Antagonistic regulation of the gibberellic acid response during stem growth in rice.

The size of plants is largely determined by growth of the stem. Stem elongation is stimulated by gibberellic acid1-3. Here we show that internode stem elongation in rice is regulated antagonistically by an 'accelerator' and a 'decelerator' in concert with gibberellic acid. Expression of a gene we name ACCELERATOR OF INTERNODE ELONGATION 1 (ACE1), which encodes a protein of unknown function, confers cells of the intercalary meristematic region with the competence for cell division, leading to internode elongation in the presence of gibberellic acid. By contrast, upregulation of DECELERATOR OF INTERNODE ELONGATION 1 (DEC1), which encodes a zinc-finger transcription factor, suppresses internode elongation, whereas downregulation of DEC1 allows internode elongation. We also show that the mechanism of internode elongation that is mediated by ACE1 and DEC1 is conserved in the Gramineae family. Furthermore, an analysis of genetic diversity suggests that mutations in ACE1 and DEC1 have historically contributed to the selection of shorter plants in domesticated populations of rice to increase their resistance to lodging, and of taller plants in wild species of rice for adaptation to growth in deep water. Our identification of these antagonistic regulatory factors enhances our understanding of the gibberellic acid response as an additional mechanism that regulates internode elongation and environmental fitness, beyond biosynthesis and gibberellic acid signal transduction.

Regulator of Awn Elongation 3, an E3 ubiquitin ligase, is responsible for loss of awns during African rice domestication.

Two species of rice have been independently domesticated from different ancestral wild species in Asia and Africa. Comparison of mutations that underlie phenotypic and physiological alterations associated with domestication traits in these species gives insights into the domestication history of rice in both regions. Asian cultivated rice, Oryza sativa, and African cultivated rice, Oryza glaberrima, have been modified and improved for common traits beneficial for humans, including erect plant architecture, nonshattering seeds, nonpigmented pericarp, and lack of awns. Independent mutations in orthologous genes associated with these traits have been documented in the two cultivated species. Contrary to this prevailing model, selection for awnlessness targeted different genes in O. sativa and O. glaberrima. We identify Regulator of Awn Elongation 3 (RAE3) a gene that encodes an E3 ubiquitin ligase and is responsible for the awnless phenotype only in O. glaberrima. A 48-bp deletion may disrupt the substrate recognition domain in RAE3 and diminish awn elongation. Sequencing analysis demonstrated low nucleotide diversity in a ~600-kb region around the derived rae3 allele on chromosome 6 in O. glaberrima compared with its wild progenitor. Identification of RAE3 sheds light on the molecular mechanism underlying awn development and provides an example of how selection on different genes can confer the same domestication phenotype in Asian and African rice.

Molecular mechanism of internode elongation in rice.

Rice plants that form ventilated tissues, such as aerenchyma in the leaves, stems, and roots, allow for growth in waterlogged conditions (paddy fields), but they cannot breathe and drown in flooded environments where the whole plant body is submerged. However, deepwater rice plants grown in flood-prone areas of Southeast Asia survive in prolonged flooded environments by taking in air through an elongated stem (internode) and leaves that emerge above the water surface, even if the water level is several meters high and flooding continues for several months. Although it has been known that plant hormones, such as ethylene and gibberellins, promote internode elongation in deepwater rice plants, the genes that control rapid internode elongation during submergence have not been identified. We recently identified several genes responsible for the quantitative trait loci involved in internode elongation in deepwater rice. Identification of the the genes revealed a molecular gene network from ethylene to gibberellins in which internode elongation is promoted by novel ethylene-responsive factors and enhances gibberellin responsiveness at the internode. In addition, elucidation of the molecular mechanism of internode elongation in deepwater rice will help our understanding of the internode elongation mechanism in normal paddy rice and contribute to improving crops through the regulation of internode elongation.

Designing rice panicle architecture via developmental regulatory genes.

