Enhanced resistance to soybean cyst nematode Heterodera glycines in transgenic soybean by silencing putative CLE receptors

CLE peptides are small extracellular proteins important in regulating plant meristematic activity through the CLE‐receptor kinase‐WOX signalling module. Stem cell pools in the SAM (shoot apical meristem), RAM (root apical meristem) and vascular cambium are controlled by CLE signalling pathways. Interestingly, plant‐parasitic cyst nematodes secrete CLE‐like effector proteins, which act as ligand mimics of plant CLE peptides and are required for successful parasitism. Recently, we demonstrated that Arabidopsis CLE receptors CLAVATA1 (CLV1), the CLAVATA2 (CLV2)/CORYNE (CRN) heterodimer receptor complex and RECEPTOR‐LIKE PROTEIN KINASE 2 (RPK2), which transmit the CLV3 signal in the SAM, are required for perception of beet cyst nematode Heterodera schachtii CLEs. Reduction in nematode infection was observed in clv1, clv2, crn, rpk2 and combined double and triple mutants. In an effort to develop nematode resistance in an agriculturally important crop, orthologues of Arabidopsis receptors including CLV1, CLV2, CRN and RPK2 were identified from soybean, a host for the soybean cyst nematode Heterodera glycines. For each of the receptors, there are at least two paralogues in the soybean genome. Localization studies showed that most receptors are expressed in the root, but vary in their level of expression and spatial expression patterns. Expression in nematode‐induced feeding cells was also confirmed. In vitro direct binding of the soybean receptors with the HgCLE peptide was analysed. Knock‐down of the receptors in soybean hairy roots showed enhanced resistance to SCN. Our findings suggest that targeted disruption of nematode CLE signalling may be a potential means to engineer nematode resistance in crop plants.     

Perception of root‐active CLE peptides requires CORYNE function in the phloem vasculature

Arabidopsis root development is orchestrated by signaling pathways that consist of different CLAVATA3/EMBRYO SURROUNDING REGION (CLE) peptide ligands and their cognate CLAVATA (CLV) and BARELY ANY MERISTEM (BAM) receptors. How and where different CLE peptides trigger specific morphological or physiological changes in the root is poorly understood. Here, we report that the receptor‐like protein CLAVATA 2 (CLV2) and the pseudokinase CORYNE (CRN) are necessary to fully sense root‐active CLE peptides. We uncover BAM3 as the CLE45 receptor in the root and biochemically map its peptide binding surface. In contrast to other plant peptide receptors, we found no evidence that SOMATIC EMBRYOGENESIS RECEPTOR KINASE (SERK) proteins act as co‐receptor kinases in CLE45 perception. CRN stabilizes BAM3 expression and thus is required for BAM3‐mediated CLE45 signaling. Moreover, protophloem‐specific CRN expression complements resistance of the crn mutant to root‐active CLE peptides, suggesting that protophloem is their principal site of action. Our work defines a genetic framework for dissecting CLE peptide signaling and CLV/BAM receptor activation in the root.     

The Cysteine Pairs in CLV2 are Not Necessary for Sensing the CLV3 Peptide in Shoot and Root Meristems

Receptor-like proteins (RLPs) are involved in both plant defense and developmental processes. Previous genetic and biochemical studies show that the leucine-rich repeat (LRR) receptor-like protein CLAVATA2 (CLV2) functions together with CLAVATA1 (CLV1) and CORYNE (CRN) in Arabidopsis to limit the stem cell number in shoot apical meristem, while in root it acts with CRN to trigger a premature differentiation of the stem cells after sensing the exogenously applied peptides of CLV3p, CLE19p or CLE40p. It has been proposed that disulfide bonds might be formed through two cysteine pairs in the extracellular LRR domains of CLV1 and CLV2 to stabilize the receptor complex. Here we tested the hypothesis by replacing these cysteines with alanines and showed that depletions of one or both of the cysteine pairs do not hamper the function of CLV2 in SAM maintenance. In vitro peptide assay also showed that removal of the cysteine pairs did not affect the perception of CLV3 peptides in roots. These observations allow us to conclude that the formation of disulfide bonds is not needed for the function of CLV2.     

