Mechanisms of Endoplasmic Reticulum Protein Homeostasis in Plants.

Maintenance of proteome integrity is essential for cell function and survival in changing cellular and environmental conditions. The endoplasmic reticulum (ER) is the major site for the synthesis of secretory and membrane proteins. However, the accumulation of unfolded or misfolded proteins can perturb ER protein homeostasis, leading to ER stress and compromising cellular function. Eukaryotic organisms have evolved sophisticated and conserved protein quality control systems to ensure protein folding fidelity via the unfolded protein response (UPR) and to eliminate potentially harmful proteins via ER-associated degradation (ERAD) and ER-phagy. In this review, we summarize recent advances in our understanding of the mechanisms of ER protein homeostasis in plants and discuss the crosstalk between different quality control systems. Finally, we will address unanswered questions in this field.

Arabidopsis 3ß-Hydroxysteroid Dehydrogenases/C4-Decarboxylases Are Essential for the Pollen and Embryonic Development.

The biosynthesis of C27-29 sterols from their C30 precursor squalene involves C24-alkylation and the removal of three methyl groups, including two at the C4 position. The two C4 demethylation reactions require a bifunctional enzyme known as 3ß-hydroxysteroid dehydrogenase/C4-decarboxylase (3ßHSD/D), which removes an oxidized methyl (carboxylic) group at C4 while simultaneously catalyzing the 3ß-hydroxyl→3-keto oxidation. Its loss-of-function mutations cause ergosterol-dependent growth in yeast and congenital hemidysplasia with ichthyosiform erythroderma and limb defect (CHILD) syndrome in humans. Although plant 3ßHSD/D enzymes were well studied enzymatically, their developmental functions remain unknown. Here we employed a CRISPR/Cas9-based genome-editing approach to generate knockout mutants for two Arabidopsis 3ßHSD/D genes, HSD1 and HSD2, and discovered the male gametophytic lethality for the hsd1 hsd2 double mutation. Pollen-specific expression of HSD2 in the heterozygous hsd1 hsd2/+ mutant not only rescued the pollen lethality but also revealed the critical roles of the two HSD genes in embryogenesis. Our study thus demonstrated the essential functions of the two Arabidopsis 3ßHSD/D genes in male gametogenesis and embryogenesis.

Assessment of Biological Activity of 28-Homobrassinolide via a Multi-Level Comparative Analysis.

Brassinosteroids (BRs) play vital roles in the plant life cycle and synthetic BRs are widely used to increase crop yield and plant stress tolerance. Among them are 24R-methyl-epibrassinolide (24-EBL) and 24S-ethyl-28-homobrassinolide (28-HBL), which differ from brassinolide (BL, the most active BR) at the C-24 position. Although it is well known that 24-EBL is 10% active as BL, there is no consensus on the bioactivity of 28-HBL. A recent outpouring of research interest in 28-HBL on major crops accompanied with a surge of industrial-scale synthesis that produces mixtures of active (22R,23R)-28-HBL and inactive (22S,23S)-28HBL, demands a standardized assay system capable of analyzing different synthetic "28-HBL" products. In this study, the relative bioactivity of 28-HBL to BL and 24-EBL, including its capacity to induce the well-established BR responses at molecular, biochemical, and physiological levels, was systematically analyzed using the whole seedlings of the wild-type and BR-deficient mutant of Arabidopsis thaliana. These multi-level bioassays consistently showed that 28-HBL exhibits a much stronger bioactivity than 24-EBL and is almost as active as BL in rescuing the short hypocotyl phenotype of the dark-grown det2 mutant. These results are consistent with the previously established structure-activity relationship of BRs, proving that this multi-level whole seedling bioassay system could be used to analyze different batches of industrially produced 28-HBL or other BL analogs to ensure the full potential of BRs in modern agriculture.

Engineered ATG8-binding motif-based selective autophagy to degrade proteins and organelles in planta.

