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Qingxi J Shen – One of the best experts on this subject based on the ideXlab platform.

  • Transcriptomic analysis of rice Aleurone Cells identified a novel abscisic acid response element.
    Plant cell & environment, 2017
    Co-Authors: Kenneth A. Watanabe, Arielle Homayouni, Kuan‐ying Huang, Qingxi J Shen
    Abstract:

    Seeds serve as a great model to study plant responses to drought stress, which is largely mediated by abscisic acid (ABA). The ABA responsive element (ABRE) is a key cis-regulatory element in ABA signaling. However, its consensus sequence, (ACGTG(G/T)C), is present in the promoters of only about 40% of ABA induced genes in rice Aleurone Cells, suggesting other ABREs may exist. To identify novel ABREs, RNA-sequencing was performed on Aleurone Cells of rice seeds treated with 20 μM ABA. Gibbs sampling was used to identify enriched elements and particle bombardment-mediated transient expression studies were performed to verify the function. Gene ontology analysis was performed to predict the roles of genes containing the novel ABREs. This study revealed 2,443 ABA-inducible genes and a novel ABRE, designated as ABREN, which was experimentally verified to mediate ABA signaling in rice Aleurone Cells. Many of the ABREN-containing genes are predicted to be involved in stress responses and transcription. Analysis of other species suggest that the ABREN may be monocot-specific. This study also revealed interesting expression patterns of genes involved in ABA metabolism and signaling. Collectively, this study advanced our understanding of diverse cis-regulatory sequences and the transcriptomes underlying ABA responses in rice Aleurone Cells.

  • rna sequencing reveals previously unannotated protein and microrna coding genes expressed in Aleurone Cells of rice seeds
    Genomics, 2014
    Co-Authors: Kenneth A. Watanabe, Patricia Ringler, Qingxi J Shen
    Abstract:

    The rice genome annotation has been greatly improved in recent years, largely due to the availability of full length cDNA sequences derived from many tissues. Among those yet to be studied is the Aleurone layer, which produces hydrolases for mobilization of seed storage reserves during seed germgermination and post germination growth. Herein, we report transcriptomes of Aleurone Cells treated with the hormones abscisic acid, gibberellic acid, or both. Using a comprehensive approach, we identified hundreds of novel genes. To minimize the number of false positives, only transcripts that did not overlap with existing annotations, had a high level of expression, and showed a high level of uniqueness within the rice genome were considered to be novel genes. This approach led to the identification of 553 novel genes that encode proteins and/or microRNAs. The transcriptome data reported here will help to further improve the annotation of the rice genome.

  • a negative regulator encoded by a rice wrky gene represses both abscisic acid and gibberellins signaling in Aleurone Cells
    Plant Molecular Biology, 2009
    Co-Authors: Zhonglin Zhang, Xiaolu Zou, Margaret Shin, Jianzhi Huang, Qingxi J Shen
    Abstract:

    Abscisic acid (ABA) and gibberellins (GAs) control several developmental processes including seed maturation, dormancy, and germination. The antagonism of these two hormones is well-documented. However, recent data from transcription profiling studies indicate that they can function as agonists in regulating the expression of many genes although the underlying mechanism is unclear. Here we report a rice WRKY gene, OsWRKY24, which encodes a protein that functions as a negative regulator of both GA and ABA signaling. Overexpression of OsWRKY24 via particle bombardment-mediated transient expression in Aleurone Cells represses the expression of two reporter constructs: the β-glucuronidase gene driven by the GA-inducible Amy32b α-amylase promoter (Amy32b-GUS) and the ABA-inducible HVA22 promoter (HVA22-GUS). OsWRKY24 is unlikely a general repressor because it has little effect on the expression of the luciferase reporter gene driven by a constitutive ubiquitin promoter (UBI-Luciferase). As to the GA signaling, OsWRKY24 differs from OsWRKY51 and −71, two negative regulators specifically function in the GA signaling pathway, in several ways. First, OsWRKY24 contains two WRKY domains while OsWRKY51 and −71 have only one; both WRKY domains are essential for the full repressing activity of OsWRKY24. Second, binding of OsWRKY24 to the Amy32b promoter appears to involve sequences in addition to the TGAC cores of the W-boxes. Third, unlike OsWRKY71, OsWRKY24 is stable upon GA treatment. Together, these data demonstrate that OsWRKY24 is a novel type of transcriptional repressor that inhibits both GA and ABA signaling.

