Regulation of Flowering Time and Other Developmental Plasticities by 3' Splicing Factor-Mediated Alternative Splicing in Arabidopsis thaliana.

Plants, as sessile organisms, show a high degree of plasticity in their growth and development and have various strategies to cope with these alterations under continuously changing environments and unfavorable stress conditions. In particular, the floral transition from the vegetative and reproductive phases in the shoot apical meristem (SAM) is one of the most important developmental changes in plants. In addition, meristem regions, such as the SAM and root apical meristem (RAM), which continually generate new lateral organs throughout the plant life cycle, are important sites for developmental plasticity. Recent findings have shown that the prevailing type of alternative splicing (AS) in plants is intron retention (IR) unlike in animals; thus, AS is an important regulatory mechanism conferring plasticity for plant growth and development under various environmental conditions. Although eukaryotes exhibit some similarities in the composition and dynamics of their splicing machinery, plants have differences in the 3' splicing characteristics governing AS. Here, we summarize recent findings on the roles of 3' splicing factors and their interacting partners in regulating the flowering time and other developmental plasticities in Arabidopsis thaliana.

Design and Fabrication of Micro Saw Enabling Root-Side Cutting of Bone.

A novel micro saw was fabricated using a combination of photolithography and electroplating techniques, resembling a miniature timing belt with sideways blades. The rotation or oscillation direction of the micro saw is designed to be perpendicular to the cutting direction so that transverse cutting of the bone is attainable to extract a preoperatively planned bone-cartilage donor for osteochondral auto-graft transplantation. The mechanical property of the fabricated micro saw obtained using the nanoindentation test shows that the mechanical properties of the micro saw are almost an order of magnitude higher than bone, which indicates its potential bone-cutting application. To demonstrate the cutting capability of the fabricated micro saw, an in vitro animal bone cutting was performed using a custom test rig consisting of a microcontroller, 3D printer, and other readily available parts.

Reduced Expression of PRX2/ATPRX1, PRX8, PRX35, and PRX73 Affects Cell Elongation, Vegetative Growth, and Vasculature Structures in Arabidopsis thaliana.

Class III peroxidases (PRXs) are involved in a broad spectrum of physiological and developmental processes throughout the life cycle of plants. However, the specific function of each PRX member in the family remains largely unknown. In this study, we selected four class III peroxidase genes (PRX2/ATPRX1, PRX8, PRX35, and PRX73) from a previous genome-wide transcriptome analysis, and performed phenotypic and morphological analyses, including histochemical staining, in PRX2RNAi, PRX8RNAi, PRX35RNAi, and PRX73RNAi plants. The reduced mRNA levels of corresponding PRX genes in PRX2RNAi, PRX8RNAi, PRX35RNAi, and PRX73RNAi seedlings resulted in elongated hypocotyls and roots, and slightly faster vegetative growth. To investigate internal structural changes in the vasculature, we performed histochemical staining, which revealed alterations in cell wall structures in the main vasculature of hypocotyls, stems, and roots of each PRXRNAi plant compared to wild-type (Col-0) plants. Furthermore, we found that PRX35RNAi plants displayed the decrease in the cell wall in vascular regions, which are involved in downregulation of lignin biosynthesis and biosynthesis-regulated genes' expression. Taken together, these results indicated that the reduced expression levels of PRX2/ATPRX1, PRX8, PRX35, and PRX73 affected hypocotyl and root elongation, vegetative growth, and the vasculature structures in hypocotyl, stem, and root tissues, suggesting that the four class III PRX genes play roles in plant developmental processes.

Profiling Cannabinoid Contents and Expression Levels of Corresponding Biosynthetic Genes in Commercial Cannabis (Cannabis sativa L.) Cultivars.

Cannabis (Cannabis sativa L.) is widely cultivated and studied for its psychoactive and medicinal properties. As the major cannabinoids are present in acidic forms in Cannabis plants, non-enzymatic processes, such as decarboxylation, are crucial for their conversion to neutral active cannabinoid forms. Herein, we detected the levels of cannabidivarin (CBDV), cannabidiol (CBD), cannabichromene (CBC), and Δ9-tetrahydrocannabinol (Δ9-THC) in the leaves and vegetative shoots of five commercial Cannabis cultivars using a combination of relatively simple extraction, decarboxylation, and high-performance liquid chromatography analyses. The CBDV, CBC, and Δ9-THC levels were 6.3-114.9, 34.4-187.2, and 57.6-407.4 µg/g, respectively, and the CBD levels were the highest, ranging between 1.2-8.9 µg/g in leaf and vegetative shoot tissues of Cannabis cultivars. Additionally, correlations were observed between cannabinoid accumulation and transcription levels of genes encoding key enzymes for cannabinoid biosynthesis, including CsCBGAS, CsCBDAS, CsCBCAS, and CsTHCAS. These data suggest that the high accumulation of cannabinoids, such as CBC, Δ9-THC, and CBD, might be derived from the transcriptional regulation of CsCBGAS and CsCBDAS in Cannabis plants.