Protein-targeting technologies represent essential approaches in biological research. Protein knockdown tools developed recently in mammalian cells by exploiting natural degradation mechanisms allow for precise determination of protein function and discovery of degrader-type drugs. However, no method to directly target endogenous proteins for degradation is currently available in plants. Here, we describe a novel method for targeted protein clearance by engineering an autophagy receptor with a binder to provide target specificity and an ATG8-binding motif (AIM) to link the targets to nascent autophagosomes, thus harnessing the autophagy machinery for degradation. We demonstrate its specificity and broad potentials by degrading various fluorescence-tagged proteins, including cytosolic mCherry, the nucleus-localized bZIP transcription factor TGA5, and the plasma membrane-anchored brassinosteroid receptor BRI1, as well as fluorescence-coated peroxisomes, using a tobacco-based transient expression system. Stable expression of AIM-based autophagy receptors in Arabidopsis further confirms the feasibility of this approach in selective autophagy of endogenous proteins. With its wide substrate scope and its specificity, our concept of engineered AIM-based selective autophagy could provide a convenient and robust research tool for manipulating endogenous proteins in plants and may open an avenue toward degradation of cytoplasmic components other than proteins in plant research.

A Predominant Role of AtEDEM1 in Catalyzing a Rate-Limiting Demannosylation Step of an Arabidopsis Endoplasmic Reticulum-Associated Degradation Process.

Endoplasmic reticulum-associated degradation (ERAD) is a key cellular process for degrading misfolded proteins. It was well known that an asparagine (N)-linked glycan containing a free α1,6-mannose residue is a critical ERAD signal created by Homologous to α-mannosidase 1 (Htm1) in yeast and ER-Degradation Enhancing α-Mannosidase-like proteins (EDEMs) in mammals. An earlier study suggested that two Arabidopsis homologs of Htm1/EDEMs function redundantly in generating such a conserved N-glycan signal. Here we report that the Arabidopsis irb1 (reversal of bri1) mutants accumulate brassinosteroid-insensitive 1-5 (bri1-5), an ER-retained mutant variant of the brassinosteroid receptor BRI1 and are defective in one of the Arabidopsis Htm1/EDEM homologs, AtEDEM1. We show that the wild-type AtEDEM1, but not its catalytically inactive mutant, rescues irb1-1. Importantly, an insertional mutation of the Arabidopsis Asparagine-Linked Glycosylation 3 (ALG3), which causes N-linked glycosylation with truncated glycans carrying a different free α1,6-mannose residue, completely nullifies the inhibitory effect of irb1-1 on bri1-5 ERAD. Interestingly, an insertional mutation in AtEDEM2, the other Htm1/EDEM homolog, has no detectable effect on bri1-5 ERAD; however, it enhances the inhibitory effect of irb1-1 on bri1-5 degradation. Moreover, AtEDEM2 transgenes rescued the irb1-1 mutation with lower efficacy than AtEDEM1. Simultaneous elimination of AtEDEM1 and AtEDEM2 completely blocks generation of α1,6-mannose-exposed N-glycans on bri1-5, while overexpression of either AtEDEM1 or AtEDEM2 stimulates bri1-5 ERAD and enhances the bri1-5 dwarfism. We concluded that, despite its functional redundancy with AtEDEM2, AtEDEM1 plays a predominant role in promoting bri1-5 degradation.

The Crucial Role of Demannosylating Asparagine-Linked Glycans in ERADicating Misfolded Glycoproteins in the Endoplasmic Reticulum.

Most membrane and secreted proteins are glycosylated on certain asparagine (N) residues in the endoplasmic reticulum (ER), which is crucial for their correct folding and function. Protein folding is a fundamentally inefficient and error-prone process that can be easily interfered by genetic mutations, stochastic cellular events, and environmental stresses. Because misfolded proteins not only lead to functional deficiency but also produce gain-of-function cellular toxicity, eukaryotic organisms have evolved highly conserved ER-mediated protein quality control (ERQC) mechanisms to monitor protein folding, retain and repair incompletely folded or misfolded proteins, or remove terminally misfolded proteins via a unique ER-associated degradation (ERAD) mechanism. A crucial event that terminates futile refolding attempts of a misfolded glycoprotein and diverts it into the ERAD pathway is executed by removal of certain terminal α1,2-mannose (Man) residues of their N-glycans. Earlier studies were centered around an ER-type α1,2-mannosidase that specifically cleaves the terminal α1,2Man residue from the B-branch of the three-branched N-linked Man9GlcNAc2 (GlcNAc for N-acetylglucosamine) glycan, but recent investigations revealed that the signal that marks a terminally misfolded glycoprotein for ERAD is an N-glycan with an exposed α1,6Man residue generated by members of a unique folding-sensitive α1,2-mannosidase family known as ER-degradation enhancing α-mannosidase-like proteins (EDEMs). This review provides a historical recount of major discoveries that led to our current understanding on the role of demannosylating N-glycans in sentencing irreparable misfolded glycoproteins into ERAD. It also discusses conserved and distinct features of the demannosylation processes of the ERAD systems of yeast, mammals, and plants.

