FERONIA: A Receptor Kinase at the Core of a Global Signaling Network.

Initially identified as a key regulator of female fertility in Arabidopsis, the FERONIA (FER) receptor kinase is now recognized as crucial for almost all aspects of plant growth and survival. FER partners with a glycosylphosphatidylinositol-anchored protein of the LLG family to act as coreceptors on the cell surface. The FER-LLG coreceptor interacts with different RAPID ALKALINIZATION FACTOR (RALF) peptide ligands to function in various growth and developmental processes and to respond to challenges from the environment. The RALF-FER-LLG signaling modules interact with molecules in the cell wall, cell membrane, cytoplasm, and nucleus and mediate an interwoven signaling network. Multiple FER-LLG modules, each anchored by FER or a FER-related receptor kinase, have been studied, illustrating the functional diversity and the mechanistic complexity of the FER family signaling modules. The challenges going forward are to distill from this complexity the unifying schemes where possible and attain precision and refinement in the knowledge of critical details upon which future investigations can be built. By focusing on the extensively characterized FER, this review provides foundational information to guide the next phase of research on FER in model as well as crop species and potential applications for improving plant growth and resilience.

Extracellular pectin-RALF phase separation mediates FERONIA global signaling function.

The FERONIA (FER)-LLG1 co-receptor and its peptide ligand RALF regulate myriad processes for plant growth and survival. Focusing on signal-induced cell surface responses, we discovered that intrinsically disordered RALF triggers clustering and endocytosis of its cognate receptors and FER- and LLG1-dependent endocytosis of non-cognate regulators of diverse processes, thus capable of broadly impacting downstream responses. RALF, however, remains extracellular. We demonstrate that RALF binds the cell wall polysaccharide pectin. They phase separate and recruit FER and LLG1 into pectin-RALF-FER-LLG1 condensates to initiate RALF-triggered cell surface responses. We show further that two frequently encountered environmental challenges, elevated salt and temperature, trigger RALF-pectin phase separation, promiscuous receptor clustering and massive endocytosis, and that this process is crucial for recovery from stress-induced growth attenuation. Our results support that RALF-pectin phase separation mediates an exoskeletal mechanism to broadly activate FER-LLG1-dependent cell surface responses to mediate the global role of FER in plant growth and survival.

Brassinosteroid recruits FERONIA to safeguard cell expansion in Arabidopsis.

Plant cell expansion is driven by turgor pressure and regulated by hormones. How plant cells avoid cell wall rupture during hormone-induced cell expansion remains a mystery. Here we show that brassinosteroid (BR), while stimulating cell elongation, promotes the plasma membrane (PM) accumulation of the receptor kinase FERONIA (FER), which monitors cell wall damage and in turn attenuates BR-induced cell elongation to prevent cell rupture. The GSK3-like kinase BIN2 phosphorylates FER, resulting in reduced FER accumulation and translocation from endoplasmic reticulum to PM. By inactivating BIN2, BR signaling promotes dephosphorylation and increases PM accumulation of FER, thereby enhancing the surveillance of cell wall integrity. Our study reveals a vital signaling circuit that coordinates hormone signaling with mechanical sensing to prevent cell bursting during hormone-induced cell expansion.

Using Hyper as a molecular probe to visualize hydrogen peroxide in living plant cells: An updated method.

Reactive oxygen species (ROS) are highly reactive reduced oxygen molecules that play a myriad of roles in animal and plant cells. In plant cells the production of ROS results from aerobic metabolism during respiration and photosynthesis. Therefore mitochondria, chloroplasts, and peroxisomes constitute an important source of ROS. However, ROS can also be produced in response to many physiological stimuli such as pathogen attack, hormone signaling, abiotic stresses or during cell wall organization and plant morphogenesis. The study of ROS in plant cells has been limited to biochemical assays and use of fluorescent probes, however, the irreversible oxidation of the fluorescent dyes prevents the visualization of dynamic changes. We have previously reported that Hyper 1 is a biosensor for H2O2 and consists of a circularly permutated YFP (cpYFP) inserted into the regulatory domain of the Escherichia coli hydrogen peroxide (H2O2) sensor protein OxyR rendering it an H2O2-specific quantitative probe (Bilan & Belousov, 2018; Hernandez-Barrera et al., 2015). Herein we describe an updated protocol for using the improved new version of Hyper 2 and Hyper 3 as a dynamic biosensor for H2O2 in Arabidopsis with virtually unlimited potential to detect H2O2 throughout the plant and under a broad range of developmental and environmental conditions (Bilan et al., 2013).