The splicing factor 1-FLOWERING LOCUS M module spatially regulates temperature-dependent flowering by modulating FLOWERING LOCUS T and LEAFY expression.

KEY MESSAGE: The AtSF1-FLM module spatially controls temperature-dependent flowering by negatively regulating the expression of FT and LFY in the leaf and shoot apex, respectively. Alternative splicing mediated by various splicing factors is important for the regulation of plant growth and development. Our recent reports have shown that a temperature-dependent interaction between Arabidopsis thaliana splicing factor 1 (AtSF1) and FLOWERING LOCUS M (FLM) pre-mRNA introns controls the differential production of FLM-ß transcripts at different temperatures, eventually resulting in temperature-responsive flowering. However, the molecular and genetic interactions between the AtSF1-FLM module and floral activator genes remain unknown. Here, we aimed to identify the interactions among AtSF1, FLM, FLOWERING LOCUS T (FT), and LEAFY (LFY) by performing molecular and genetic analyses. FT and TWIN SISTER OF FT (TSF) expression in atsf1-2 mutants significantly increased in the morning and middle of the night at 16 and 23 °C, respectively, under long-day conditions. In addition, ft mutation suppressed the early flowering of atsf1-2 and atsf1-2 flm-3 mutants and masked the temperature response of atsf1-2 flm-3 mutants, suggesting that FT is a downstream target gene of the AtSF1-FLM module. LFY expression significantly increased in the diurnal samples of atsf1-2 mutants and in the shoot apex regions of atsf1-2 ft-10 mutants at different temperatures. The chromatin immunoprecipitation (ChIP) assay revealed that FLM directly binds to the genomic regions of LFY but not of APETALA1 (AP1). Moreover, lfy mutation suppressed the early flowering of flm-3 mutants, suggesting that LFY is another target of the AtSF1-FLM module. Our results reveal that the AtSF1-FLM module spatially modulates temperature-dependent flowering by regulating FT and LFY expressions.

Applications and Major Achievements of Genome Editing in Vegetable Crops: A Review.

The emergence of genome-editing technology has allowed manipulation of DNA sequences in genomes to precisely remove or replace specific sequences in organisms resulting in targeted mutations. In plants, genome editing is an attractive method to alter gene functions to generate improved crop varieties. Genome editing is thought to be simple to use and has a lower risk of off-target effects compared to classical mutation breeding. Furthermore, genome-editing technology tools can also be applied directly to crops that contain complex genomes and/or are not easily bred using traditional methods. Currently, highly versatile genome-editing tools for precise and predictable editing of almost any locus in the plant genome make it possible to extend the range of application, including functional genomics research and molecular crop breeding. Vegetables are essential nutrient sources for humans and provide vitamins, minerals, and fiber to diets, thereby contributing to human health. In this review, we provide an overview of the brief history of genome-editing technologies and the components of genome-editing tool boxes, and illustrate basic modes of operation in representative systems. We describe the current and potential practical application of genome editing for the development of improved nutritious vegetables and present several case studies demonstrating the potential of the technology. Finally, we highlight future directions and challenges in applying genome-editing systems to vegetable crops for research and product development.

Critical enzymes for biosynthesis of cucurbitacin derivatives in watermelon and their biological significance.

Various cucurbitacins have been isolated, and their structures have been elucidated. Owing to their economic potential and importance as active pharmacological compounds, their cytotoxicity in various cancer cells has been assessed. Here, we mined several candidate genes with potential involvement in cucurbitacin biosynthesis in watermelon (Citrullus lanatus) and performed in vitro enzymatic assays and instrumental analyses using various substrates to identify cucurbitacin functions and products. Enzymatic activities of two acetyltransferases (ACTs) and one UDP-glucosyltransferase (UGT) against cucurbitacins were confirmed, resulting in the synthesis of novel cucurbitacins in vivo and/or in vitro to our knowledge. As ACTs and UGT are involved in the dynamic conversion of cucurbitacins by catalyzing acetylation and glucosylation at moieties in the cucurbitacins skeleton, these findings improve our knowledge on how these genes contribute to the diversity of cucurbitacins.