PAWH1 and PAWH2 are plant-specific components of an Arabidopsis endoplasmic reticulum-associated degradation complex.

Endoplasmic reticulum-associated degradation (ERAD) is a unique mechanism to degrade misfolded proteins via complexes containing several highly-conserved ER-anchored ubiquitin ligases such as HMG-CoA reductase degradation1 (Hrd1). Arabidopsis has a similar Hrd1-containing ERAD machinery; however, our knowledge of this complex is limited. Here we report two closely-related Arabidopsis proteins, Protein Associated With Hrd1-1 (PAWH1) and PAWH2, which share a conserved domain with yeast Altered Inheritance of Mitochondria24. PAWH1 and PAWH2 localize to the ER membrane and associate with Hrd1 via EMS-mutagenized Bri1 Suppressor7 (EBS7), a plant-specific component of the Hrd1 complex. Simultaneously elimination of two PAWHs constitutively activates the unfolded protein response and compromises stress tolerance. Importantly, the pawh1 pawh2 double mutation reduces the protein abundance of EBS7 and Hrd1 and inhibits degradation of several ERAD substrates. Our study not only discovers additional plant-specific components of the Arabidopsis Hrd1 complex but also reveals a distinct mechanism for regulating the Hrd1 stability.

Communications Between the Endoplasmic Reticulum and Other Organelles During Abiotic Stress Response in Plants.

To adapt to constantly changing environmental conditions, plants have evolved sophisticated tolerance mechanisms to integrate various stress signals and to coordinate plant growth and development. It is well known that inter-organellar communications play important roles in maintaining cellular homeostasis in response to environmental stresses. The endoplasmic reticulum (ER), extending throughout the cytoplasm of eukaryotic cells, is a central organelle involved in lipid metabolism, Ca2+ homeostasis, and synthesis and folding of secretory and transmembrane proteins crucial to perceive and transduce environmental signals. The ER communicates with the nucleus via the highly conserved unfolded protein response pathway to mitigate ER stress. Importantly, recent studies have revealed that the dynamic ER network physically interacts with other intracellular organelles and endomembrane compartments, such as the Golgi complex, mitochondria, chloroplast, peroxisome, vacuole, and the plasma membrane, through multiple membrane contact sites between closely apposed organelles. In this review, we will discuss the signaling and metabolite exchanges between the ER and other organelles during abiotic stress responses in plants as well as the ER-organelle membrane contact sites and their associated tethering complexes.

Author Correction: Nitrate-NRT1.1B-SPX4 cascade integrates nitrogen and phosphorus signalling networks in plants.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Nitrate-NRT1.1B-SPX4 cascade integrates nitrogen and phosphorus signalling networks in plants.

To ensure high crop yields in a sustainable manner, a comprehensive understanding of the control of nutrient acquisition is required. In particular, the signalling networks controlling the coordinated utilization of the two most highly demanded mineral nutrients, nitrogen and phosphorus, are of utmost importance. Here, we reveal a mechanism by which nitrate activates both phosphate and nitrate utilization in rice (Oryza sativa L.). We show that the nitrate sensor NRT1.1B interacts with a phosphate signalling repressor SPX4. Nitrate perception strengthens the NRT1.1B-SPX4 interaction and promotes the ubiquitination and degradation of SPX4 by recruiting NRT1.1B interacting protein 1 (NBIP1), an E3 ubiquitin ligase. This in turn allows the key transcription factor of phosphate signalling, PHR2, to translocate to the nucleus and initiate the transcription of phosphorus utilization genes. Interestingly, the central transcription factor of nitrate signalling, NLP3, is also under the control of SPX4. Thus, nitrate-triggered degradation of SPX4 activates both phosphate- and nitrate-responsive genes, implementing the coordinated utilization of nitrogen and phosphorus.

Big Grain3, encoding a purine permease, regulates grain size via modulating cytokinin transport in rice.