Stigma receptors control intraspecies and interspecies barriers in Brassicaceae.

Flowering plants have evolved numerous intraspecific and interspecific prezygotic reproductive barriers to prevent production of unfavourable offspring1. Within a species, self-incompatibility (SI) is a widely utilized mechanism that rejects self-pollen2,3 to avoid inbreeding depression. Interspecific barriers restrain breeding between species and often follow the SI × self-compatible (SC) rule, that is, interspecific pollen is unilaterally incompatible (UI) on SI pistils but unilaterally compatible (UC) on SC pistils1,4-6. The molecular mechanisms underlying SI, UI, SC and UC and their interconnections in the Brassicaceae remain unclear. Here we demonstrate that the SI pollen determinant S-locus cysteine-rich protein/S-locus protein 11 (SCR/SP11)2,3 or a signal from UI pollen binds to the SI female determinant S-locus receptor kinase (SRK)2,3, recruits FERONIA (FER)7-9 and activates FER-mediated reactive oxygen species production in SI stigmas10,11 to reject incompatible pollen. For compatible responses, diverged pollen coat protein B-class12-14 from SC and UC pollen differentially trigger nitric oxide, nitrosate FER to suppress reactive oxygen species in SC stigmas to facilitate pollen growth in an intraspecies-preferential manner, maintaining species integrity. Our results show that SRK and FER integrate mechanisms underlying intraspecific and interspecific barriers and offer paths to achieve distant breeding in Brassicaceae crops.

The RAC/ROP GTPase activator OsRopGEF10 functions in crown root development by regulating cytokinin signaling in rice.

RAC/Rho of plant (ROP) GTPases are major molecular switches that control diverse signaling cascades for plant growth, development, and defense. Here, we discovered a signaling node that connects RAC/ROPs to cytokinins. Rice (Oryza sativa) plants develop a fibrous root system mainly composed of crown roots. Cytokinin signaling via a phosphorelay system is critical for crown root development. We show that OsRopGEF10, which activates RAC/ROPs, acts upstream of the cytoplasmic-nuclear shuttling phosphotransfer proteins AHPs of the cytokinin signaling pathway to promote crown root development. Mutations of OsRopGEF10 induced hypersensitivity to cytokinin, whereas overexpressing this gene reduced the cytokinin response. Loss of OsRopGEF10 function reduced the expression of the response regulator gene OsRR6, a repressor of cytokinin signaling, and impaired crown root development. Mutations in OsAHP1/2 led to increased crown root production and rescued the crown root defect of Osropgef10. Furthermore, auxin activates the ROP GTPase OsRAC3, which attenuates cytokinin signaling for crown root initiation. Molecular interactions between OsRopGEF10, OsRAC3, and OsAHP1/2 implicate a mechanism whereby OsRopGEF10-activated OsRAC3 recruits OsAHP1/2 to the cortical cytoplasm, sequestering them from their phosphorelay function in the nucleus. Together, our findings uncover the OsRopGEF10-OsRAC3-OsAHP1/2 signaling module, establish a link between RAC/ROPs and cytokinin, and reveal molecular crosstalk between auxin and cytokinin during crown root development.

Evolutionary analysis of the LORELEI gene family in plants reveals regulatory subfunctionalization.

A signaling complex comprising members of the LORELEI (LRE)-LIKE GPI-anchored protein (LLG) and Catharanthus roseus RECEPTOR-LIKE KINASE 1-LIKE (CrRLK1L) families perceive RAPID ALKALINIZATION FACTOR (RALF) peptides and regulate growth, reproduction, immunity, and stress responses in Arabidopsis (Arabidopsis thaliana). Genes encoding these proteins are members of multigene families in most angiosperms and could generate thousands of signaling complex variants. However, the links between expansion of these gene families and the functional diversification of this critical signaling complex as well as the evolutionary factors underlying the maintenance of gene duplicates remain unknown. Here, we investigated LLG gene family evolution by sampling land plant genomes and explored the function and expression of angiosperm LLGs. We found that LLG diversity within major land plant lineages is primarily due to lineage-specific duplication events, and that these duplications occurred both early in the history of these lineages and more recently. Our complementation and expression analyses showed that expression divergence (i.e. regulatory subfunctionalization), rather than functional divergence, explains the retention of LLG paralogs. Interestingly, all but one monocot and all eudicot species examined had an LLG copy with preferential expression in male reproductive tissues, while the other duplicate copies showed highest levels of expression in female or vegetative tissues. The single LLG copy in Amborella trichopoda is expressed vastly higher in male compared to in female reproductive or vegetative tissues. We propose that expression divergence plays an important role in retention of LLG duplicates in angiosperms.