Zhonglin Zhang – One of the best experts on this subject based on the ideXlab platform.

  • a negative regulator encoded by a rice wrky gene represses both abscisic acid and gibberellins signaling in Aleurone Cells
    Plant Molecular Biology, 2009
    Co-Authors: Zhonglin Zhang, Xiaolu Zou, Margaret Shin, Jianzhi Huang, Qingxi J Shen
    Abstract:

    Abscisic acid (ABA) and gibberellins (GAs) control several developmental processes including seed maturation, dormancy, and germination. The antagonism of these two hormones is well-documented. However, recent data from transcription profiling studies indicate that they can function as agonists in regulating the expression of many genes although the underlying mechanism is unclear. Here we report a rice WRKY gene, OsWRKY24, which encodes a protein that functions as a negative regulator of both GA and ABA signaling. Overexpression of OsWRKY24 via particle bombardment-mediated transient expression in Aleurone Cells represses the expression of two reporter constructs: the β-glucuronidase gene driven by the GA-inducible Amy32b α-amylase promoter (Amy32b-GUS) and the ABA-inducible HVA22 promoter (HVA22-GUS). OsWRKY24 is unlikely a general repressor because it has little effect on the expression of the luciferase reporter gene driven by a constitutive ubiquitin promoter (UBI-Luciferase). As to the GA signaling, OsWRKY24 differs from OsWRKY51 and −71, two negative regulators specifically function in the GA signaling pathway, in several ways. First, OsWRKY24 contains two WRKY domains while OsWRKY51 and −71 have only one; both WRKY domains are essential for the full repressing activity of OsWRKY24. Second, binding of OsWRKY24 to the Amy32b promoter appears to involve sequences in addition to the TGAC cores of the W-boxes. Third, unlike OsWRKY71, OsWRKY24 is stable upon GA treatment. Together, these data demonstrate that OsWRKY24 is a novel type of transcriptional repressor that inhibits both GA and ABA signaling.

  • interactions of two abscisic acid induced wrky genes in repressing gibberellin signaling in Aleurone Cells
    Plant Journal, 2006
    Co-Authors: Zhen Xie, Zhonglin Zhang, Xiaolu Zou, Guangxiao Yang, Setsuko Komatsu, Qingxi J Shen
    Abstract:

    Gibberellins (GA) promote while abscisic acid (ABA) inhibits seed germgermination and post-germination growth. To address the cross-talk of GA and ABA signaling, we studied two rice WRKY genes (OsWRKY51 and OsWRKY71) that are ABA-inducible and GA-repressible in embryos and Aleurone Cells. Over-expression of these two genes in Aleurone Cells specifically and synergistically represses induction of the ABA-repressible and GA-inducible Amy32b alpha-amylamylase promoter reporter construct (Amy32b-GUS) by GA or the GA-inducible transcriptional actiactivator, GAMYB. The physical interactions of OsWRKY71 proteins themselves and that of OsWRKY71 and OsWRKY51 are revealed in the nuclei of Aleurone Cells using bimolecular fluorescence complementation (BiFC) assays. Although OsWRKY51 itself does not bind to the Amy32b promoter in vitro, it interacts with OsWRKY71 and enhances the binding affinity of OsWRKY71 to W boxes in the Amy32b promoter. The binding activity of OsWRKY71 is abolished by deleting the C-terminus containing the WRKY domain or substituting the key amino acids in the WRKY motif and the zinc finger region. However, two of these non-DNA-binding mutants are still able to repress GA induction by enhancing the binding affinity of the wild-type DNA-binding OsWRKY71 repressors. In contrast, the third non-DNA-binding mutant enhances GA induction of Amy32b-GUS, by interfering with the binding of the wild-type OsWRKY71 or the OsWRKY71/OsWRKY51 repressing complex. These data demonstrate the synergistic interaction of ABA-inducible WRKY genes in regulating GAMYB-mediated GA signaling in Aleurone Cells, thereby establishing a novel mechanism for ABA and GA signaling cross-talk.

