Yanru Hu

Jasmonate Regulates the INDUCER OF CBF EXPRESSION–C-REPEAT BINDING FACTOR/DRE BINDING FACTOR1 Cascade and Freezing Tolerance in Arabidopsis

This study reveals that jasmonate has a positive role in regulating freezing stress responses in Arabidopsis. JAZ proteins, the repressors of jasmonate signaling, physically interact with ICE1 and ICE2 transcription factors, thereby repressing the ICE-CBF/DREB1 cold signaling pathway. The INDUCER OF CBF EXPRESSION (ICE)–C-REPEAT BINDING FACTOR/DRE BINDING FACTOR1 (CBF/DREB1) transcriptional pathway plays a critical role in modulating cold stress responses in Arabidopsis thaliana. Dissecting crucial upstream regulatory signals or components of the ICE-CBF/DREB1 cascade will enhance our understanding of plant cold-tolerance mechanisms. Here, we show that jasmonate positively regulates plant responses to freezing stress in Arabidopsis. Exogenous application of jasmonate significantly enhanced plant freezing tolerance with or without cold acclimation. By contrast, blocking endogenous jasmonate biosynthesis and signaling rendered plants hypersensitive to freezing stress. Consistent with the positive role of jasmonate in freezing stress, production of endogenous jasmonate was triggered by cold treatment. In addition, cold induction of genes acting in the CBF/DREB1 signaling pathway was upregulated by jasmonate. Further investigation revealed that several JASMONATE ZIM-DOMAIN (JAZ) proteins, the repressors of jasmonate signaling, physically interact with ICE1 and ICE2 transcription factors. JAZ1 and JAZ4 repress the transcriptional function of ICE1, thereby attenuating the expression of its regulon. Consistent with this, overexpression of JAZ1 or JAZ4 represses freezing stress responses of Arabidopsis. Taken together, our study provides evidence that jasmonate functions as a critical upstream signal of the ICE-CBF/DREB1 pathway to positively regulate Arabidopsis freezing tolerance.

Arabidopsis WRKY46 coordinates with WRKY70 and WRKY53 in basal resistance against pathogen Pseudomonas syringae

The WRKY transcription factors are involved in plant resistance against both biotrophic and necrotrophic pathogens. Arabidopsis WRKY46 is specifically induced by salicylic acid (SA) and biotrophic pathogen Pseudomonas syringae infection. To determine its possible roles in plant defense and elucidate potential functional redundancy with structurally related WRKY70 and WRKY53, we examined loss-of-function T-DNA insertion single, double and triple mutants, as well as gain-of-function transgenic WRKY46 over-expressing plants in response to P. syringae. WRKY46 over-expressing plants were more resistant to P. syringae. In contrast, pathogen-infected wrky46wrky70, wrky46wrky53 double mutants and wrky46wrky70wrky53 triple mutants showed increased susceptibility to this pathogen, with increased bacterial growth and more severe disease symptoms. The contrasting responses of gain-of-function plants and loss-of-function mutants were correlated with increased or reduced expression of defense-related PR1 gene. Expression studies of WRKY46, WRKY70, and WRKY53 in various defense-signaling mutants suggested that they are partially involved in SA-signaling pathway. In addition, our findings demonstrated negative cross-regulation among these three genes. These results indicate that WRKY46, WRKY70, and WRKY53 positively regulate basal resistance to P. syringae; and that they play overlapping and synergetic roles in plant basal defense.

Arabidopsis transcription factor WRKY8 functions antagonistically with its interacting partner VQ9 to modulate salinity stress tolerance

The WRKY transcription factors have been demonstrated to play crucial roles in regulating stress responses; however, the exact mechanisms underlying their involvement in stress responses are not fully understood. Arabidopsis WRKY8 was predominantly expressed in roots and was highly upregulated by salt treatment. Disruption of WRKY8 rendered plants hypersensitive to salt, showing delayed germination, inhibited post-germination development and accelerated chlorosis. Further investigation revealed that WRKY8 interacted with VQ9, and their interaction decreased the DNA-binding activity of WRKY8. The VQ9 protein was exclusively localized in the nucleus, and VQ9 expression was strongly responsive to NaCl treatment. Mutation of VQ9 enhanced tolerance to salt stress, indicating that VQ9 acts antagonistically with WRKY8 to mediate responses to salt stress. The antagonist functions of WRKY8 and VQ9 were consistent with an increased or reduced Na⁺/K⁺ concentration ratio, as well as contrasting expression patterns of downstream stress-responsive genes in salt-stressed wrky8 and vq9 mutants. Moreover, chromatin immunoprecipitation (ChIP) assays showed that WRKY8 directly bound the promoter of RD29A under salt conditions. These results provided strong evidence that the VQ9 protein acts as a repressor of the WRKY8 factor to maintain an appropriate balance of WRKY8-mediated signaling pathways to establish salinity stress tolerance.

