Assembly of cell wall polysaccharides into specific patterns is required for plant growth. A complex of RAPID ALKALINIZATION FACTOR 4 (RALF4) and its cell wall–anchored LEUCINE-RICH REPEAT EXTENSIN 8 (LRX8)–interacting protein is crucial for cell wall integrity during pollen tube growth, but its molecular connection with the cell wall is unknown. Here, we show that LRX8-RALF4 complexes adopt a heterotetrametric configuration in vivo, displaying a dendritic distribution. The LRX8-RALF4 complex specifically interacts with demethylesterified pectins in a charge-dependent manner through RALF4’s polycationic surface. The LRX8-RALF4-pectin interaction exerts a condensing effect, patterning the cell wall’s polymers into a reticulated network essential for wall integrity and expansion. Our work uncovers a dual structural and signaling role for RALF4 in pollen tube growth and in the assembly of complex extracellular polymers.

The embryonic cuticle is necessary for normal seed development and seedling establishment in Arabidopsis. Although mutants with defective embryonic cuticles have been identified, neither the deposition of cuticle material, nor its regulation, has been described during embryogenesis. Here we use electron microscopy, cuticle staining and permeability assays to show that cuticle deposition initiates de novo in patches on globular embryos. By combining these techniques with genetics and gene expression analysis, we show that successful patch coalescence to form a continuous cuticle requires a signalling involving the endosperm-specific subtilisin protease ALE1 and the receptor kinases GSO1 and GSO2, which are expressed in the developing embryonic epidermis. Transcriptome analysis shows that this pathway regulates stress-related gene expression in seeds. Consistent with these findings we show genetically, and through activity analysis, that the stress-associated MPK6 protein acts downstream of GSO1 and GSO2 in the developing embryo. We propose that a stress-related signalling pathway has been hijacked in some angiosperm seeds through the recruitment of endosperm-specific components. Our work reveals the presence of an inter-compartmental dialogue between the endosperm and embryo that ensures the formation of an intact and functional cuticle around the developing embryo through an “auto-immune” type interaction. Plant embryogenesis occurs deep within the tissues of the developing seed, and leads to the production of the mature embryo. In Arabidopsis and many other plant species embryo-derive structure (such as the cotyledons) are suddenly exposed to environmental stresses such as low humidity. In these species the embryonic cuticle provides a primary defence against environmental stress, and particularly dehydration, at germination. The formation of an intact and functional cuticle during embryogenesis is thus of key importance for seedling survival. Our work shows that a signalling pathway involving receptor-kinases expressed in the embryo epidermis, and a protease expressed in the endosperm tissue surrounding the embryo, is critical for ensuring the production of an intact cuticle. Furthermore, we show that a component of stress-related MAP-Kinase signalling in plants acts downstream in this pathway, possibly to mediate transcriptional responses characteristic of responses to stress. We propose that plants have redeployed a signalling pathway associated with stress resistance to ensure the formation of an intact embryonic cuticle prior to germination, and thus ensure seedling survival at germination.

Complex cell-to-cell communication between the male pollen tube and the female reproductive organs is required for plant fertilization. A family ofCatharanthus roseusreceptor kinase 1-like (CrRLK1L) membrane receptors has been genetically implicated in this process. Here, crystal structures of theCrRLK1Ls ANXUR1 and ANXUR2 are reported at 1.48 and 1.1 Å resolution, respectively. The structures reveal a novel arrangement of two malectin-like domains connected by a short β-hairpin linker and stabilized by calcium ions. The canonical carbohydrate-interaction surfaces of related animal and bacterial carbohydrate-binding modules are not conserved in plantCrRLK1Ls. In line with this, the binding of chemically diverse oligosaccharides to ANXUR1 and HERCULES1 could not be detected. Instead,CrRLK1Ls have evolved a protein–protein interface between their malectin domains which forms a deep cleft lined by highly conserved aromatic and polar residues. Analysis of the glycosylation patterns of differentCrRLK1Ls and their oligomeric states suggests that this cleft could resemble a binding site for a ligand required for receptor activation ofCrRLK1Ls.

The embryonic cuticle is necessary for normal seed development and seedling establishment in Arabidopsis. Although mutants with defective embryonic cuticles have been identified, neither the deposition of cuticle material, nor its regulation, has been described during embryogenesis. Here we use electron microscopy, lipid staining and permeability assays to show that cuticle deposition initiates de novo in patches on globular embryos. By combining these techniques with genetics and gene expression analysis, we show that successful patch coalescence to form a continuous cuticle requires a signalling involving the endosperm-specific subtilisin protease ALE1 and the receptor kinases GSO1 and GSO2, which are expressed in the developing embryonic epidermis. Transcriptome analysis shows that this pathway regulates stress-related gene expression in seeds. Consistent with these findings we show genetically, and through activity analysis, that the stress-associated MPK6 protein acts downstream of GSO1 and GSO2 in the developing embryo. We propose that a stress-related signalling pathway has been hijacked in some angiosperm seeds through the recruitment of endosperm-specific components. Our work reveals the presence of an inter-compartmental dialogue between the endosperm and embryo that ensures the formation of an intact and functional cuticle around the developing embryo through an “auto-immune” type interaction.

A complex cell-to-cell communication between the male pollen tube and the female reproductive organs is required for plant fertilization. A family of Catharanthus roseus Receptor Kinase-1 ( Cr RLK1L) membrane receptors has been genetically implicated in this process. Here we present crystal structures of the CrRLK1Ls ANXUR1 and 2 at 1.48 and 1.1 Å resolution, respectively. Our structures reveal a novel arrangement of two malectin-like domains connected by a short βhairpin linker and stabilized by calcium ions. The canonical carbohydrate interaction surfaces of related animal and bacterial carbohydrate binding modules are not conserved among plant CrRLK1Ls. In line with this, we failed to detect binding of chemically diverse oligosaccharides to ANXUR1 and HERCULES1. Instead, CrRLK1Ls have evolved a protein-protein interface between their malectin domains, which forms a deep cleft lined by highly conserved aromatic and polar residues. Analysis of the glycosylation pattern of different CrRLK1Ls and their oligomeric states together suggests that this cleft could resemble a binding site for a ligand required for Cr RLK1Ls receptor activation.