Grain size is an important agronomic trait affecting grain yield, but the underlying molecular mechanisms remain to be elucidated. Here, we isolated a dominant mutant, big grain3 (bg3-D), which exhibits a remarkable increase of grain size caused by activation of the PURINE PERMEASE gene, OsPUP4. BG3/OsPUP4 is predominantly expressed in vascular tissues and is specifically suppressed by exogenous cytokinin application. Hormone profiling revealed that the distribution of different cytokinin forms, in roots and shoots of the bg3-D mutant, is altered. Quantitative reverse transcription-PCR (qRT-PCR) analysis indicated that expression of rice cytokinin type-A RESPONSE REGULATOR (OsRR) genes is enhanced in the roots of the bg3-D mutant. These results suggest that OsPUP4 might contribute to the long-distance transport of cytokinin, by reinforcing cytokinin loading into vascular bundle cells. Furthermore, plants overexpressing OsPUP7, the closest homolog of OsPUP4, also exhibited a similar phenotype to the bg3-D mutant. Interestingly, subcellular localization demonstrated that OsPUP4 was localized on the plasma membrane, whereas OsPUP7 was localized to the endoplasmic reticulum. Based on these findings, we propose that OsPUP4 and OsPUP7 function in a linear pathway to direct cytokinin cell-to-cell transport, affecting both its long-distance movement and local allocation.

A Temperature-Sensitive Misfolded bri1-301 Receptor Requires Its Kinase Activity to Promote Growth.

BRASSINOSTEROID-INSENSITIVE1 (BRI1) is a leucine-rich-repeat receptor-like kinase that functions as the cell surface receptor for brassinosteroids (BRs). Previous studies showed that BRI1 requires its kinase activity to transduce the extracellular BR signal into the nucleus. Among the many reported mutant bri1 alleles, bri1-301 is unique, as its glycine-989-to-isoleucine mutation completely inhibits its kinase activity in vitro but only gives rise to a weak dwarf phenotype compared with strong or null bri1 alleles, raising the question of whether kinase activity is essential for the biological function of BRI1. Here, we show that the Arabidopsis (Arabidopsis thaliana) bri1-301 mutant receptor exhibits weak BR-triggered phosphorylation in vivo and absolutely requires its kinase activity for the limited growth that occurs in the bri1-301 mutant. We also show that bri1-301 is a temperature-sensitive misfolded protein that is rapidly degraded in the endoplasmic reticulum and at the plasma membrane by yet unknown mechanisms. A temperature increase from 22°C to 29°C reduced the protein stability and biochemical activity of bri1-301, likely due to temperature-enhanced protein misfolding. The bri1-301 protein could be used as a model to study the degradation machinery for misfolded membrane proteins with cytosolic structural lesions and the plasma membrane-associated protein quality-control mechanism.

Correction to: OsbZIP71, a bZIP transcription factor, confers salinity and drought tolerance in rice.

Due to an error in combining the figure, an incorrect version of Fig. 9e was presented in the original publication.

EBS7 is a plant-specific component of a highly conserved endoplasmic reticulum-associated degradation system in Arabidopsis.

Endoplasmic reticulum (ER)-associated degradation (ERAD) is an essential part of an ER-localized protein quality-control system for eliminating terminally misfolded proteins. Recent studies have demonstrated that the ERAD machinery is conserved among yeast, animals, and plants; however, it remains unknown if the plant ERAD system involves plant-specific components. Here we report that the Arabidopsis ethyl methanesulfonate-mutagenized brassinosteroid-insensitive 1 suppressor 7 (EBS7) gene encodes an ER membrane-localized ERAD component that is highly conserved in land plants. Loss-of-function ebs7 mutations prevent ERAD of brassinosteroid insensitive 1-9 (bri1-9) and bri1-5, two ER-retained mutant variants of the cell-surface receptor for brassinosteroids (BRs). As a result, the two mutant receptors accumulate in the ER and consequently leak to the plasma membrane, resulting in the restoration of BR sensitivity and phenotypic suppression of the bri1-9 and bri1-5 mutants. EBS7 accumulates under ER stress, and its mutations lead to hypersensitivity to ER and salt stresses. EBS7 interacts with the ER membrane-anchored ubiquitin ligase Arabidopsis thaliana HMG-CoA reductase degradation 1a (AtHrd1a), one of the central components of the Arabidopsis ERAD machinery, and an ebs7 mutation destabilizes AtHrd1a to reduce polyubiquitination of bri1-9. Taken together, our results uncover a plant-specific component of a plant ERAD pathway and also suggest its likely biochemical function.