Rice panicle architecture displays remarkable diversity in branch number, branch length, and grain arrangement; however, much remains unknown about how such diversity in patterns is generated. Although several genes related to panicle branch number and panicle length have been identified, how panicle branch number and panicle length are coordinately regulated is unclear. Here, we show that panicle length and panicle branch number are independently regulated by the genes Prl5/OsGA20ox4, Pbl6/APO1, and Gn1a/OsCKX2. We produced near-isogenic lines (NILs) in the Koshihikari genetic background harboring the elite alleles for Prl5, regulating panicle rachis length; Pbl6, regulating primary branch length; and Gn1a, regulating panicle branching in various combinations. A pyramiding line carrying Prl5, Pbl6, and Gn1a showed increased panicle length and branching without any trade-off relationship between branch length or number. We successfully produced various arrangement patterns of grains by changing the combination of alleles at these three loci. Improvement of panicle architecture raised yield without associated negative effects on yield-related traits except for panicle number. Three-dimensional (3D) analyses by X-ray computed tomography (CT) of panicles revealed that differences in panicle architecture affect grain filling. Importantly, we determined that Prl5 improves grain filling without affecting grain number.

Fine mapping and characterization of two novel quantitative trait loci for early seedling growth in rice.

MAIN CONCLUSION: Two novel QTLs for early seedling growth in rice were fine mapped, with one of which to a 4-kb identical to the known GW6a gene, and another one to a 43-kb region that contains six candidate genes. Leaves are extremely important for plant photosynthesis: the size and shape of which determine the rate of transpiration, carbon fixation and light interception, and their robust growth at seedling stage endow crops with the ability to compete with weeds. So far, many genes for the traits have been cloned with mutants; however, identification of those quantitative trait loci (QTLs) that control early seedling growth has seldom been reported. In this study, we report the identification of two QTLs, qLBL1 and qLBL2 on the rice chromosome 6 for leaf blade length at early seedling stage. Fine mapping revealed that qLBL1 was placed into a 4-kb, and qLBL2 was delimited to a 43-kb genomic interval. We further found that LBL1 was equivalent to the known grain-size gene GW6a and the qLBL2 region contains 6 candidate genes. Genetic analysis using nearly isogenic lines and transgenic rice plants revealed that both genetic factors were positive regulators. The genetic effects were mainly due to alterations of cell division by cytological observations. RT-qPCR results showed that LBL1 was preferentially expressed in leaf blades, and consistently, histochemical staining of pGW6a::GUS plants showed that GUS signal was strong in the vascular tissues of leaf blade of seedlings. Thus, we fine mapped and characterized two QTLs for early seedling growth and provided useful information to improve crop breeding.

Noninvasive imaging of hollow structures and gas movement revealed the gas partial-pressure-gradient-driven long-distance gas movement in the aerenchyma along the leaf blade to submerged organs in rice.

Rice (Oryza sativa) plants have porous or hollow organs consisting of aerenchyma, which is presumed to function as a low-resistance diffusion pathway for air to travel from the foliage above the water to submerged organs. However, gas movement in rice plants has yet to be visualized in real time. In this study involving partially submerged rice plants, the leaves emerging from the water were fed nitrogen-13-labeled nitrogen ([13 N]N2 ) tracer gas, and the gas movement downward along the leaf blade, leaf sheath, and internode over time was monitored. The [13 N]N2 gas arrived at the bottom of the plant within 10 min, which was 20 min earlier than carbon-11 photoassimilates. The [13 N]N2 gas movement was presumably mediated by diffusion along the aerenchyma network from the leaf blade to the root via nodes functioning as junctions, which were detected by X-ray computed tomography. These findings imply the diffusion of gas along the aerenchyma, which does not consume energy, has enabled plants to adapt to aquatic environments. Additionally, there were no major differences in [13 N]N2 gas movement between paddy rice and deepwater rice plants, indicative of a common aeration mechanism in the two varieties, despite the difference in their response to flooding.

Diel O2 Dynamics in Partially and Completely Submerged Deepwater Rice: Leaf Gas Films Enhance Internodal O2 Status, Influence Gene Expression and Accelerate Stem Elongation for 'Snorkelling' during Submergence.