CLAVATA2 forms a distinct CLE‐binding receptor complex regulating Arabidopsis stem cell specification

CLAVATA1 (CLV1), CLV2, CLV3, CORYNE (CRN), BAM1 and BAM2 are key regulators that function at the shoot apical meristem (SAM) of plants to promote differentiation by limiting the size of the organizing center that maintains stem cell identity in neighboring cells. Previous results have indicated that the extracellular domain of the receptor kinase CLV1 binds to the CLV3‐derived CLE ligand. The biochemical role of the receptor‐like protein CLV2 has remained largely unknown. Although genetic analysis suggested that CLV2, together with the membrane kinase CRN, acts in parallel with CLV1, recent studies using transient expression indicated that CLV2 and CRN from a complex with CLV1. Here, we report detection of distinct CLV2‐CRN heteromultimeric and CLV1‐BAM multimeric complexes in transient expression in tobacco and in Arabidopsis meristems. Weaker interactions between the two complexes were detectable in transient expression. We also find that CLV2 alone generates a membrane‐localized CLE binding activity independent of CLV1. CLV2, CLV1 and the CLV1 homologs BAM1 and BAM2 all bind to the CLV3‐derived CLE peptide with similar kinetics, but BAM receptors show a broader range of interactions with different CLE peptides. Finally, we show that BAM and CLV1 overexpression can compensate for the loss of CLV2 function in vivo. These results suggest two parallel ligand‐binding receptor complexes controlling stem cell specification in Arabidopsis.     

Evidence for functional conservation, sufficiency, and proteolytic processing of the CLAVATA3 CLE domain

Arabidopsis (Arabidopsis thaliana) CLAVATA3 (CLV3) is hypothesized to act as a ligand for the CLV1 receptor kinase in the regulation of stem cell specification at shoot and flower meristems. CLV3 is a secreted protein, with an amino-terminal signal sequence and a conserved C-terminal domain of 15 amino acids, termed the CLE (CLV3/ESR-related) domain, based on its similarity to a largely unstudied protein family broadly present in land plants. We have tested the function of 13 Arabidopsis CLEs in vivo and found a significant variability in the ability of CLEs to replace CLV3, ranging from complete to no complementation. The best rescuing CLE depends on CLV1 for function, while other CLEs act independently of CLV1. Domain-swap experiments indicate that differences in function can be traced to the CLE domain within these proteins. Indeed, when the CLE domain of CLV3 is placed downstream of an unrelated signal sequence, it is capable of fully replacing CLV3 function. Finally, we have detected proteolytic activity in extracts from cauliflower (Brassica oleracea) that process both CLV3 and CLE1 at their C termini. For CLV3, processing appears to occur at the absolutely conserved arginine-70 found at the beginning of the CLE domain. We propose that CLV3 and other CLEs are C-terminally processed to generate an active CLE peptide.     

CLE14/CLE20 peptides may interact with CLAVATA2/CORYNE receptor-like kinases to irreversibly inhibit cell division in the root meristem of Arabidopsis

Towards an understanding of the interacting nature of the CLAVATA (CLV) complex, we predicted the 3D structures of CLV3/ESR-related (CLE) peptides and the ectodomain of their potential receptor proteins/kinases, and docking models of these molecules. The results show that the ectodomain of CLV1 can form homodimers and that the 12-/13-amino-acid CLV3 peptide fits into the binding clefts of the CLV1 dimers. Our results also demonstrate that the receptor domain of CORYNE (CRN), a recently identified receptor-like kinase, binds tightly to the ectodomain of CLV2, and this likely leads to an increased possibility for docking with CLV1. Furthermore, our docking models reveal that two CRN-CLV2 ectodomain heterodimers are able to form a tetramer receptor complex. Peptides of CLV3, CLE14, CLE19, and CLE20 are also able to bind a potential CLV2-CRN heterodimer or heterotetramer complex. Using a cell-division reporter line, we found that synthetic 12-amino-acid CLE14 and CLE20 peptides inhibit, irreversibly, root growth by reducing cell division rates in the root apical meristem, resulting in a short-root phenotype. Intriguingly, we observed that exogenous application of cytokinin can partially rescue the short-root phenotype induced by over-expression of either CLE14 or CLE20 in planta. However, cytokinin treatment does not rescue the short-root phenotype caused by exogenous application of the synthetic CLE14/CLE20 peptides, suggesting a requirement for a condition provided only in living plants. These results therefore imply that the CLE14/CLE20 peptides may act through the CLV2-CRN receptor kinase, and that their availabilities and/or abundances may be affected by cytokinin activity in planta.     

