Models of sequestration and receptor cross-talk for explaining multiple mutants in plant stem cell regulation

Background  

Stem cells reside in a plant's shoot meristem throughout its life and are main regulators of above-ground plant development. The stem cell maintenance depends on a feedback network between the CLAVATA and WUSCHEL genes. The CLAVATA3 peptide binds to the CLAVATA1 receptor leading to WUSCHEL inhibition. WUSCHEL, on the other hand, activates CLAVATA3 expression. Recent experiments suggest a second pathway where CLAVATA3 inhibits WUSCHEL via the CORYNE receptor pathway. An interesting question, central for understanding the receptor signaling, is why the clavata1-11 null mutant has a weaker phenotype compared with the clavata1-1 non-null mutant. It has been suggested that this relies on interference from the mutated CLAVATA1 acting on the CORYNE pathway.     

CLAVATA1-LIKE, a leucine-rich-repeat protein receptor kinase gene differentially expressed during adventitious caulogenesis in Pinus pinaster and Pinus pinea

Molecular cloning and characterization of a CLAVATA1-LIKE gene in Pinus pinaster and Pinus pinea is reported. CLAVATA1 is a well-characterized gene in Arabidopsis integral to the balance between primordial differentiation and meristem proliferation. Currently, it is not known if the Arabidopsis model of in vitro caulogenesis is applicable to conifers. In this work, we study the putative role of the CLAVATA1-LIKE gene during caulogenic induction in cotyledons of P. pinea and P. pinaster after exposure to benzyladenine. Expression analysis showed that the gene was differentially expressed during the first phases of adventitious caulogenesis in cotyledons from both species, suggesting that CLAVATA1-LIKE may play a role in caulogenesis in conifers. A binary vector carrying CLAVATA1-LIKE promoter driven GFP:GUS expression was constructed. Results of genetic transformation showed GUS activity during somatic embryogenic mass proliferation and embryo maturation.     

Genetic control of shoot and flower meristem behavior

Flowers and shoots are derived from specialized groups of stem cells termed meristems. Recent studies in Arabidopsis have identified factors that contribute to meristem structure and identity, such as CLAVATA1, CLAVATA3, and SHOOTMERISTEMLESS, which act in both shoot and flower meristems, as well as LEAFY and APETALA1 which specifically determine a floral fate.     

Shaping up: the genetic control of leaf shape

Leaf initiation at the shoot apical meristem involves a balance between cell proliferation and commitment to make primordia. Several genes, such as CLAVATA1, CLAVATA3, WUSCHEL, KNOTTED1, and PHANTASTICA, play key roles in these processes. When expressed in the leaf primordium, however, these 'meristem' genes can profoundly affect leaf shape and size, possibly by regulating hormone gradients and transport. The KNOTTED1-like genes are involved in regulating changes in hormonal levels. Recent studies have elaborated on the role that hormones, such as auxin, play in releasing biophysical constraints on leaf initiation and growth. Final leaf form is elaborated by a coordination of these hormonally regulated processes, cell division and cellular differentiation.     

thick tassel dwarf1 encodes a putative maize ortholog of the Arabidopsis CLAVATA1 leucine-rich repeat receptor-like kinase

Development in higher plants depends on the activity of meristems, formative regions that continuously initiate new organs at their flanks. Meristems must maintain a balance between stem cell renewal and organ initiation. In fasciated mutants, organ initiation fails to keep pace with meristem proliferation. The thick tassel dwarf1 (td1) mutation of maize affects both male and female inflorescence development. The female inflorescence, which results in the ear, is fasciated, with extra rows of kernels. The male inflorescence, or tassel, shows an increase in spikelet density. Floral meristems are also affected in td1 mutants; for example, male florets have an increase in stamen number. These results suggest that td1 functions in the inflorescence to limit meristem size. In addition, td1 mutants are slightly shorter than normal siblings, indicating that td1 also plays a role in vegetative development. td1 encodes a leucine-rich repeat receptor-like kinase (LRR-RLK) that is a putative ortholog of the Arabidopsis CLAVATA1 protein. These results complement previous work showing that fasciated ear2 encodes a CLAVATA2-like protein, and suggest that the CLAVATA signaling pathway is conserved in monocots. td1 maps in the vicinity of quantitative trait loci that affect seed row number, spikelet density and plant height. We discuss the possible selection pressures on td1 during maize domestication.     

The ULTRAPETALA gene controls shoot and floral meristem size in Arabidopsis

The regulation of proper shoot and floral meristem size during plant development is mediated by a complex interaction of stem cell promoting and restricting factors. The phenotypic effects of mutations in the ULTRAPETALA gene, which is required to control shoot and floral meristem cell accumulation in Arabidopsis thaliana, are described. ultrapetala flowers contain more floral organs and whorls than wild-type plants, phenotypes that correlate with an increase in floral meristem size preceding organ initiation. ultrapetala plants also produce more floral meristems than wild-type plants, correlating with an increase in inflorescence meristem size without visible fasciation. Expression analysis indicates that ULTRAPETALA controls meristem cell accumulation partly by limiting the domain of CLAVATA1 expression. Genetic studies show that ULTRAPETALA acts independently of ERA1, but has overlapping functions with PERIANTHIA and the CLAVATA signal transduction pathway in controlling shoot and floral meristem size and meristem determinacy. Thus ULTRAPETALA defines a novel locus that restricts meristem cell accumulation in Arabidopsis shoot and floral meristems.     

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.     