Activation of Big Grain1 significantly improves grain size by regulating auxin transport in rice.

Grain size is one of the key factors determining grain yield. However, it remains largely unknown how grain size is regulated by developmental signals. Here, we report the identification and characterization of a dominant mutant big grain1 (Bg1-D) that shows an extra-large grain phenotype from our rice T-DNA insertion population. Overexpression of BG1 leads to significantly increased grain size, and the severe lines exhibit obviously perturbed gravitropism. In addition, the mutant has increased sensitivities to both auxin and N-1-naphthylphthalamic acid, an auxin transport inhibitor, whereas knockdown of BG1 results in decreased sensitivities and smaller grains. Moreover, BG1 is specifically induced by auxin treatment, preferentially expresses in the vascular tissue of culms and young panicles, and encodes a novel membrane-localized protein, strongly suggesting its role in regulating auxin transport. Consistent with this finding, the mutant has increased auxin basipetal transport and altered auxin distribution, whereas the knockdown plants have decreased auxin transport. Manipulation of BG1 in both rice and Arabidopsis can enhance plant biomass, seed weight, and yield. Taking these data together, we identify a novel positive regulator of auxin response and transport in a crop plant and demonstrate its role in regulating grain size, thus illuminating a new strategy to improve plant productivity.

Variation in NRT1.1B contributes to nitrate-use divergence between rice subspecies.

Asian cultivated rice (Oryza sativa L.) consists of two main subspecies, indica and japonica. Indica has higher nitrate-absorption activity than japonica, but the molecular mechanisms underlying that activity remain elusive. Here we show that variation in a nitrate-transporter gene, NRT1.1B (OsNPF6.5), may contribute to this divergence in nitrate use. Phylogenetic analysis revealed that NRT1.1B diverges between indica and japonica. NRT1.1B-indica variation was associated with enhanced nitrate uptake and root-to-shoot transport and upregulated expression of nitrate-responsive genes. The selection signature of NRT1.1B-indica suggests that nitrate-use divergence occurred during rice domestication. Notably, field tests with near-isogenic and transgenic lines confirmed that the japonica variety carrying the NRT1.1B-indica allele had significantly improved grain yield and nitrogen-use efficiency (NUE) compared to the variety without that allele. Our results show that variation in NRT1.1B largely explains nitrate-use divergence between indica and japonica and that NRT1.1B-indica can potentially improve the NUE of japonica.

Control of grain size and rice yield by GL2-mediated brassinosteroid responses.

Given the continuously growing population and decreasing arable land, food shortage is becoming one of the most serious global problems in this century(1). Grain size is one of the determining factors for grain yield and thus is a prime target for genetic breeding(2,3). Although a number of quantitative trait loci (QTLs) associated with rice grain size have been identified in the past decade, mechanisms underlying their functions remain largely unknown(4,5). Here we show that a grain-length-associated QTL, GL2, has the potential to improve grain weight and grain yield up to 27.1% and 16.6%, respectively. We also show that GL2 is allelic to OsGRF4 and that it contains mutations in the miR396 targeting sequence. Because of the mutation, GL2 has a moderately increased expression level, which consequently activates brassinosteroid responses by upregulating a large number of brassinosteroid-induced genes to promote grain development. Furthermore, we found that GSK2, the central negative regulator of rice brassinosteroid signalling, directly interacts with OsGRF4 and inhibits its transcription activation activity to mediate the specific regulation of grain length by the hormone. Thus, this work demonstrates the feasibility of modulating specific brassinosteroid responses to improve plant productivity.

Brassinosteroid regulates cell elongation by modulating gibberellin metabolism in rice.