Pollen-pistil interactions: It takes two to tangle but a molecular cast of many to deliver.

Explosive advances have been made in the molecular understanding of pollen-pistil interactions that underlie reproductive success in flowering plants in the past three decades. Among the most notable is the discovery of pollen tube attractants [1∗,2∗]. The roles these molecules play in facilitating conspecific precedence thus promoting interspecific genetic isolation are also emerging [3-5]. Male-female interactions during the prezygotic phase and contributions from the male and female gametophytes have been comprehensively reviewed recently. Here, we focus on key advances in understanding the mechanistic underpinnings of how these interactions overcome barriers at various pollen-pistil interfaces along the pollen tube growth pathway to facilitate fertilization by desirable mates.

RALF peptide signaling controls the polytubey block in Arabidopsis.

Fertilization of an egg by multiple sperm (polyspermy) leads to lethal genome imbalance and chromosome segregation defects. In Arabidopsis thaliana, the block to polyspermy is facilitated by a mechanism that prevents polytubey (the arrival of multiple pollen tubes to one ovule). We show here that FERONIA, ANJEA, and HERCULES RECEPTOR KINASE 1 receptor-like kinases located at the septum interact with pollen tube-specific RALF6, 7, 16, 36, and 37 peptide ligands to establish this polytubey block. The same combination of RALF (rapid alkalinization factor) peptides and receptor complexes controls pollen tube reception and rupture inside the targeted ovule. Pollen tube rupture releases the polytubey block at the septum, which allows the emergence of secondary pollen tubes upon fertilization failure. Thus, orchestrated steps in the fertilization process in Arabidopsis are coordinated by the same signaling components to guarantee and optimize reproductive success.

A rich and bountiful harvest: Key discoveries in plant cell biology.

The field of plant cell biology has a rich history of discovery, going back to Robert Hooke's discovery of cells themselves. The development of microscopes and preparation techniques has allowed for the visualization of subcellular structures, and the use of protein biochemistry, genetics, and molecular biology has enabled the identification of proteins and mechanisms that regulate key cellular processes. In this review, seven senior plant cell biologists reflect on the development of this research field in the past decades, including the foundational contributions that their teams have made to our rich, current insights into cell biology. Topics covered include signaling and cell morphogenesis, membrane trafficking, cytokinesis, cytoskeletal regulation, and cell wall biology. In addition, these scientists illustrate the pathways to discovery in this exciting research field.

Malectin/Malectin-like domain-containing proteins: A repertoire of cell surface molecules with broad functional potential.

Cell walls are at the front line of interactions between walled-organisms and their environment. They support cell expansion, ensure cell integrity and, for multicellular organisms such as plants, they provide cell adherence, support cell shape morphogenesis and mediate cell-cell communication. Wall-sensing, detecting perturbations in the wall and signaling the cell to respond accordingly, is crucial for growth and survival. In recent years, plant signaling research has suggested that a large family of receptor-like kinases (RLKs) could function as wall sensors partly because their extracellular domains show homology with malectin, a diglucose binding protein from the endoplasmic reticulum of animal cells. Studies of several malectin/malectin-like (M/ML) domain-containing RLKs (M/MLD-RLKs) from the model plant Arabidopsis thaliana have revealed an impressive array of biological roles, controlling growth, reproduction and stress responses, processes that in various ways rely on or affect the cell wall. Malectin homologous sequences are widespread across biological kingdoms, but plants have uniquely evolved a highly expanded family of proteins with ML domains embedded within various protein contexts. Here, we present an overview on proteins with malectin homologous sequences in different kingdoms, discuss the chromosomal organization of Arabidopsis M/MLD-RLKs and the phylogenetic relationship between these proteins from several model and crop species. We also discuss briefly the molecular networks that enable the diverse biological roles served by M/MLD-RLKs studied thus far.

FERONIA receptor kinase-regulated reactive oxygen species mediate self-incompatibility in Brassica rapa.