BRASSINOSTEROID INSENSITIVE2 Interacts with ABSCISIC ACID INSENSITIVE5 to Mediate the Antagonism of Brassinosteroids to Abscisic Acid during Seed Germination in Arabidopsis

A kinase regulates the integration of brassinosteroid and abscisic acid signaling during seed germination by phosphorylating and stabilizing an abscisic acid-responsive transcription factor. Seed germination and postgerminative growth are regulated by a delicate hormonal balance. Abscisic acid (ABA) represses Arabidopsis thaliana seed germination and postgerminative growth, while brassinosteroids (BRs) antagonize ABA-mediated inhibition and promote these processes. However, the molecular mechanism underlying BR-repressed ABA signaling remains largely unknown. Here, we show that the Glycogen Synthase Kinase 3-like kinase BRASSINOSTEROID INSENSITIVE2 (BIN2), a critical repressor of BR signaling, positively regulates ABA responses during seed germination and postgerminative growth. Mechanistic investigation revealed that BIN2 physically interacts with ABSCISIC ACID INSENSITIVE5 (ABI5), a bZIP transcription factor. Further genetic analysis demonstrated that the ABA-hypersensitive phenotype of BIN2-overexpressing plants requires ABI5. BIN2 was found to phosphorylate and stabilize ABI5 in the presence of ABA, while application of epibrassinolide (the active form of BRs) inhibited the regulation of ABI5 by BIN2. Consistently, the ABA-induced accumulation of ABI5 was affected in BIN2-related mutants. Moreover, mutations of the BIN2 phosphorylation sites on ABI5 made the mutant protein respond to ABA improperly. Additionally, the expression of several ABI5 regulons was positively modulated by BIN2. These results provide evidence that BIN2 phosphorylates and stabilizes ABI5 to mediate ABA response during seed germination, while BRs repress the BIN2-ABI5 cascade to antagonize ABA-mediated inhibition.

Jasmonate regulates leaf senescence and tolerance to cold stress: crosstalk with other phytohormones.

Plants are challenged with numerous abiotic stresses, such as drought, cold, heat, and salt stress. These environmental stresses are major causes of crop failure and reduced yields worldwide. Phytohormones play essential roles in regulating various plant physiological processes and alleviating stressful perturbations. Jasmonate (JA), a group of oxylipin compounds ubiquitous in the plant kingdom, acts as a crucial signal to modulate multiple plant processes. Recent studies have shown evidence supporting the involvement of JA in leaf senescence and tolerance to cold stress. Concentrations of JA are much higher in senescent leaves compared with those in non-senescent ones. Treatment with exogenous JA induces leaf senescence and expression of senescence-associated genes. In response to cold stress, exogenous application of JA enhances Arabidopsis freezing tolerance with or without cold acclimation. Consistently, biosynthesis of endogenous JA is activated in response to cold exposure. JA positively regulates the CBF (C-REPEAT BINDING FACTOR) transcriptional pathway to up-regulate downstream cold-responsive genes and ultimately improve cold tolerance. JA interacts with other hormone signaling pathways (such as auxin, ethylene, and gibberellin) to regulate leaf senescence and tolerance to cold stress. In this review, we summarize recent studies that have provided insights into JA-mediated leaf senescence and cold-stress tolerance.

The DELLA-CONSTANS Transcription Factor Cascade Integrates Gibberellic Acid and Photoperiod Signaling to Regulate Flowering

DELLA proteins physically and genetically interact with CO to modulate flowering under long-days in Arabidopsis. Gibberellin (GA) and photoperiod pathways have recently been demonstrated to collaboratively modulate flowering under long days (LDs). However, the molecular mechanisms underlying this collaboration remain largely unclear. In this study, we found that GA-induced expression of FLOWERING LOCUS T (FT) under LDs was dependent on CONSTANS (CO), a critical transcription factor positively involved in photoperiod signaling. Mechanistic investigation revealed that DELLA proteins, a group of crucial repressors in GA signaling, physically interacted with CO. The DELLA-CO interactions repressed the transcriptional function of CO protein. Genetic analysis demonstrated that CO acts downstream of DELLA proteins to regulate flowering. Disruption of CO rescued the earlier flowering phenotype of the gai-t6 rga-t2 rgl1-1 rgl2-1 mutant (dellap), while a gain-of-function mutation in GA INSENSITIVE (GAI, a member of the DELLA gene) repressed the earlier flowering phenotype of CO-overexpressing plants. In addition, the accumulation of DELLA proteins and mRNAs was rhythmic, and REPRESSOR OF GA1-3 protein was noticeably decreased in the long-day afternoon, a time when CO protein is abundant. Collectively, these results demonstrate that the DELLA-CO cascade inhibits CO/FT-mediated flowering under LDs, which thus provide evidence to directly integrate GA and photoperiod signaling to synergistically modulate flowering under LDs.

