Roles of Plant Hormones and Their Interplay in Rice Immunity

ABSTRACT Plant hormones have been extensively studied for their importance in innate immunity particularly in the dicotyledonous model plant Arabidopsis thaliana. However, only in the last decade, plant hormones were demonstrated to play conserved and divergent roles in fine-tuning immune responses in rice （Oryza sativa L.）, a monocotyledonous model crop plant. Emerging evidence showed that salicylic acid （SA） plays a role in rice basal defense but is differentially required by rice pattern recognition receptor （PRR） and resistance （R） protein-mediated immunity, and its function is likely dependent on the signaling pathway rather than the change of endogenous levels. Jasmonate （JA） plays an important role in rice basal defense against bacterial and fungal infection and may be involved in the SA-mediated resistance. Ethylene （ET） can act as a positive or negative modulator of disease resistance, depending on the pathogen type and environmental conditions. Brassinosteroid （BR） signaling and abscisic acid （ABA） either promote or defend against infection of pathogens with distinct infection/colonization strategies. Auxin and gibberellin （GA） are generally thought of as negative regulators of innate immunity in rice. Moreover, GA interacts antagonistically with JA signaling in rice development and immunity through the DELLA protein as a master regulator of the two hormone pathways. In this review, we summarize the roles of plant hormones in rice immunity and discuss their interplay/crosstalk mechanisms and the complex regulatory network of plant hormone pathways in fine-tuning rice immunity and growth.

Diverse Transcriptional Programs Associated with Environmental Stress and Hormones in the Arabidopsis Receptor-Like Kinase Gene Family

The genome ofArabidopsis thaliana encodes more than 600 receptor-like kinase （RLK） genes, by far the dominant class of receptors found in land plants. Although similar to the mammalian receptor tyrosine kinases, plant RLKs are serine/threonine kinases that represent a novel signaling innovation unique to plants and, consequently, an excellent opportunity to understand how extracellular signaling evolved and functions in plants as opposed to animals. RLKs are predicted to be major components of the signaling pathways that allow plants to respond to environmental and developmental conditions. However, breakthroughs in identifying these processes have been limited to only a handful of individual RLKs. Here, we used a Syngenta custom Arabidopsis GeneChip array to compile a detailed profile of the tran- scriptional activity of 604 receptor-like kinase genes after exposure to a cross-section of known signaling factors in plants, including abiotic stresses, biotic stresses, and hormones. In the 68 experiments comprising the study, we found that 582 of the 604 RLK genes displayed a two-fold or greater change in expression to at least one of 12 types of treatments, thereby providing a large body of experimental evidence for targeted functional screens of individual RLK genes. We investigated whether particular subfamilies of RLK genes are responsive to specific types of signals and found that each subfamily displayed broad ranges of expression, as opposed to being targeted towards particular signal classes. Finally, by analyzing the divergence of sequence and gene expression among the RLK subfamilies, we present evidence as to the functional basis for the expansion of the RLKs and how this expansion may have affected conservation and divergences in their function. Taken as a whole, our study represents a preliminary, working model of processes and interactions in which the members of the RLK gene family may be involved, where such information has remained elusive for so many of its members.

Auxin Biosynthesis： A Simple Two-Step Pathway Converts Tryptophan to Indole-3-Acetic Acid in Plants

Indole-3-acetic acid （IAA）, the main naturally occurring auxin, is essential for almost every aspect of plant growth and development. However, only recently have studies finally established the first complete auxin biosynthesis pathway that converts tryptophan （Trp） to IAA in plants. Trp is first converted to indole-3-pyruvate （IPA） by the TAA family of amino transferases and subsequently IAA is produced from IPA by the YUC family of flavin monooxygenases. The two- step conversion of Trp to IAA is the main auxin biosynthesis pathway that plays an essential role in many developmental processes.

A DTX/MATE-Type Transporter Facilitates Abscisic Acid Efflux and Modulates ABA Sensitivity and Drought Tolerance in Arabidopsis

Abscisic acid （ABA） regulates numerous physiological and developmental processes in plants. Recent studies identify intracellular ABA receptors, implicating the transport of ABA across cell membranes as crucial for ABA sensing and response. Here, we report that a DTX/Multidrug and Toxic Compound Extrusion （MATE） family member in Arabidopsis thaliana, AtDTX50, functions as an ABA efflux transporter. When expressed heterologously in both an Escherichia coli strain and Xenopus oocyte cells, AtDTX50 was found to facilitate ABA efflux. Furthermore, dtx50 mutant mesophyll cells preloaded with ABA released less ABA compared with the wild-type （WT）. The AtDTX50 gene was expressed mainly in the vascular tissues and guard ceils and its expression was strongly up-regulated by exogenous ABA. The AtDTX50：：GFP fusion protein was localized predominantly to the plasma membrane. The dtx50 mutant plants were observed to be more sensitive to ABA in growth inhibition. In addition, compared with the WT, dtx50 mutant plants were more tolerant to drought with lower stomatal conductance, consistent with its function as an ABA efflux carrier in guard cells.