Most plants in the Brassicaceae evolve self-incompatibility (SI) to avoid inbreeding and generate hybrid vigor. Self-pollen is recognized by the S-haplotype-specific interaction of the pollen ligand S-locus protein 11 (SP11) (also known as S-locus cysteine-rich protein [SCR]) and its stigma-specific S-locus receptor kinase (SRK). However, mechanistically much remains unknown about the signaling events that culminate in self-pollen rejection. Here, we show that self-pollen triggers high levels of reactive oxygen species (ROS) in stigma papilla cells to mediate SI in heading Chinese cabbage (Brassica rapa L. ssp. pekinensis). We found that stigmatic ROS increased after self-pollination but decreased after compatible(CP)- pollination. Reducing stigmatic ROS by scavengers or suppressing the expression of respiratory burst oxidase homologs (Rbohs), which encode plant NADPH oxidases that produce ROS, both broke down SI. On the other hand, increasing the level of ROS inhibited the germination and penetration of compatible pollen on the stigma, mimicking an incompatible response. Furthermore, suppressing a B. rapa FERONIA (FER) receptor kinase homolog or Rac/Rop guanosine triphosphatase (GTPase) signaling effectively reduced stigmatic ROS and interfered with SI. Our results suggest that FER-Rac/Rop signaling-regulated, NADPH oxidase-produced ROS is an essential SI response leading to self-pollen rejection.

Pollen PCP-B peptides unlock a stigma peptide-receptor kinase gating mechanism for pollination.

Sexual reproduction in angiosperms relies on precise communications between the pollen and pistil. The molecular mechanisms underlying these communications remain elusive. We established that in Arabidopsis, a stigmatic gatekeeper, the ANJEA-FERONIA (ANJ-FER) receptor kinase complex, perceives the RAPID ALKALINIZATION FACTOR peptides RALF23 and RALF33 to induce reactive oxygen species (ROS) production in the stigma papillae, whereas pollination reduces stigmatic ROS, allowing pollen hydration. Upon pollination, the POLLEN COAT PROTEIN B-class peptides (PCP-Bs) compete with RALF23/33 for binding to the ANJ-FER complex, leading to a decline of stigmatic ROS that facilitates pollen hydration. Our results elucidate a molecular gating mechanism in which distinct peptide classes from pollen compete with stigma peptides for interaction with a stigmatic receptor kinase complex, allowing the pollen to hydrate and germinate.

Auxin efflux controls orderly nucellar degeneration and expansion of the female gametophyte in Arabidopsis.

The nucellus tissue in flowering plants provides nutrition for the development of the female gametophyte (FG) and young embryo. The nucellus degenerates as the FG develops, but the mechanism controlling the coupled process of nucellar degeneration and FG expansion remains largely unknown. The degeneration process of the nucellus and spatiotemporal auxin distribution in the developing ovule before fertilization were investigated in Arabidopsis thaliana. Nucellar degeneration before fertilization occurs through vacuolar cell death and in an ordered degeneration fashion. This sequential nucellar degeneration is controlled by the signalling molecule auxin. Auxin efflux plays the core role in precisely controlling the spatiotemporal pattern of auxin distribution in the nucellus surrounding the FG. The auxin efflux carrier PIN1 transports maternal auxin into the nucellus while PIN3/PIN4/PIN7 further delivers auxin to degenerating nucellar cells and concurrently controls FG central vacuole expansion. Notably, auxin concentration and auxin efflux are controlled by the maternal tissues, acting as a key communication from maternal to filial tissue.

Programmed Cell Death in Stigmatic Papilla Cells Is Associated With Senescence-Induced Self-Incompatibility Breakdown in Chinese Cabbage and Radish.

Self-incompatibility (SI) is a genetic mechanism flowering plants adopted to reject self-pollen and promote outcrossing. In the Brassicaceae family plants, the stigma tissue plays a key role in self-pollen recognition and rejection. We reported earlier in Chinese cabbage (Brassica rapa) that stigma tissue showed upregulated ethylene responses and programmed cell death (PCD) upon compatible pollination, but not in SI responses. Here, we show that SI is significantly compromised or completely lost in senescent flowers and young flowers of senescent plants. Senescence upregulates senescence-associated genes in B. rapa. Suppressing their expression in young stigmas by antisense oligodeoxyribonucleotide abolishes compatible pollination-triggered PCD and inhibits the growth of compatible pollen tubes. Furthermore, ethylene biosynthesis genes and response genes are upregulated in senescent stigmas, and increasing the level of ethylene or inhibiting its response increases or decreases the expression of senescence-associated genes, respectively. Our results show that senescence causes PCD in stigmatic papilla cells and is associated with the breakdown of SI in Chinese cabbage and in radish.