Heterologous Expression of AtWRKY57 Confers Drought Tolerance in Oryza sativa

Drought stress is a severe environmental factor that greatly restricts plant distribution and crop production. Recently, we have found that overexpressing AtWRKY57 enhanced drought tolerance in Arabidopsis thaliana. In this study, we further reported that the Arabidopsis WRKY57 transcription factor was able to confer drought tolerance to transgenic rice (Oryza sativa) plants. The enhanced drought tolerance of transgenic rice was resulted from the lower water loss rates, cell death, malondialdehyde contents and relative electrolyte leakage while a higher proline content and reactive oxygen species-scavenging enzyme activities was observed during stress conditions. Moreover, further investigation revealed that the expression levels of several stress-responsive genes were up-regulated in drought-tolerant transgenic rice plants, compared with those in wild-type plants. In addition to the drought tolerance, the AtWRKY57 over-expressing plants also had enhanced salt and PEG stress tolerances. Taken together, our study indicates that over-expressing AtWRKY57 in rice improved not only drought tolerance but also salt and PEG tolerance, demonstrating its potential role in crop improvement.

Arabidopsis VQ motif-containing proteins VQ12 and VQ29 negatively modulate basal defense against Botrytis cinerea

Arabidopsis VQ motif-containing proteins have recently been demonstrated to interact with several WRKY transcription factors; however, their specific biological functions and the molecular mechanisms underlying their involvement in defense responses remain largely unclear. Here, we showed that two VQ genes, VQ12 and VQ29, were highly responsive to the necrotrophic fungal pathogen Botrytis cinerea. To characterize their roles in plant defense, we generated amiR-vq12 transgenic plants by using an artificial miRNA approach to suppress the expression of VQ12, and isolated a loss-of-function mutant of VQ29. Phenotypic analysis showed that decreasing the expression of VQ12 and VQ29 simultaneously rendered the amiR-vq12 vq29 double mutant plants resistant against B. cinerea. Consistently, the B. cinerea-induced expression of defense-related PLANT DEFENSIN1.2 (PDF1.2) was increased in amiR-vq12 vq29. In contrast, constitutively-expressing VQ12 or VQ29 confered transgenic plants susceptible to B. cinerea. Further investigation revealed that VQ12 and VQ29 physically interacted with themselves and each other to form homodimers and heterodimer. Moreover, expression analysis of VQ12 and VQ29 in defense-signaling mutants suggested that they were partially involved in jasmonate (JA)-signaling pathway. Taken together, our study indicates that VQ12 and VQ29 negatively regulate plant basal resistance against B. cinerea.

The bHLH Transcription Factors MYC2, MYC3, and MYC4 Are Required for Jasmonate-Mediated Inhibition of Flowering in Arabidopsis

In plants, the floral transition is flexibly controlled by various environmental conditions and endogenous developmental cues. In Arabidopsis, six major flowering pathways respond to changes in these factors (Fornara et al., 2010). The photoperiod, vernalization, and ambient pathways monitor exogenous signals from the environment such as day length, minimum winter temperature, and ambient temperature (Fornara et al., 2010). By contrast, the autonomous, gibberellin, and age pathways respond to endogenous cues linked to developmental status (Fornara et al., 2010). Accumulating evidence indicates that the six flowering pathways converge in a network to regulate floral integrator genes FLOWERING LOCUS T (FT), TWIN SISTER OF FT (TSF), and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 (Fornara et al., 2010).

Jasmonate negatively regulates stomatal development in Arabidopsis cotyledons

Jasmonate and MYC transcription factors negatively regulate stomatal development in Arabidopsis cotyledons. Stomata are ports that facilitate gas and water vapor exchange between plants and their environment. Stomatal development is strictly regulated by endogenous signals and environmental cues. Jasmonate is an important signal that modulates multiple physiological processes in plants, yet the molecular mechanisms underlying its interactions with other developmental signaling pathways remain poorly understood. Here, we show that jasmonate negatively regulates stomatal development in Arabidopsis (Arabidopsis thaliana) cotyledons. Cotyledons of the wild type and stomata-overproliferating mutants (such as too many mouths-1 and stomatal density and distribution1-1) treated with methyl jasmonate exhibit a clear reduction in stomata number. By contrast, blocking endogenous jasmonate biosynthesis or perception enhanced stomatal development. Moreover, three MYC transcription factors involved in jasmonate signaling, MYC2, MYC3, and MYC4, were found to redundantly modulate jasmonate-inhibited stomatal development. A genetic analysis showed that these MYC proteins act upstream of the SPEECHLESS and FAMA transcription factors to mediate stomatal development. Furthermore, jasmonate repression of stomatal development is dependent on these three MYC transcription factors, as stomatal development of the myc2 myc3 myc4 triple mutant was insensitive to methyl jasmonate treatment. Collectively, our study demonstrates that jasmonate and MYC transcription factors negatively regulate stomatal development in Arabidopsis cotyledons.