Identification of potential host plant mimics of CLAVATA3/ESR (CLE)-like peptides from the plant-parasitic nematode Heterodera schachtii

In this article, we present the cloning of two CLAVATA3/ESR (CLE)-like genes, HsCLE1 and HsCLE2, from the beet cyst nematode Heterodera schachtii, a plant-parasitic cyst nematode with a relatively broad host range that includes the model plant Arabidopsis. CLEs are small secreted peptide ligands that play important roles in plant growth and development. By secreting peptide mimics of plant CLEs, the nematode can developmentally reprogramme root cells for the formation of unique feeding sites within host roots for its own benefit. Both HsCLE1 and HsCLE2 encode small secreted polypeptides with a conserved C-terminal CLE domain sharing highest similarity to Arabidopsis CLEs 1-7. Moreover, HsCLE2 contains a 12-amino-acid CLE motif that is identical to AtCLE5 and AtCLE6. Like all other plant and nematode CLEs identified to date, HsCLEs caused wuschel-like phenotypes when overexpressed in Arabidopsis, and this activity was abolished when the proteins were expressed without the CLE motif. HsCLEs could also function in planta without a signal peptide, highlighting the unique, yet conserved function of nematode CLE variable domains in trafficking CLE peptides for secretion. In a direct comparison of HsCLE2 overexpression phenotypes with those of AtCLE5 and AtCLE6, similar shoot and root phenotypes were observed. Exogenous application of 12-amino-acid synthetic peptides corresponding to the CLE motifs of HsCLEs and AtCLE5/6 suggests that the function of this class of CLEs may be subject to complex endogenous regulation. When seedlings were grown on high concentrations of peptide (10 μm), root growth was suppressed; however, when seedlings were grown on low concentrations of peptide (0.1 μm), root growth was stimulated. Together, these findings indicate that AtCLEs1-7 may be the target peptides mimicked by HsCLEs to promote parasitism.     

Analysis of interactions among the CLAVATA3 receptors reveals a direct interaction between CLAVATA2 and CORYNE in Arabidopsis

In Arabidopsis, CORYNE (CRN), a new member of the receptor kinase family, was recently isolated as a key player involved in the CLAVATA3 (CLV3) signaling pathway, thereby playing an important role in regulating the development of shoot and root apical meristems. However, the precise relationships among CLAVATA1 (CLV1), CLAVATA2 (CLV2), and CRN receptors remain unclear. Here, we demonstrate the subcellular localization of CRN and analyze the interactions among CLV1, CLV2, and CRN using firefly luciferase complementation imaging (LCI) assays in both Arabidopsis mesophyll protoplasts and Nicotiana benthamiana leaves. Fluorescence targeting showed that CRN was localized to the plasma membrane. The LCI assays coupled with co‐immunoprecipitation assays demonstrated that CLV2 can directly interact with CRN in the absence of CLV3. Additional LCI assays showed that CLV1 did not interact with CLV2, but can interact weakly with CRN. We also found that CLV1 can interact with CLV2–CRN heterodimers, implying that these three proteins may form a complex. Moreover, CRN, rather than CLV1 and CLV2, was able to form homodimers without CLV3 stimulation. Taken together, our results add direct evidence to the newly proposed two‐parallel receptor pathways model and therefore provide new insights into the CLV3 signaling pathway.     