Deepwater rice has a remarkable shoot elongation response to partial submergence. Shoot elongation to maintain air-contact enables 'snorkelling' of O2 to submerged organs. Previous research has focused on partial submergence of deepwater rice. We tested the hypothesis that leaf gas films enhance internode O2 status and stem elongation of deepwater rice when completely submerged. Diel patterns of O2 partial pressure (pO2) were measured in internodes of deepwater rice when partially or completely submerged, and with or without gas films on leaves, for the completely submerged plants. We also took measurements for paddy rice. Deepwater rice elongated during complete submergence and the shoot tops emerged. Leaf gas films improved O2 entry during the night, preventing anoxia in stems, which is of importance for elongation of the submerged shoots. Expressions of O2 deprivation inducible genes were upregulated in completely submerged plants during the night, and more so when gas films were removed from the leaves. Diel O2 dynamics showed similar patterns in paddy and deepwater rice. We demonstrated that shoot tops in air enabled 'snorkelling' and increased O2 in internodes of both rice ecotypes; however, 'snorkelling' was achieved only by rapid shoot elongation by deepwater rice, but not by paddy rice.

Time-Course Transcriptomics Analysis Reveals Key Responses of Submerged Deepwater Rice to Flooding.

Water submergence is an environmental factor that limits plant growth and survival. Deepwater rice (Oryza sativa) adapts to submergence by rapidly elongating its internodes and thereby maintaining its leaves above the water surface. We performed a comparative RNA sequencing transcriptome analysis of the shoot base region, including basal nodes, internodes, and shoot apices of seedlings at two developmental stages from two varieties with contrasting deepwater growth responses. A transcriptomic comparison between deepwater rice cv C9285 and nondeepwater rice cv Taichung 65 revealed both similar and differential expression patterns between the two genotypes during submergence. The expression of genes related to gibberellin biosynthesis, trehalose biosynthesis, anaerobic fermentation, cell wall modification, and transcription factors that include ethylene-responsive factors was significantly different between the varieties. Interestingly, in both varieties, the jasmonic acid content at the shoot base decreased during submergence, while exogenous jasmonic acid inhibited submergence-induced internode elongation in cv C9285, suggesting that jasmonic acid plays a role in the submergence response of rice. Furthermore, a targeted de novo transcript assembly revealed transcripts that were specific to cv C9285, including submergence-induced biotic stress-related genes. Our multifaceted transcriptome approach using the rice shoot base region illustrates a differential response to submergence between deepwater and nondeepwater rice. Jasmonic acid metabolism appears to participate in the submergence-mediated internode elongation response of deepwater rice.

Correction to: Sucrose affects the developmental transition of rhizomes in Oryza longistaminata.

The article Sucrose affects the developmental transition of rhizomes in Oryza longistaminata, written by Kanako Bessho-Uehara, Jovano Erris Nugroho, Hirono Kondo, Rosalyn B. Angeles-Shim, Motoyuki Ashikari, was originally published electronically on the publisher's internet portal (currently SpringerLink) on 8 May 2018 without open access.

Loss of function at RAE2, a previously unidentified EPFL, is required for awnlessness in cultivated Asian rice.

Domestication of crops based on artificial selection has contributed numerous beneficial traits for agriculture. Wild characteristics such as red pericarp and seed shattering were lost in both Asian (Oryza sativa) and African (Oryza glaberrima) cultivated rice species as a result of human selection on common genes. Awnedness, in contrast, is a trait that has been lost in both cultivated species due to selection on different sets of genes. In a previous report, we revealed that at least three loci regulate awn development in rice; however, the molecular mechanism underlying awnlessness remains unknown. Here we isolate and characterize a previously unidentified EPIDERMAL PATTERNING FACTOR-LIKE (EPFL) family member named REGULATOR OF AWN ELONGATION 2 (RAE2) and identify one of its requisite processing enzymes, SUBTILISIN-LIKE PROTEASE 1 (SLP1). The RAE2 precursor is specifically cleaved by SLP1 in the rice spikelet, where the mature RAE2 peptide subsequently induces awn elongation. Analysis of RAE2 sequence diversity identified a highly variable GC-rich region harboring multiple independent mutations underlying protein-length variation that disrupt the function of the RAE2 protein and condition the awnless phenotype in Asian rice. Cultivated African rice, on the other hand, retained the functional RAE2 allele despite its awnless phenotype. Our findings illuminate the molecular function of RAE2 in awn development and shed light on the independent domestication histories of Asian and African cultivated rice.