Brassinosteroid (BR) and gibberellin (GA) are two predominant hormones regulating plant cell elongation. A defect in either of these leads to reduced plant growth and dwarfism. However, their relationship remains unknown in rice (Oryza sativa). Here, we demonstrated that BR regulates cell elongation by modulating GA metabolism in rice. Under physiological conditions, BR promotes GA accumulation by regulating the expression of GA metabolic genes to stimulate cell elongation. BR greatly induces the expression of D18/GA3ox-2, one of the GA biosynthetic genes, leading to increased GA1 levels, the bioactive GA in rice seedlings. Consequently, both d18 and loss-of-function GA-signaling mutants have decreased BR sensitivity. When excessive active BR is applied, the hormone mostly induces GA inactivation through upregulation of the GA inactivation gene GA2ox-3 and also represses BR biosynthesis, resulting in decreased hormone levels and growth inhibition. As a feedback mechanism, GA extensively inhibits BR biosynthesis and the BR response. GA treatment decreases the enlarged leaf angles in plants with enhanced BR biosynthesis or signaling. Our results revealed a previously unknown mechanism underlying BR and GA crosstalk depending on tissues and hormone levels, which greatly advances our understanding of hormone actions in crop plants and appears much different from that in Arabidopsis thaliana.

OsNAP connects abscisic acid and leaf senescence by fine-tuning abscisic acid biosynthesis and directly targeting senescence-associated genes in rice.

It has long been established that premature leaf senescence negatively impacts the yield stability of rice, but the underlying molecular mechanism driving this relationship remains largely unknown. Here, we identified a dominant premature leaf senescence mutant, prematurely senile 1 (ps1-D). PS1 encodes a plant-specific NAC (no apical meristem, Arabidopsis ATAF1/2, and cup-shaped cotyledon2) transcriptional activator, Oryza sativa NAC-like, activated by apetala3/pistillata (OsNAP). Overexpression of OsNAP significantly promoted senescence, whereas knockdown of OsNAP produced a marked delay of senescence, confirming the role of this gene in the development of rice senescence. OsNAP expression was tightly linked with the onset of leaf senescence in an age-dependent manner. Similarly, ChIP-PCR and yeast one-hybrid assays demonstrated that OsNAP positively regulates leaf senescence by directly targeting genes related to chlorophyll degradation and nutrient transport and other genes associated with senescence, suggesting that OsNAP is an ideal marker of senescence onset in rice. Further analysis determined that OsNAP is induced specifically by abscisic acid (ABA), whereas its expression is repressed in both aba1 and aba2, two ABA biosynthetic mutants. Moreover, ABA content is reduced significantly in ps1-D mutants, indicating a feedback repression of OsNAP on ABA biosynthesis. Our data suggest that OsNAP serves as an important link between ABA and leaf senescence. Additionally, reduced OsNAP expression leads to delayed leaf senescence and an extended grain-filling period, resulting in a 6.3% and 10.3% increase in the grain yield of two independent representative RNAi lines, respectively. Thus, fine-tuning OsNAP expression should be a useful strategy for improving rice yield in the future.

OsbZIP71, a bZIP transcription factor, confers salinity and drought tolerance in rice.

The bZIP transcription factor (TF) family plays an important role in the abscisic acid (ABA) signaling pathway of abiotic stress in plants. We here report the cloning and characterization of OsbZIP71, which encodes a rice bZIP TF. Functional analysis showed that OsbZIP71 is a nuclear-localized protein that specifically binds to the G-box motif, but has no transcriptional activity both in yeast and rice protoplasts. In yeast two-hybrid assays, OsbZIP71 can form both homodimers and heterodimers with Group C members of the bZIP gene family. Expression of OsbZIP71 was strongly induced by drought, polyethylene glycol (PEG), and ABA treatments, but repressed by salt treatment. OsbZIP71 overexpressing (p35S::OsbZIP71) rice significantly improved tolerance to drought, salt and PEG osmotic stresses. In contrast, RNAi knockdown transgenic lines were much more sensitive to salt, PEG osmotic stresses, and also ABA treatment. Inducible expression (RD29A::OsbZIP71) lines were significantly improved their tolerance to PEG osmotic stresses, but hypersensitivity to salt, and insensitivity to ABA. Real-time PCR analysis revealed that the abiotic stress-related genes, OsVHA-B, OsNHX1, COR413-TM1, and OsMyb4, were up-regulated in overexpressing lines, while these same genes were down-regulated in RNAi lines. Chromatin immunoprecipitation analysis confirmed that OsbZIP71 directly binds the promoters of OsNHX1 and COR413-TM1 in vivo. These results suggest that OsbZIP71 may play an important role in ABA-mediated drought and salt tolerance in rice.