Bending Strain and Bending Fatigue Lifetime of Flexible Metal Electrodes on Polymer Substrates.

As the technology of flexible electronics has remarkably advanced, the long-term reliability of flexible devices has attracted much attention, as it is an important factor for such devices in reaching real commercial viability. To guarantee the bending fatigue lifetime, the exact evaluation of bending strain and the change in electrical resistance is required. In this study, we investigated the bending strains of Cu thin films on flexible polyimide substrates with different thicknesses using monolayer and bilayer bending models and monitored the electrical resistance of the metal electrode during a bending fatigue test. For a thin metal electrode, the bending strain and fatigue lifetime were similar regardless of substrate thickness, but for a thick metal film, the fatigue lifetime was changed by different bending strains in the metal electrode according to substrate thickness. To obtain the exact bending strain distribution, we conducted a finite-element simulation and compared the bending strains of thin and thick metal structures. For thick metal electrodes, the real bending strain obtained from a bilayer model or simulation showed values much different from those from a simple monolayer model. This study can provide useful guidelines for developing highly reliable flexible electronics.

Two types of mutations in the HEUKCHEEM gene functioning in cucumber spine color development can be used as signatures for cucumber domestication.

MAIN CONCLUSION: The HEUKCHEEM gene plays an important role in spine color formation. A white spine occurs due to two mutations in HEUKCHEEM and is closely related to the regional distribution of these mutants. Mapping analysis revealed that the HEUKCHEEM gene is co-segregated with the B locus in the regulation of black spine color development in cucumber fruit. HEUKCHEEM induced the expression of the genes involved in the anthocyanin biosynthetic pathway, leading to the accumulation of anthocyanins in black spines. The transiently over-expressed HEUKCHEEM in cucumber and tobacco plants enhanced the expression of anthocyanin biosynthesis-related genes, leading to anthocyanin accumulation. However, two mutations-insertion of the 6994 bp mutator-like transposable element (MULE) sequence into the second intron and one single-nucleotide polymorphism (SNP) of C to T in the second exon of HEUKCHEEM-were identified in white spines, leading to no accumulation of anthocyanin biosynthesis-related gene transcripts and anthocyanins. Furthermore, association analysis using 104 cucumber accessions with different geographical origins revealed that the types of mutations in HEUKCHEEM are strongly linked to geographical origins. The MULE insertion is found extensively in cucumbers with white spines in East Asia and Australia. However, cucumbers with white spines in other areas could be significantly influenced by a single SNP mutation. Our results provide fundamental information on spine color development in cucumber fruits and spine color-based cucumber breeding programs.

Identification of quantitative trait loci governing subgynoecy in cucumber.

KEY MESSAGE: QTL-seq analysis identified three major QTLs conferring subgynoecy in cucumbers. Furthermore, sequence and expression analyses predicted candidate genes controlling subgynoecy. The cucumber (Cucumis sativus L.) is a typical monoecious having individual male and female flowers, and sex differentiation is an important developmental process that directly affects its fruit yield. Subgynoecy represents a sex form with a high degree of femaleness and would have alternative use as gynoecy under limited resource conditions. Recently, many studies have been reported that QTL-seq, which integrates the advantages of bulked segregant analysis and high-throughput whole-genome resequencing, can be a rapid and cost-effective way of mapping QTLs. Segregation analysis in the F2 and BC1 populations derived from a cross between subgynoecious LOSUAS and monoecious BMB suggested the quantitative nature of subgynoecy in cucumbers. Both genome-wide SNP profiling of subgynoecious and monoecious bulks constructed from F2 and BC1 plants consistently identified three significant genomic regions, one on chromosome 3 (sg3.1) and another two on short and long arms of chromosome 1 (sg1.1 and sg1.2). Classical QTL analysis using the F2 confirmed sg3.1 (R2 = 42%), sg1.1 (R2 = 29%) and sg1.2 (R2 = 18%) as major QTLs. These results revealed the unique genetic inheritance of subgynoecious line LOSUAS through two distinct major QTLs, sg3.1 and sg1.1, which mainly increase degree of femaleness, while another QTL, sg1.2, contributes to decrease it. This study demonstrated that QTL-seq allows rapid and powerful detection of QTLs using preliminary generation mapping populations such as F2 or BC1 population and further that the identified QTLs could be useful for molecular breeding of cucumber lines with high yield potential.