In Planta Processing and Glycosylation of a Nematode CLAVATA3/ENDOSPERM SURROUNDING REGION-Like Effector and Its Interaction with a Host CLAVATA2-Like Receptor to Promote Parasitism

Like other biotrophic plant pathogens, plant-parasitic nematodes secrete effector proteins into host cells to facilitate infection. Effector proteins that mimic plant CLAVATA3/ENDOSPERM SURROUNDING REGION-related (CLE) proteins have been identified in several cyst nematodes, including the potato cyst nematode (PCN); however, the mechanistic details of this cross-kingdom mimicry are poorly understood. Plant CLEs are posttranslationally modified and proteolytically processed to function as bioactive ligands critical to various aspects of plant development. Using ectopic expression coupled with nanoliquid chromatography-tandem mass spectrometry analysis, we show that the in planta mature form of proGrCLE1, a multidomain CLE effector secreted by PCN during infection, is a 12-amino acid arabinosylated glycopeptide (named GrCLE1-1Hyp4,7g) with striking structural similarity to mature plant CLE peptides. This glycopeptide is more resistant to hydrolytic degradation and binds with higher affinity to a CLAVATA2-like receptor (StCLV2) from potato (Solanum tuberosum) than its nonglycosylated forms. We further show that StCLV2 is highly up-regulated at nematode infection sites and that transgenic potatoes with reduced StCLV2 expression are less susceptible to PCN infection, indicating that interference of the CLV2-mediated signaling pathway confers nematode resistance in crop plants. These results strongly suggest that phytonematodes have evolved to utilize host cellular posttranslational modification and processing machinery for the activation of CLE effectors following secretion into plant cells and highlight the significance of arabinosylation in regulating nematode CLE effector activity. Our finding also provides evidence that multidomain CLEs are modified and processed similarly to single-domain CLEs, adding new insight into CLE maturation in plants.Plants are vulnerable to attack by plant-parasitic nematodes. The cyst-forming endoparasitic nematodes (Globodera and Heterodera spp.) are among the most damaging plant pathogens, causing tremendous crop losses globally (Chitwood, 2003). Cyst nematodes have evolved an intimate parasitic relationship with their hosts by transforming normal root cells into a unique feeding structure called a syncytium that serves as the sole nutritive source required for subsequent growth and development (Hussey and Grundler, 1998; Davis et al., 2004). Cyst nematodes are soil-borne pathogens. Once infective juveniles hatch in the soil, they penetrate into the roots of host plants and select a single cell near the root vasculature to initiate a syncytium. The syncytium forms by the fusion of cells adjacent to the initial syncytial cell through extensive cell wall dissolution and develops into a large fused cell that is highly metabolically active and contains numerous enlarged nuclei and nucleoli (Endo, 1964). Like other plant pathogens, cyst nematodes use secreted effector proteins to facilitate plant parasitism. Effector proteins, originating from the nematode esophageal gland cells (two subventral and one dorsal) and secreted into root tissues through the nematode stylet (a mouth spear), represent important molecular signals that manipulate various host cellular processes to redifferentiate normal root cells into a syncytium (Davis et al., 2004; Mitchum et al., 2008, 2013).Genes encoding effector proteins with sequence similarity to plant CLAVATA3/ENDOSPERM SURROUNDING REGION-related (CLE) proteins have recently been cloned from several cyst nematode species, including the potato cyst nematode (PCN [Globodera rostochiensis; Gr]; Wang et al., 2001, 2011; Gao et al., 2003; Lu et al., 2009), a regulated and devastating pest of potato (Solanum tuberosum [St]) and tomato (Solanum lycopersicum) crops. Plant CLE proteins, identified from diverse monocot and dicot species (Cock and McCormick, 2001; Oelkers et al., 2008), are a class of peptide hormones that regulate many aspect of plant growth and development (Yamada and Sawa, 2013). Plant CLE genes encode small proteins that contain an N-terminal signal peptide, an internal variable domain, and either a single or multiple conserved C-terminal CLE domain(s) (Cock and McCormick, 2001; Kinoshita et al., 2007; Oelkers et al., 2008). The Arabidopsis (Arabidopsis thaliana [At]) genome encodes at least 32 single-domain CLEs, of which CLAVATA3 (CLV3) is the best characterized member. CLV3 is found to interact with three major membrane-associated receptor complexes, CLV1, CLV2-CORYNE (CRN), and RECEPTOR LIKE PROTEIN KINASE2 (RPK2; Clark et al., 1993; Jeong et al., 1999; Müller et al., 2008; Kinoshita et al., 2010; Zhu et al., 2010), to control the fate of stem cells in the shoot apical meristem (Fletcher et al., 1999). Among the three CLV3 receptors, CLV1 and RPK2 are leucine-rich repeat (LRR) receptor-like kinases, whereas CLV2 is an LRR