Is ABP1 an Auxin Receptor Yet？

AUXIN BINDING PROTEIN 1 （ABP1） has long been proposed as an auxin receptor to regulate cell expansion. The embryo lethality of ABP1-null mutants demonstrates its fundamental role in plant development, but also hinders investigation of its involvement in post-embryonic processes and its mode of action. By taking advantage of weak alleles and inducible systems, several recent studies have revealed a role for ABP1 in organ development, cell polarization, and shape formation. In addition to its role in the regulation of auxin-induced gene expression, ABP1 has now been shown to modulate non-transcriptional auxin responses. ABP1 is required for activating two antagonizing ROP GTPase signaling pathways involved in cytoskeletal reorganization and cell shape formation, and participates in the regulation of clathrinmediated endocytosis to subsequently affect PIN protein distribution. These exciting discoveries provide indisputable evidence for the auxin-induced signaling pathways that are downstream of ABP1 function, and suggest intriguing mechanisms for ABPl-mediated polar cell expansion and spatial coordination in response to auxin.

Abscisic Acid Signal off the STARTing Block

The year 2009 marked a real turnaround in our understanding of the mode of abscisic acid （ABA） action. Nearly 25 years had elapsed since the first biochemical detection of ABA-binding proteins in the plasmalemma of Vicia guard cells was reported. This recent--and laudable--achievement is owed largely to the discovery of the soluble ABA receptors whose major interacting proteins happen to be some of the most well-established components of earliest steps in ABA signaling. These soluble receptors, with the double name of PYRABACTIN RESISTANCE （PYR） or REGULATORY COMPONENT OF ABA RECEPTOR （RCAR）, are a family of Arabidopsis proteins of about 150-200 amino acids that share a conserved START domain. The ABA signal transduction circuitry under non-stress conditions is muted by the clade A protein phosphatases 2C （PP2C） （notably HAB1, ABI1, and ABI2）. During the initial steps of ABA signaling, the binding of the hormone to the receptor induces a conformational change in the latter that allows it to sequester the PP2Cs. This excludes them from the negative regulation of the downstream ABA-activated kinases （OST1/SnRK2.6/SRK2E, SnRK2.2, and SnRK2.3）, thus unleashing the pathway by freeing them to phosphorylate downstream targets that now include several b-ZIP transcription factors, ion channels （SLAC1, KAT1）, and a NADPH oxidase （AtrbohF）. The discovery of this family of soluble receptors and the rich insight already gained from structural studies of their complexes with different isoforms of ABA, PP2C, and the synthetic agonist pyrabactin lay the foundation towards rational design of chemical switches that can bolster drought hardiness in plants.

A Novel ABA Insensitive Mutant of Lotus japonicus with a Wilty Phenotype Displays Unaltered Nodulation Regulation

An ABA insensitive mutant, Beyma, was isolated in Lotus japonicus MG-20 from an EMS mutagenesis population using root growth inhibition to applied ABA as the screening criterion. （The name ＂Beyma＂ was taken from the Australian Aboriginal language, Wagiman, beyma, meaning ‘drying up＇.） The stable mutant that segregates as a dominant Mendelian mutation is insensitive to ABA induced inhibition of germination, vegetative growth, stomatal opening, as well as nodulation. Tissue ABA levels were normal, suggesting a sensitivity rather than biosynthesis mutation. It is slow-growing （50-70% of wild-type MG-20） and has a near-constitutive wilty phenotype associated with its inability to regulate stomatal opening. Whilst showing a wide range of ABA insensitive phenotypes, Beyma did not show alteration of nodule number control, as, in the absence of added ABA, the number and patterning （but not size） of nodules formed in the mutant were similar to that of MG-20. Split root experiments on MG-20 showed that application of ABA on one side of the root inhibited nodulation locally but not systemically. We propose that ABA is not involved directly in systemic autoregulation of nodulation （AON）.