Molecular mechanisms controlling legume autoregulation of nodulation

Background

High input costs and environmental pressures to reduce nitrogen use in agriculture have increased the competitive advantage of legume crops. The symbiotic relationship that legumes form with nitrogen-fixing soil bacteria in root nodules is central to this advantage.

Scope

Understanding how legume plants maintain control of nodulation to balance the nitrogen gains with their energy needs and developmental costs will assist in increasing their productivity and relative advantage. For this reason, the regulation of nodulation has been extensively studied since the first mutants exhibiting increased nodulation were isolated almost three decades ago.

Conclusions

Nodulation is regulated primarily via a systemic mechanism known as the autoregulation of nodulation (AON), which is controlled by a CLAVATA1-like receptor kinase. Multiple components sharing homology with the CLAVATA signalling pathway that maintains control of the shoot apical meristem in arabidopsis have now been identified in AON. This includes the recent identification of several CLE peptides capable of activating nodule inhibition responses, a low molecular weight shoot signal and a role for CLAVATA2 in AON. Efforts are now being focused on directly identifying the interactions of these components and to identify the form that long-distance transport molecules take.     

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.     

The CLAVATA1 receptor-like kinase requires CLAVATA3 for its assembly into a signaling complex that includes KAPP and a Rho-related protein

The CLAVATA1 (CLV1) and CLAVATA3 (CLV3) genes are required to maintain the balance between cell proliferation and organ formation at the Arabidopsis shoot and flower meristems. CLV1 encodes a receptor-like protein kinase. We have found that CLV1 is present in two protein complexes in vivo. One is approximately 185 kD, and the other is approximately 450 kD. In each complex, CLV1 is part of a disulfide-linked multimer of approximately 185 kD. The 450-kD complex contains the protein phosphatase KAPP, which is a negative regulator of CLV1 signaling, and a Rho GTPase-related protein. In clv1 and clv3 mutants, CLV1 is found primarily in the 185-kD complex. We propose that CLV1 is present as an inactive disulfide-linked heterodimer and that CLV3 functions to promote the assembly of the active 450-kD complex, which then relays signal transduction through a Rho GTPase.     

Orthologs of arabidopsis CLAVATA 1 gene in cultivated Brassicaceae plants

In arabidopsis (Arabidopsis thaliana), the CLAVATA1 (CLV1) gene is involved in maintaining the balance between the stem cells in the central zone of the stem apical meristem and the determined cells at its periphery. However, CLV1 has not been previously characterized in other Brassicaceae. Using the direct amplification of genomic DNA, we obtained a full-length CLV1 ortholog from canola plants (Brassica napus), and also three CLV1 fragments from rape (B. rapa), canola (B. napus), and false flax (Camelina sativa), which corresponded to the transmembrane domain and a part of the kinase domain of the CLAVATA1 protein. The nucleotide and deduced amino acid sequences of the full-size CLV1 ortholog from B. napus were similar by 81 and 87% to the prototype gene from arabidopsis; in the case of shorter gene fragments, the similarity was as high as 91-93 and 98%, respectively. By their primary structure, the CLV1 genes in the Brassicaceae considerably differ from its putative structural homologs beyond this family.     

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.     

Plant signalling peptides: some recent developments.

The subject of this review is plant signalling peptides, peptides of a new generation which regulate growth, differentiation, and other plant physiological functions. These peptides include systemin, the phytosulfokines (PSKs), ENOD40, CLAVATA3, Locus-S, POLARIS, IDA, and ROT4. On the basis of literature data and our own results we discuss their structure, biological properties, and structure/biological function relationship, especially for the more studied systemin and PSK-alpha.     

Gain-of-function phenotypes of chemically synthetic CLAVATA3/ESR-related (CLE) peptides in Arabidopsis thaliana and Oryza sativa

Using 26 chemically synthetic CLAVATA3/ESR (CLE) peptides, which correspond to the predicted products of the 31 Arabidopsis CLE genes, we investigated the CLE peptide function in Arabidopsis and rice. Treatment with some CLE peptides inhibited root elongation in rice as well as in Arabidopsis. It also reduced the size of the shoot apical meristem in Arabidopsis but not in rice. Database searches revealed 47 putative CLE genes in the rice genome and multiple CLE domains in some CLE genes, indicating diverse CLE function in these plants.     

Plant cytokine or phytocytokine

Peptide hormones play an important role in plant growth and development. Some of them are secreted by stem cells and also regulate plant immunity through cell-cell communication and reprogramming the expression of immune related genes, such as CLAVATA3p (CLV3p) and phytosulfokine (PSK). These peptides play similar roles as cytokines in plant innate immunity. As explosive progress of plant omics, more and more such functional peptides will be discovered. I recommend that they should be named as plant cytokines or phytocytokines. This nomenclature will be convenient for study of plant secretory peptides and plant innate immunity.     

CLE Peptides in Plants: Proteolytic Processing,Structure-Activity Relationship,and Ligand-Receptor Interaction

Ligand-receptor signaling initiated by the CLAVATA3/ ENDOSPERM SURROUNDING REGION (CLE) family peptides is critical in regulating cell division and differentiation in meristematic tissues in plants. Biologically active CLE peptides are released from precursor proteins via proteolytic processing. The mature form of CLE ligands consists of 12–13 amino acids with several post-translational modifications. This review summarizes recent progress toward understanding the proteolytic activities that cleave precursor proteins to release CLE peptides, the molecular structure and function of mature CLE ligands, and interactions between CLE ligands and corresponding leucine-rich repeat (LRR) receptor-like kinases (RLKs).