Rice leaf hydrophobicity and gas films are conferred by a wax synthesis gene (LGF1) and contribute to flood tolerance.

Floods impede gas (O2 and CO2 ) exchange between plants and the environment. A mechanism to enhance plant gas exchange under water comprises gas films on hydrophobic leaves, but the genetic regulation of this mechanism is unknown. We used a rice mutant (dripping wet leaf 7, drp7) which does not retain gas films on leaves, and its wild-type (Kinmaze), in gene discovery for this trait. Gene complementation was tested in transgenic lines. Functional properties of leaves as related to gas film retention and underwater photosynthesis were evaluated. Leaf Gas Film 1 (LGF1) was identified as the gene determining leaf gas films. LGF1 regulates C30 primary alcohol synthesis, which is necessary for abundant epicuticular wax platelets, leaf hydrophobicity and gas films on submerged leaves. This trait enhanced underwater photosynthesis 8.2-fold and contributes to submergence tolerance. Gene function was verified by a complementation test of LGF1 expressed in the drp7 mutant background, which restored C30 primary alcohol synthesis, wax platelet abundance, leaf hydrophobicity, gas film retention, and underwater photosynthesis. The discovery of LGF1 provides an opportunity to better understand variation amongst rice genotypes for gas film retention ability and to target various alleles in breeding for improved submergence tolerance for yield stability in flood-prone areas.

Sucrose affects the developmental transition of rhizomes in Oryza longistaminata.

Oryza longistaminata, the African wild rice, can propagate vegetatively through rhizomes. Rhizomes elongate horizontally underground as sink organs, however, they undergo a developmental transition that shifts their growth to the surface of the ground to become aerial stems. This particular stage is essential for the establishment of new ramets. While several determinants such as abiotic stimuli and plant hormones have been reported as key factors effecting developmental transition in aerial stem, the cause of this phenomenon in rhizome remains elusive. This study shows that depletion of nutrients, particularly sucrose, is the key stimulus that induces the developmental transition in rhizomes, as indicated by the gradient of sugars from the base to the tip of the rhizome. Sugar treatments revealed that sucrose specifically represses the developmental transition from rhizome to aerial stem by inhibiting the expression of sugar metabolism and hormone synthesis genes at the bending point. Sucrose depletion affected several factors contributing to the developmental transition of rhizome including signal transduction, transcriptional regulation and plant hormone balance.

Rare allele of a previously unidentified histone H4 acetyltransferase enhances grain weight, yield, and plant biomass in rice.

Grain weight is an important crop yield component; however, its underlying regulatory mechanisms are largely unknown. Here, we identify a grain-weight quantitative trait locus (QTL) encoding a new-type GNAT-like protein that harbors intrinsic histone acetyltransferase activity (OsglHAT1). Our genetic and molecular evidences pinpointed the QTL-OsglHAT1's allelic variations to a 1.2-kb region upstream of the gene body, which is consistent with its function as a positive regulator of the traits. Elevated OsglHAT1 expression enhances grain weight and yield by enlarging spikelet hulls via increasing cell number and accelerating grain filling, and increases global acetylation levels of histone H4. OsglHAT1 localizes to the nucleus, where it likely functions through the regulation of transcription. Despite its positive agronomical effects on grain weight, yield, and plant biomass, the rare allele elevating OsglHAT1 expression has so far escaped human selection. Our findings reveal the first example, to our knowledge, of a QTL for a yield component trait being due to a chromatin modifier that has the potential to improve crop high-yield breeding.

Hormone Distribution and Transcriptome Profiles in Bamboo Shoots Provide Insights on Bamboo Stem Emergence and Growth.