The overexpression of cucumber (Cucumis sativus L.) genes that encode the branched-chain amino acid transferase modulate flowering time in Arabidopsis thaliana.

KEY MESSAGE: The overexpression of CsBCATs promotes flowering in Arabidopsis by regulating the expression of flowering time genes. The branched-chain amino acid transferases (BCATs) play an important role in the metabolism of branched-chain amino acids (BCAAs), such as isoleucine, leucine, and valine. They function in both the synthesis and the degradation of this class of amino acids. We identified and characterized the three BCAT genes in cucumber (Cucumis sativus L.). The tissue-specific expression profiling in cucumber plants revealed that CsBCAT2 and CsBCAT7 were highly expressed in the reproductive tissues, whereas CsBCAT3 expression was highly detected in the vegetative tissues. The subcellular localization patterns of three CsBCATs were observed in the mitochondria. The functional analyses of CsBCATs showed that CsBCAT2 and CsBCAT3 restored the growth of bat1Δ/bat2Δ double knockout yeast (Saccharomyces cerevisiae), and CsBCAT3 and CsBCAT7 with different substrate preferences acted in a reverse reaction. The transgenic approach demonstrated that the overexpression of the three CsBCATs resulted in early flowering phenotypes, which were associated with the upregulation of FLOWERING LOCUS T (FT) and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 (SOC1) in a manner in which they were dependent on GIGANTEA (GI)/CONSTANS (CO) and SHORT VEGETATIVE PHASE (SVP)/FLOWERING LOCUS C (FLC) modules. Our results, which are observed in conjunction, suggest that there is an interconnection between BCAT genes that function in BCAA metabolism and the flowering time in plants.

The Arabidopsis Cysteine-Rich Receptor-Like Kinase CRK36 Regulates Immunity through Interaction with the Cytoplasmic Kinase BIK1.

Receptor-like kinases are important signaling components that regulate a variety of cellular processes. In this study, an Arabidopsis cDNA microarray analysis led to the identification of the cysteine-rich receptor-like kinase CRK36 responsive to the necrotrophic fungal pathogen, Alternaria brassicicola. To determine the function of CRK36 in plant immunity, T-DNA-insertion knockdown (crk36) and overexpressing (CRK36OE) plants were prepared. CRK36OE plants exhibited increased hypersensitive cell death and ROS burst in response to avirulent pathogens. Treatment with a typical pathogen-associated molecular pattern, flg22, markedly induced pattern-triggered immune responses, notably stomatal defense, in CRK36OE plants. The immune responses were weakened in crk36 plants. Protein-protein interaction assays revealed the in vivo association of CRK36, FLS2, and BIK1. CRK36 enhanced flg22-triggered BIK1 phosphorylation, which showed defects with Cys mutations in the DUF26 motifs of CRK36. Disruption of BIK1 and RbohD/RbohF genes further impaired CRK36-mediated stomatal defense. We propose that CRK36, together with BIK1 and NADPH oxidases, may form a positive activation loop that enhances ROS burst and leads to the promotion of stomatal immunity.

Wall-thickness-dependent strength of nanotubular ZnO.

We fabricate nanotubular ZnO with wall thickness of 45, 92, 123 nm using nanoporous gold (np-Au) with ligament diameter at necks of 1.43 µm as sacrificial template. Through micro-tensile and micro-compressive testing of nanotubular ZnO structures, we find that the exponent m in [Formula: see text], where [Formula: see text] is the relative strength and [Formula: see text] is the relative density, for tension is 1.09 and for compression is 0.63. Both exponents are lower than the value of 1.5 in the Gibson-Ashby model that describes the relation between relative strength and relative density where the strength of constituent material is independent of external size, which indicates that strength of constituent ZnO increases as wall thickness decreases. We find, based on hole-nanoindentation and glazing incidence X-ray diffraction, that this wall-thickness-dependent strength of nanotubular ZnO is not caused by strengthening of constituent ZnO by size reduction at the nanoscale. Finite element analysis suggests that the wall-thickness-dependent strength of nanotubular ZnO originates from nanotubular structures formed on ligaments of np-Au.