receptor-like protein that acts together with a membrane-associated protein kinase, CRN, to transmit the CLV3 signal. The 96-amino acid CLV3 precursor is proteolytically processed into a mature 13-amino acid arabinosylated glycopeptide derived from its CLE domain, in which one (at position 7) of the two Hyp residues (at positions 4 and 7) is further modified by the addition of three units of l-Ara (Ohyama et al., 2009). The mature CLV3 glycopeptide exhibits full biological activity and binds to the LRR domain of CLV1 more strongly than nonarabinosylated forms (Ohyama et al., 2009). Hyp arabinosylation, a posttranslational modification unique to plants, also has been observed in mature CLE2 and CLE9 peptides from Arabidopsis as well as in CLE-ROOT SIGNAL2, an Arabidopsis CLE2 ortholog that controls nodulation in Lotus japonicus (Lj; Ohyama et al., 2009; Shinohara et al., 2012; Okamoto et al., 2013), where the arabinoside chains are revealed to have important roles in biological activity, receptor binding, and peptide conformation (Shinohara and Matsubayashi, 2013). Many Arabidopsis CLE genes are expressed in roots (Sharma et al., 2003; Jun et al., 2010), and evidence is emerging that CLE-receptor signaling pathways regulate root meristem function (Stahl et al., 2009, 2013; Meng and Feldman, 2010).Nematode CLE genes are expressed exclusively within the dorsal gland cell and encode secreted proteins with the characteristic CLE motif(s) at their C termini (Mitchum et al., 2008; Lu et al., 2009; Wang et al., 2011). Outside the conserved CLE motif, there is no sequence similarity between nematode and plant CLE proteins. The dramatic up-regulation of CLE genes in parasitic stages of the nematode life cycle (Wang et al., 2001, 2010b, 2011; Gao et al., 2003; Lu et al., 2009), along with the observation that transgenic plants expressing double-stranded RNA complementary to nematode CLE genes are less susceptible to nematode infection (Patel et al., 2008), have made it clear that CLE effectors play a critical role in nematode parasitism. Nematode-encoded CLE genes are the only CLE genes that have been identified outside the plant kingdom. Several lines of evidence suggest that nematode CLEs function as peptide mimics of endogenous plant CLEs. First, overexpression of nematode CLE genes in Arabidopsis generated phenotypes similar to those of plant CLE gene overexpression (Wang et al., 2005, 2011; Lu et al., 2009). Second, expression of nematode-encoded CLE genes in the shoot apical meristem of an Arabidopsis clv3-2 null mutant partially or completely rescued the mutant phenotypes (Lu et al., 2009; Wang et al., 2010b). Lastly, recent studies showed that Arabidopsis receptors, including CLV1, CLV2-CRN, and RPK2, are expressed in syncytia induced by the beet cyst nematode (BCN; Heterodera schachtii) and that receptor mutants fail to respond to BCN CLE peptides and show increased resistance to BCN infection (Replogle et al., 2011, 2013), further bolstering the notion of nematode-secreted CLE effectors as peptide mimics of endogenous plant CLEs and the importance of nematode CLE signaling in plant parasitism.Plant CLE precursors undergo posttranslational modifications and proteolytic processing to become bioactive CLE peptides (Shinohara and Matsubayashi, 2010; Shinohara et al., 2012; Okamoto et al., 2013). To fulfill a role as peptide mimics of plant CLEs, nematode CLEs are presumably recognized by the existing host modification and processing machinery for maturation. However, until now, the bioactive form of nematode-secreted CLEs that acts in planta has not been described. In addition, cyst nematodes are specialist feeders. Many agriculturally important nematode species, such as PCN, the soybean cyst nematode (Heterodera glycines), and the cereal cyst nematode (Heterodera avenae), fail to infect Arabidopsis. The mechanism of perception of nematode-secreted CLEs in crop plants still awaits investigation. In this study, we explored the molecular basis of CLE mimicry in the PCN-potato pathosystem. Using ectopic expression coupled with nanoliquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) analysis, we determined that the in planta mature form of proGrCLE1, a representative and multidomain CLE effector secreted from PCN during infection (Lu et al., 2009), is a 12-amino acid arabinosylated glycopeptide (hereafter referred to as GrCLE1-1Hyp4,7g) similar in structure to bioactive plant CLE peptides. We further cloned a CLV2-like gene from potato (hereafter referred to as StCLV2). We found that the GrCLE1-1Hyp4,7g glycopeptide binds directly to the StCLV2 ectodomain with high affinity and that transgenic potato lines with reduced StCLV2 expression are less susceptible to PCN infection. Our data provide direct evidence that nematode-secreted CLE effectors can be recognized by existing host cellular machinery to become bioactive mimics of endogenous plant CLE signals and suggest that cyst nematodes tap into the conserved CLV2 signaling pathway to promote successful infection of crop plants.     