  • Interactions of two abscisic‐acid induced WRKY genes in repressing gibberellin signaling in Aleurone Cells
    The Plant journal : for cell and molecular biology, 2006
    Co-Authors: Zhen Xie, Zhonglin Zhang, Xiaolu Zou, Guangxiao Yang, Setsuko Komatsu, Qingxi J Shen
    Abstract:

    Gibberellins (GA) promote while abscisic acid (ABA) inhibits seed germgermination and post-germination growth. To address the cross-talk of GA and ABA signaling, we studied two rice WRKY genes (OsWRKY51 and OsWRKY71) that are ABA-inducible and GA-repressible in embryos and Aleurone Cells. Over-expression of these two genes in Aleurone Cells specifically and synergistically represses induction of the ABA-repressible and GA-inducible Amy32b alpha-amylamylase promoter reporter construct (Amy32b-GUS) by GA or the GA-inducible transcriptional actiactivator, GAMYB. The physical interactions of OsWRKY71 proteins themselves and that of OsWRKY71 and OsWRKY51 are revealed in the nuclei of Aleurone Cells using bimolecular fluorescence complementation (BiFC) assays. Although OsWRKY51 itself does not bind to the Amy32b promoter in vitro, it interacts with OsWRKY71 and enhances the binding affinity of OsWRKY71 to W boxes in the Amy32b promoter. The binding activity of OsWRKY71 is abolished by deleting the C-terminus containing the WRKY domain or substituting the key amino acids in the WRKY motif and the zinc finger region. However, two of these non-DNA-binding mutants are still able to repress GA induction by enhancing the binding affinity of the wild-type DNA-binding OsWRKY71 repressors. In contrast, the third non-DNA-binding mutant enhances GA induction of Amy32b-GUS, by interfering with the binding of the wild-type OsWRKY71 or the OsWRKY71/OsWRKY51 repressing complex. These data demonstrate the synergistic interaction of ABA-inducible WRKY genes in regulating GAMYB-mediated GA signaling in Aleurone Cells, thereby establishing a novel mechanism for ABA and GA signaling cross-talk.

Frank Gubler – One of the best experts on this subject based on the ideXlab platform.

Xiaolu Zou – One of the best experts on this subject based on the ideXlab platform.

  • a negative regulator encoded by a rice wrky gene represses both abscisic acid and gibberellins signaling in Aleurone Cells
    Plant Molecular Biology, 2009
    Co-Authors: Zhonglin Zhang, Xiaolu Zou, Margaret Shin, Jianzhi Huang, Qingxi J Shen
    Abstract:

    Abscisic acid (ABA) and gibberellins (GAs) control several developmental processes including seed maturation, dormancy, and germination. The antagonism of these two hormones is well-documented. However, recent data from transcription profiling studies indicate that they can function as agonists in regulating the expression of many genes although the underlying mechanism is unclear. Here we report a rice WRKY gene, OsWRKY24, which encodes a protein that functions as a negative regulator of both GA and ABA signaling. Overexpression of OsWRKY24 via particle bombardment-mediated transient expression in Aleurone Cells represses the expression of two reporter constructs: the β-glucuronidase gene driven by the GA-inducible Amy32b α-amylase promoter (Amy32b-GUS) and the ABA-inducible HVA22 promoter (HVA22-GUS). OsWRKY24 is unlikely a general repressor because it has little effect on the expression of the luciferase reporter gene driven by a constitutive ubiquitin promoter (UBI-Luciferase). As to the GA signaling, OsWRKY24 differs from OsWRKY51 and −71, two negative regulators specifically function in the GA signaling pathway, in several ways. First, OsWRKY24 contains two WRKY domains while OsWRKY51 and −71 have only one; both WRKY domains are essential for the full repressing activity of OsWRKY24. Second, binding of OsWRKY24 to the Amy32b promoter appears to involve sequences in addition to the TGAC cores of the W-boxes. Third, unlike OsWRKY71, OsWRKY24 is stable upon GA treatment. Together, these data demonstrate that OsWRKY24 is a novel type of transcriptional repressor that inhibits both GA and ABA signaling.