Growth and development are tightly co-ordinated events in the lifetime of living organisms. In temperate bamboo plants, spring is the season when environmental conditions are suitable for the emergence of new shoots. Previous studies demonstrated that bamboo plants undergo an energy-consuming 'fast stem growth' phase. However, the events during the initiation of stem elongation in bamboo are poorly understood. To understand the onset of bamboo stem growth, we performed hormone and transcriptome profiling of tissue regions in newly elongating shoots of the Moso bamboo Phyllostachys edulis. The growth hormones auxins, cytokinins and gibberellins accumulated in the shoot apex, while the stress hormones ABA, salicylic acid (SA) and jasmonic acid (JA) are predominantly found in the lower part of the stem. The mature basal part of the stem showed enrichment of transcripts associated with cell wall metabolism and biosynthesis of phenylpropanoid metabolites, such as lignin. In the young upper stem region, expression of cell formation- and DNA synthesis-related genes was enriched. Moreover, the apical region showed enhanced expression of genes involved in meristem maintenance, leaf differentiation and development, abaxial/adaxial polarity and flowering. Our findings integrate the spatial regulation of hormones and transcriptome programs during the initiation of bamboo stem growth.

A single base change explains the independent origin of and selection for the nonshattering gene in African rice domestication.

Reduced seed shattering was a critical evolutionary step in crop domestication. Two cultivated rice species, Oryza sativa and Oryza glaberrima, were independently domesticated from the wild species Oryza rufipogon in Asia and Oryza barthii in Africa, respectively. A single nucleotide polymorphism (SNP) in the c gene, which encodes a trihelix transcription factor, causes nonshattering in O. sativa. However, the genetic mechanism of nonshattering in O. glaberrima is poorly understood. We conducted an association analysis for the coding sequences of SH3/SH4 in AA- genome rice species and the mutation suggested to cause nonshattering was demonstrated to do so using a positional-cloning approach in the O. sativa genetic background. We found that the loss of seed shattering in O. glaberrima was caused by an SNP resulting in a truncated SH3/SH4 protein. This mutation appears to be endemic and to have spread in the African gene pool by hybridization with some O. barthii accessions. We showed that interaction between the O. sativa and O. glaberrima domestication alleles of SH3 in heterozygotes induces a 'throwback' seed-shattering phenotype similar to that in the wild species. Identification of the causative SNP provides new insights into the molecular basis of seed shattering in crops and may facilitate investigation of the history of African rice domestication.

Construction of rice chromosome segment substitution lines harboring Oryza barthii genome and evaluation of yield-related traits.

Rice (Oryza sativa L.) is one of the most important staple food in the world. To meet the increasing demand for food, a strategy for improving rice yield is needed. Alleles of wild relatives are useful because they confer adaptation to plants under diverse harsh environments and have the potential to improve rice. O. barthii is a wild rice species endemic to Africa and the known progenitor of the African cultivated rice, O. glaberrima. To explore the genetic potential of the O. barthii as a genetic resource, 40 chromosome segment substitution lines (CSSL) of O. barthii in the background of the elite japonica cultivar Koshihikari were developed and evaluated to identify QTLs associated with 10 traits related to flag leaf morphology, grain yield and other agronomic traits. More than 90% of the entire genome of the donor parent was represented in contiguous or overlapping chromosome segments in the CSSLs. Evaluation of the CSSLs for several agriculturally important traits identified candidate chromosome segments that harbors QTLs associated with yield and yield-related traits. These results suggest that alleles from O. barthii might be used as a novel genetic resource for improving the yield-related traits in cultivars of O. sativa.

Analysis of Rhizome Development in Oryza longistaminata, a Wild Rice Species.

