associated receptor kinase protein), W5HY42_WHEAT and W5I0R4_WHEAT (potential G-type lectin S-receptor-like serine/threonine-protein kinase), have the potential to be COS receptors.

De Novo Transcriptome Study Identifies Candidate Genes Involved in Resistance to Austropuccinia psidii (Myrtle Rust) in Syzygium luehmannii (Riberry).

Austropuccinia psidii, causal agent of myrtle rust, was discovered in Australia in 2010 and has since become established on a wide range of species within the family Myrtaceae. Syzygium luehmannii, endemic to Australia, is an increasingly valuable berry crop. Plants were screened for responses to A. psidii inoculation, and specific resistance, in the form of localized necrosis, was determined in 29% of individuals. To understand the molecular basis underlying this response, mRNA was sequenced from leaf samples taken preinoculation, and at 24 and 48 h postinoculation, from four resistant and four susceptible plants. Analyses, based on de novo transcriptome assemblies for all plants, identified significant expression changes in resistant plants (438 transcripts) 48 h after pathogen exposure compared with susceptible plants (three transcripts). Most significantly up-regulated in resistant plants were gene homologs for transcription factors, receptor-like kinases, and enzymes involved in secondary metabolite pathways. A putative G-type lectin receptor-like kinase was exclusively expressed in resistant individuals and two transcripts incorporating toll/interleukin-1, nucleotide binding site, and leucine-rich repeat domains were up-regulated in resistant plants. The results of this study provide the first early gene expression profiles for a plant of the family Myrtaceae in response to the myrtle rust pathogen.

Proteomic analysis of Arabidopsis plasma membranes reveals lipopolysaccharide-responsive changes

Plant plasma membranes (PMs) contain pattern recognition receptors (PRRs), lately believed to be associated within multicomponent complexes, which perceive microbe-associated molecular pattern (MAMP) molecules like lipopolysaccharides (LPSs) and result in signal transduction events that lead to activated immune defense responses. As such, Arabidopsis thaliana leaves were treated with LPS from Escherichia coli (LPSE.coli) over time, and PM fractions isolated and evaluated using gel-based and subsequent mass spectrometry approaches for identification of LPS-responsive proteins. From the identified protein bands and spots, it is concluded that perception of hexaacylated LPS and resulting signal transduction occurs via PM-associated protein(s), amongst others, receptor-like kinases (RLKs) including G-type lectin S-receptor kinase and BAK1, and mostly likely within specialized perception domains.

GsSRK, a G-type lectin S-receptor-like serine/threonine protein kinase, is a positive regulator of plant tolerance to salt stress

Receptor-like protein kinases (RLKs) play vital roles in sensing outside signals, yet little is known about RLKs functions and roles in stress signal perception and transduction in plants, especially in wild soybean. Through the microarray analysis, GsSRK was identified as an alkaline (NaHCO3)-responsive gene, and was subsequently isolated from Glycine soja by homologous cloning. GsSRK encodes a 93.22kDa protein with a highly conserved serine/threonine protein kinase catalytic domain, a G-type lectin region, and an S-locus region. Real-time PCR results showed that the expression levels of GsSRK were largely induced by ABA, salt, and drought stresses. Over expression of GsSRK in Arabidopsis promoted seed germination, as well as primary root and rosette leaf growth during the early stages of salt stress. Compared to the wild type Arabidopsis, GsSRK overexpressors exhibited enhanced salt tolerance and higher yields under salt stress, with higher chlorophyll content, lower ion leakage, higher plant height, and more siliques at the adult developmental stage. Our studies suggest that GsSRK plays a crucial role in plant response to salt stress.

The Nicotiana attenuata LECTIN RECEPTOR KINASE 1 is involved in the perception of insect feeding

The Nicotiana attenuata LECTIN RECEPTOR KINASE 1 (LecRK1) has been recently identified as a component of the mechanism used by plants to suppress the Manduca sexta-triggered accumulation of salicylic acid (SA). The suppression of the SA burst by LecRK1 allows for the unfettered induction of jasmonic acid (JA)-mediated defense responses against M. sexta herbivory. LecRK1 contains a multi-domain extracellular region composed of a G-type Lectin domain and a PAN-AP domain separated by a variable sequence with low similarity to an EGF domain. The LecRK1 intracellular region is composed of a single domain structure with predicted Ser/Thr protein kinase activity. The multi-domain structure of the extracellular region of LecRK1 adds a level of complexity in terms of the potential ligands that this receptor protein could recognize.