Patterns and Timing in Expression of Early Auxin-Induced Genes Imply Involvement of Phospholipases A （pPLAs） in the Regulation of Auxin Responses

While it is known that patatin-related phospholipase A （pPLA） activity is rapidly activated within 3min by auxin, hardly anything is known about how this signal influences downstream responses like transcription of early auxin-induced genes or other physiological responses. We screened mutants with T-DNA insertions in members of the pPLA gene family for molecular and physiological phenotypes related to auxin. Only one in nine Arabidopsis thaliana ppla knockdown mutants displayed an obvious constitutive auxin-related phenotype. Compared to wild-type, ppla-IIlδ mutant seedlings had decreased main root lengths and increased lateral root numbers. We tested auxin-induced gene expression as a molecular readout for primary molecular auxin responses in nine ppla mutants and found delayed up- regulation of auxin-responsive gene exRression in all of themL Thirty minutes after auxin treatment, up-regulation of up to 40% of auxin-induced genes was delayed in mutant seedlings. We observed only a few cases with hypersensitive auxin-induced gene expression in ppla mutants. While, in three ppla mutants, which were investigated in detail, rapid up- regulation （as early as 10 min after auxin stimulus） of auxin-regulated genes was impaired, late transcriptional responses were wild-type-like. This regulatory or dynamic phenotype was consistently observed in different ppla mutants with delayed up-regulation that frequently affected the same genes. This defect was not affected by pPLA transcript levels which remained constant. This indicates aposttranslational mechanism as a functional link of pPLAs to auxin signaling. The need for a receptor triggering an auxin response without employing transcription regulation is discussed.

Prediction and Validation of Promoters Involved in the Abscisic Acid Response in Physcomitrella patens

Detection of cis-regulatory elements, such as transcription factor binding sites （TFBS）, through utilization of ortholog conservation is possible only if genomic data from closely related organisms are available. An alternative approach is the detection of TFBS based on their overrepresentation in promoters of co-regulated genes. However, this approach usually suffers from a high rate of false-positive prediction. Here, we have conducted a case study using promoters of genes known to be strongly induced by the phytohormone abscisic acid （ABA） in the model plant Physcomitrella patens, a moss. Putative TFBS were detected using three de novo motif detection tools in a strict consensus approach. The resulting motifs were validated using data from microarray expression profiling and were able to predict ABA-induced genes with high specificity （90.48%） at mediocre sensitivity （33.33%）. In addition, 27 genes predicted to contain ABA-responsive TFBS were validated using real-time PCR. Here, a total of 37% of the genes could be shown to be induced upon ABA treatment, while 70% were found to be regulated by ABA. We conclude that the consensus approach for motif detection using coregulation information can be used to identify genes that are regulated under a given stimulus. In terms of evolution, we find that the ABA response has apparently been conserved since the first land plants on the level of families involved in transcriptional regulation.

Long-Distance Signaling in bypass1 Mutants： Bioassay Development Reveals the bps Signal to Be a Metabolite

Root-to-shoot signaling is used by plants to coordinate shoot development with the conditions experienced by the roots. A mobile and biologically active compound, the bps signal, is over-produced in roots of an Arabidopsis thaliana mutant called bypass1 （bpsl）, and might also be a normally produced signaling molecule in wild-type plants. Our goal is to identify the bps signal chemically, which will then allow us to assess its production in normal plants. To identify any signaling molecule, a bioassay is required, and here we describe the development of a robust, simple, and quantitative bioassay for the bps signal. The developed bioassay follows the growth-reducing activity of the bps signal using the pCYCB1;I：：GUS cell cycle marker. Wild-type plants carrying this marker, and provided the bps signal through either grafts or metabolite extracts, showed reduced cell division. By contrast, control grafts and treatment with control extracts showed no change in pCYCB1;I：：GUS expression. To determine the chemical nature of the bps signal, extracts were treated with RNase A, Proteinase K, or heat. None of these treatments diminished the activity of bpsl extracts, sug- gesting that the active molecule might be a metabolite. This bioassay will be useful for future biochemical fractionation and analysis directed toward bps signal identification.

Farnesylcysteine Lyase Regulation of Abscisic Arabidopsis is Involved in Negative Acid Signaling in

The Arabidopsis FCLY gene encodes a specific farnesylcysteine （FC） lyase, which is responsible for the oxidative metabolism of FC to farnesal and cysteine. In addition, fcly mutants with quantitative decreases in FC lyase activity exhibit an enhanced response to ABA. However, the enzymological properties of the FCLY-encoded enzyme and its precise role in ABA signaling remain unclear. Here, we show that recombinant Arabidopsis FC lyase expressed in insect cells exhib- its high selectivity for FC as a substrate and requires FAD and molecular oxygen for activity. Arabidopsis FC lyase is also shown to undergo post-translational N-glycosylation. FC, which is a competitive inhibitor of isoprenylcysteine methyltransferase （ICMT）, accumulates in fcly mutants. Moreover, the enhanced response of fcly mutants to ABA is reversed by ICMToverexpression. These observations support the hypothesis that the ABA hypersensitive phenotype of fcly plants is the result of FC accumulation and inhibition of ICMT.