Vegetative reproduction is a form of asexual propagation in plants. A wide range of plants develop rhizomes, modified stems that grow underground horizontally, as a means of vegetative reproduction. In rhizomatous species, despite their distinct developmental patterns, both rhizomes and aerial shoots derive from axillary buds. Therefore, it is of interest to understand the basis of rhizome initiation and development. Oryza longistaminata, a wild rice species, develops rhizomes. We analyzed bud initiation and growth of O. longistaminata rhizomes using various methods of morphological observation. We show that, unlike aerial shoot buds that contain a few leaves only, rhizome buds initiate several leaves and bend to grow at right angles to the original rhizome. Rhizomes are maintained in the juvenile phase irrespective of the developmental phase of the aerial shoot. Stem elongation and reproductive transition are tightly linked in the aerial shoots, but are uncoupled in the rhizome. Our findings indicate that developmental programs operate independently in the rhizomes and aerial shoots. Temporal modification of the developmental pathways that are common to rhizomes and aerial shoots may be the source of developmental plasticity. Furthermore, the creation of new developmental systems appears to be necessary for rhizome development.

Oryza sativa H+-ATPase (OSA) is Involved in the Regulation of Dumbbell-Shaped Guard Cells of Rice.

The stomatal apparatus consists of a pair of guard cells and regulates gas exchange between the leaf and atmosphere. In guard cells, blue light (BL) activates H(+)-ATPase in the plasma membrane through the phosphorylation of its penultimate threonine, mediating stomatal opening. Although this regulation is thought to be widely adopted among kidney-shaped guard cells in dicots, the molecular basis underlying that of dumbbell-shaped guard cells in monocots remains unclear. Here, we show that H(+)-ATPases are involved in the regulation of dumbbell-shaped guard cells. Stomatal opening of rice was promoted by the H(+)-ATPase activator fusicoccin and by BL, and the latter was suppressed by the H(+)-ATPase inhibitor vanadate. Using H(+)-ATPase antibodies, we showed the presence of phosphoregulation of the penultimate threonine in Oryza sativa H(+)-ATPases (OSAs) and localization of OSAs in the plasma membrane of guard cells. Interestingly, we identified one H(+)-ATPase isoform, OSA7, that is preferentially expressed among the OSA genes in guard cells, and found that loss of function of OSA7 resulted in partial insensitivity to BL. We conclude that H(+)-ATPase is involved in BL-induced stomatal opening of dumbbell-shaped guard cells in monocotyledon species.

Development of chromosome segment substitution lines harboring Oryza nivara genomic segments in Koshihikari and evaluation of yield-related traits.

Chromosome segment substitution lines (CSSLs) are rich genetic resources that can be mined for novel, agriculturally useful loci or that can be used directly as materials for breeding. To date, a number of rice CSSLs have been developed by crossing rice cultivars with its wild relatives as a means to tap into the potential of wild alleles in rice improvement. Oryza nivara is a wild relative of rice that is thought to be a progenitor of O. sativa spp. indica. In the present study, 26 CSSLs that covers the entire genome of O. nivara as contiguous, overlapping segments in the genomic background of a japonica cultivar, O. sativa cv. Koshihikari were developed. Evaluation of the CSSLs for several agriculturally important traits identified candidate chromosome segments that harbors QTLs associated with yield and yield-related traits. The results of the study revealed the potential of O. nivara as a source of novel alleles that can be used to improve the existing japonica cultivar.

A mathematical model of phloem sucrose transport as a new tool for designing rice panicle structure for high grain yield.

Rice (Oryza sativa) is one of the most important food crops in the world. Numerous quantitative trait loci or genes controlling panicle architecture have been identified to increase grain yield. Yet grain yield, defined as the product of the number of well-ripened grains and their weight, is a complex trait that is determined by multiple factors such as source, sink and translocation capacity. Mechanistic modelling capturing capacities of source, sink and transport will help in the theoretical design of crop ideotypes that guarantee high grain yield. Here we present a mathematical model simulating sucrose transport and grain growth within a complex phloem network. The model predicts that the optimal panicle structure for high yield shows a simple grain arrangement with few higher order branches. In addition, numerical analyses revealed that inefficient delivery of carbon to panicles with higher order branches prevails regardless of source capacity, indicating the importance of designing grain arrangement and phloem structure. Our model highlights the previously unexplored effect of grain arrangement on the yield, and provides numerical solutions for optimal panicle structure under various source and sink capacities.