Fully-integrated, bezel-less transistor arrays using reversibly foldable interconnects and stretchable origami substrates.

Here we demonstrate fully-integrated, bezel-less transistor arrays using stretchable origami substrates and foldable conducting interconnects. Reversible folding of these arrays is enabled by origami substrates which are composed of rigid support fixtures and foldable elastic joints. In addition, hybrid structures of thin metal films and metallic nanowires worked as foldable interconnects which are located on the elastomeric joints.

Stretchable, Transparent Electrodes as Wearable Heaters Using Nanotrough Networks of Metallic Glasses with Superior Mechanical Properties and Thermal Stability.

Mechanical robustness, electrical and chemical reliabilities of devices against large deformations such as bending and stretching have become the key metrics for rapidly emerging wearable electronics. Metallic glasses (MGs) have high elastic limit, electrical conductivity, and corrosion resistance, which can be promising for applications in wearable electronics. However, their applications in wearable electronics or transparent electrodes have not been extensively explored so far. Here, we demonstrate stretchable and transparent electrodes using CuZr MGs in the form of nanotrough networks. MG nanotroughs are prepared by electrospinning and cosputtering process, and they can be transferred to various desired substrates, including stretchable elastomeric substrates. The resulting MG nanotrough network is first utilized as a stretchable transparent electrode, presenting outstanding optoelectronic (sheet resistance of 3.8 Ω/sq at transmittance of 90%) and mechanical robustness (resistance change less than 30% up to a tensile strain of 70%) as well as excellent chemical stability against hot and humid environments (negligible degradation in performance for 240 h in 85% relative humidity and 85 °C). A stretchable and transparent heater based on the MG nanotrough network is also demonstrated with a wide operating temperature range (up to 180 °C) and excellent stretchability (up to 70% in the strain). The excellent mechanical robustness of these stretchable transparent electrode and heater is ascribed to the structural configuration (i.e., a nanotrough network) and inherent high elastic limit of MGs, as supported by experimental results and numerical analysis. We demonstrate their real-time operations on human skin as a wearable, transparent thermotherapy patch controlled wirelessly using a smartphone as well as a transparent defroster for an automobile side-view mirror, suggesting a promising strategy toward next-generation wearable electronics or automobile applications.

Identification and origin of N-linked ß-D-N-acetylglucosamine monosaccharide modifications on Arabidopsis proteins.

Many plant proteins are modified with N-linked oligosaccharides at asparagine-X-serine/threonine sites during transit through the endoplasmic reticulum and the Golgi. We have identified a number of Arabidopsis (Arabidopsis thaliana) proteins with modifications consisting of an N-linked N-acetyl-D-glucosamine monosaccharide (N-GlcNAc). Electron transfer dissociation mass spectrometry analysis of peptides bearing this modification mapped the modification to asparagine-X-serine/threonine sites on proteins that are predicted to transit through the endoplasmic reticulum and Golgi. A mass labeling method was developed and used to study N-GlcNAc modification of two thioglucoside glucohydrolases (myrosinases), TGG1 and TGG2 (for thioglucoside glucohydrolase). These myrosinases are also modified with high-mannose (Man)-type glycans. We found that N-GlcNAc and high-Man-type glycans can occur at the same site. It has been hypothesized that N-GlcNAc modifications are generated when endo-ß-N-acetylglucosaminidase (ENGase) cleaves N-linked glycans. We examined the effects of mutations affecting the two known Arabidopsis ENGases on N-GlcNAc modification of myrosinase and found that modification of TGG2 was greatly reduced in one of the single mutants and absent in the double mutant. Surprisingly, N-GlcNAc modification of TGG1 was not affected in any of the mutants. These data support the hypothesis that ENGases hydrolyze high-Man glycans to produce some of the N-GlcNAc modifications but also suggest that some N-GlcNAc modifications are generated by another mechanism. Since N-GlcNAc modification was detected at only one site on each myrosinase, the production of the N-GlcNAc modification may be regulated.

O-GlcNAcylation of the Plum pox virus capsid protein catalyzed by SECRET AGENT: characterization of O-GlcNAc sites by electron transfer dissociation mass spectrometry.