The receptor-like kinase SOL2 mediates CLE signaling in Arabidopsis

Arabidopsis sol2 mutants showed CLV3 peptide resistance. Twenty-six synthetic CLE peptides were examined in the clv1, clv2 and sol2 mutants. sol2 showed different levels of resistance to the various peptides, and the spectrum of peptide resistance was quite similar to that of clv2. SOL2 encoded a receptor-like kinase protein which is identical to CORYNE (CRN). GeneChip analysis revealed that the expression of several genes was altered in the sol2 root tip. Here, we suggest that SOL2, together with CLV2, plays an important role in the regulation of root meristem development through the CLE signaling pathway.     

Variable domain I of nematode CLEs directs post-translational targeting of CLE peptides to the extracellular space

Effector proteins expressed in the esophageal gland cells of cyst nematodes are delivered into plant cells through a hollow, protrusible stylet. Although evidence indicates that effector proteins function in the cytoplasm of the syncytium,^1–3 technical constraints have made it difficult to directly determine where nematode effector proteins are initially delivered: cytoplasm, extracellular space, or both. Recently, we demonstrated that soybean cyst nematode CLE (HgCLE) propeptides are delivered to the cytoplasm of syncytial cells. Genetic and biochemical analyses indicate that the variable domain (VD) sequence is then required for targeting cytoplasmically delivered nematode CLEs to the apoplast where they function as ligand mimics of endogenous plant CLE peptides.⁴ The fact that nematode CLEs are targeted through the gland cell secretory pathway and delivered as mature propeptides into plant cells makes it impossible for these proteins to be subsequently delivered to the extracellular space via co-translational translocation through the endoplasmic reticulum (ER) secretory pathway of the host cell. However, when expressed in transgenic plants, if the mature nematode CLE propeptide harbored a functional cryptic signal peptide, it could possibly traffic to the apoplast through the ER secretory pathway by co-translational translocation. Here, we present evidence that VDI, the N-terminal sequence of the VD of HgCLE2,⁴ is sufficient for trafficking CLE peptides to the apoplast and that trafficking is indeed through an alternative pathway other than co-translational translocation.Key words: cyst nematode, effector, CLE, variable domain, trafficking, endoplasmic reticulum, co-translational translocation, post-translational     

The 14-amino acid CLV3, CLE19, and CLE40 peptides trigger consumption of the root meristem in Arabidopsis through a CLAVATA2-dependent pathway

CLAVATA3 (CLV3), CLV3/ESR19 (CLE19), and CLE40 belong to a family of 26 genes in Arabidopsis thaliana that encode putative peptide ligands with unknown identity. It has been shown previously that ectopic expression of any of these three genes leads to a consumption of the root meristem. Here, we show that in vitro application of synthetic 14-amino acid peptides, CLV3p, CLE19p, and CLE40p, corresponding to the conserved CLE motif, mimics the overexpression phenotype. The same result was observed when CLE19 protein was applied externally. Interestingly, clv2 failed to respond to the peptide treatment, suggesting that CLV2 is involved in the CLE peptide signaling. Crossing of the CLE19 overexpression line with clv mutants confirms the involvement of CLV2. Analyses using tissue-specific marker lines revealed that the peptide treatments led to a premature differentiation of the ground tissue daughter cells and misspecification of cell identity in the pericycle and endodermis layers. We propose that these 14-amino acid peptides represent the major active domain of the corresponding CLE proteins, which interact with or saturate an unknown cell identity-maintaining CLV2 receptor complex in roots, leading to consumption of the root meristem.     

Mechanisms of molecular mimicry of plant CLE peptide ligands by the parasitic nematode Globodera rostochiensis

Nematodes that parasitize plant roots cause huge economic losses and have few mechanisms for control. Many parasitic nematodes infect plants by reprogramming root development to drive the formation of feeding structures. How nematodes take control of plant development is largely unknown. Here, we identify two host factors involved in the function of a receptor ligand mimic, GrCLE1, secreted by the potato cyst nematode Globodera rostochiensis. GrCLE1 is correctly processed to an active form by host plant proteases. Processed GrCLE1 peptides bind directly to the plant CLE receptors CLV2, BAM1, and BAM2. Involvement of these receptors in the ligand-mimicking process is also supported by the fact that the ability of GrCLE1 peptides to alter plant root development in Arabidopsis (Arabidopsis thaliana) is dependent on these receptors. Critically, we also demonstrate that GrCLE1 maturation can be entirely carried out by plant factors and that the availability of CLE processing activity may be essential for successful ligand mimicry.     

The receptor kinase CORYNE of Arabidopsis transmits the stem cell-limiting signal CLAVATA3 independently of CLAVATA1

Stem cells in shoot and floral meristems of Arabidopsis thaliana secrete the signaling peptide CLAVATA3 (CLV3) that restricts stem cell proliferation and promotes differentiation. The CLV3 signaling pathway is proposed to comprise the receptor kinase CLV1 and the receptor-like protein CLV2. We show here that the novel receptor kinase CORYNE (CRN) and CLV2 act together, and in parallel with CLV1, to perceive the CLV3 signal. Mutations in CRN cause stem cell proliferation, similar to clv1, clv2, and clv3 mutants. CRN has additional functions during plant development, including floral organ development, that are shared with CLV2. The CRN protein lacks a distinct extracellular domain, and we propose that CRN and CLV2 interact via their transmembrane domains to establish a functional receptor.     

Diverse and conserved roles of CLE peptides

The function of plant CLAVATA3 (CLV3)/ENDOSPERM SURROUNDING REGION (ESR) (CLE) peptides in shoot meristem differentiation has been expanded in recent years to implicate roles in root growth and vascular development among different CLE family members. Recent evidence suggests that nematode pathogens within plant roots secrete ligand mimics of plant CLE peptides to modify selected host cells into multinucleate feeding sites. This discovery demonstrated an unprecedented adaptation of an animal gene product to functionally mimic a plant peptide involved in cellular signaling for parasitic benefit. This review highlights the diverse and conserved role of CLE peptides in these different contexts.