The capsid protein of Plum pox virus (PPV-CP) is modified with O-linked ß-N-acetylglucosamine (O-GlcNAc). In Arabidopsis thaliana this modification is made by an O-GlcNAc transferase named SECRET AGENT (SEC). Modification of PPV-CP by SEC is hypothesized to have a direct role in the infection process, because virus titer and rate of spread are reduced in SEC mutants. Previous studies used deletion mapping and site-directed mutagenesis to identify four O-GlcNAc sites on the capsid protein that are modified by Escherichia coli-expressed SEC. The infection process was not affected when two of these sites were mutated suggesting that O-GlcNAcylation of these sites does not have a significant role in the infection process or that a subset of the modifications is sufficient. Since it is possible that the mutational mapping approach missed or incorrectly identified O-GlcNAc sites, the modifications produced by E. coli-expressed SEC were characterized using mass spectrometry. O-GlcNAcylated peptides were enzymatically tagged with galactose, the products were enriched on immobilized Ricinus communis agglutinin I and sequenced by electron transfer dissociation (ETD) mass spectrometry. Five O-GlcNAc sites on PPV-CP were identified. Two of these sites were not identified in by the previous mutational mapping. In addition, one site previously predicted by mutation mapping was not detected, but modification of this site was not supported when the mutation mapping was repeated. This study suggests that mapping modification sites by ETD mass spectrometry is more comprehensive and accurate than mutational mapping.

Multiple loss-of-function of Arabidopsis gibberellin receptor AtGID1s completely shuts down a gibberellin signal.

Arabidopsis carries three receptor genes for the phytohormone gibberellin (GA), AtGID1a, AtGID1b and AtGID1c. Expression of each gene in the rice gid1-1 mutant for GA receptors causes reversion of its severely dwarfed phenotype and GA insensitivity to a normal level, even though each loss-of-function mutant shows no clear phenotype in Arabidopsis (Nakajima et al., 2006). In this paper, we report the functional redundancy and specificity of each AtGID1 by analyzing the multiple mutants for loss of function. Seeds of the double knockout mutants atgid1a atgid1b, atgid1a atgid1c and atgid1b atgid1c germinated normally. The double knockout mutant atgid1a atgid1c showed a dwarf phenotype, while other double mutants were of normal height compared to the wild-type. The stamens of the double knockout mutant atgid1a atgid1b were significantly shorter than those of the wild-type, and this leads to low fertility. A severe disarrangement of the pattern on its seed surface was also observed. The triple knockout mutant atgid1a atgid1b atgid1c did not germinate voluntarily, and only started to grow when the seed coat was peeled off after soaking. Seedlings of the triple knockout mutants were severe dwarfs, only a few millimeters high after growing for 1 month. Moreover, the triple knockout seedlings completely lost their ability to respond to exogenously applied GA. These results show that all AtGID1s function as GA receptors in Arabidopsis, but have specific role(s) for growth and development.

Identification and characterization of Arabidopsis gibberellin receptors.

Three gibberellin (GA) receptor genes (AtGID1a, AtGID1b and AtGID1c), each an ortholog of the rice GA receptor gene (OsGID1), were cloned from Arabidopsis, and the characteristics of their recombinant proteins were examined. The GA-binding activities of the three recombinant proteins were confirmed by an in vitro assay. Biochemical analyses revealed similar ligand selectivity among the recombinants, and all recombinants showed higher affinity to GA(4) than to other GAs. AtGID1b was unique in its binding affinity to GA(4) and in its pH dependence when compared with the other two, by only showing binding in a narrow pH range (pH 6.4-7.5) with 10-fold higher affinity (apparent K(d) for GA(4) = 3 x 10(-8) m) than AtGID1a and AtGID1c. A two-hybrid yeast system only showed in vivo interaction in the presence of GA(4) between each AtGID1 and the Arabidopsis DELLA proteins (AtDELLAs), negative regulators of GA signaling. For this interaction with AtDELLAs, AtGID1b required only one-tenth of the amount of GA(4) that was necessary for interaction between the other AtGID1s and AtDELLAs, reflecting its lower K(d) value. AtDELLA boosted the GA-binding activity of AtGID1 in vitro, which suggests the formation of a complex between AtDELLA and AtGID1-GA that binds AtGID1 to GA more tightly. The expression of each AtGID1 clone in the rice gid1-1 mutant rescued the GA-insensitive dwarf phenotype. These results demonstrate that all three AtGID1s functioned as GA receptors in Arabidopsis.