  • interactions of two transcriptional repressors and two transcriptional activators in modulating gibberellin signaling in Aleurone Cells
    Plant Physiology, 2008
    Co-Authors: Xiaolu Zou, Dawn S Neuman, Qingxi J Shen
    Abstract:

    Gibberellins (GAs) regulate many aspects of plant development, such as germination, growth, and flowering. The barley (Hordeum vulgare) Amy32b α-amylase promoter contains at least five cis-acting elements that govern its GA-induced expression. Our previous studies indicate that a barley WRKY gene, HvWRKY38, and its rice (Oryza sativa) ortholog, OsWRKY71, block GA-induced expression of Amy32b-GUS. In this work, we investigated the functional and physical interactions of HvWRKY38 with another repressor and two activators in barley. HvWRKY38 blocks the inductive activities of SAD (a DOF protein) and HvGAMYB (a R2R3 MYB protein) when either of these proteins is present individually. However, SAD and HvGAMYB together overcome the inhibitory effect of HvWRKY38. Yet, the combination of HvWRKY38 and BPBF (another DOF protein) almost diminishes the synergistic effect of SAD and HvGAMYB transcriptional activators. Electrophoretic mobility shift assays indicate that HvWRKY38 blocks the GA-induced expression of Amy32b by interfering with the binding of HvGAMYB to the cis-acting elements in the α-amylase promoter. The physical interaction of HvWRKY38 and BPBF repressors is demonstrated via bimolecular fluorescence complementation assays. These data suggest that the expression of Amy32b is modulated by protein complexes that contain either activators (e.g. HvGAMYB and SAD) or repressors (e.g. HvWRKY38 and BPBF). The relative amounts of the repressor or activator complexes binding to the Amy32b promoter regulate its expression level in barley Aleurone Cells.

  • interactions of two abscisic acid induced wrky genes in repressing gibberellin signaling in Aleurone Cells
    Plant Journal, 2006
    Co-Authors: Zhen Xie, Zhonglin Zhang, Xiaolu Zou, Guangxiao Yang, Setsuko Komatsu, Qingxi J Shen
    Abstract:

    Gibberellins (GA) promote while abscisic acid (ABA) inhibits seed germination and post-germination growth. To address the cross-talk of GA and ABA signaling, we studied two rice WRKY genes (OsWRKY51 and OsWRKY71) that are ABA-inducible and GA-repressible in embryos and Aleurone Cells. Over-expression of these two genes in Aleurone Cells specifically and synergistically represses induction of the ABA-repressible and GA-inducible Amy32b alpha-amylase promoter reporter construct (Amy32b-GUS) by GA or the GA-inducible transcriptional activator, GAMYB. The physical interactions of OsWRKY71 proteins themselves and that of OsWRKY71 and OsWRKY51 are revealed in the nuclei of Aleurone Cells using bimolecular fluorescence complementation (BiFC) assays. Although OsWRKY51 itself does not bind to the Amy32b promoter in vitro, it interacts with OsWRKY71 and enhances the binding affinity of OsWRKY71 to W boxes in the Amy32b promoter. The binding activity of OsWRKY71 is abolished by deleting the C-terminus containing the WRKY domain or substituting the key amino acids in the WRKY motif and the zinc finger region. However, two of these non-DNA-binding mutants are still able to repress GA induction by enhancing the binding affinity of the wild-type DNA-binding OsWRKY71 repressors. In contrast, the third non-DNA-binding mutant enhances GA induction of Amy32b-GUS, by interfering with the binding of the wild-type OsWRKY71 or the OsWRKY71/OsWRKY51 repressing complex. These data demonstrate the synergistic interaction of ABA-inducible WRKY genes in regulating GAMYB-mediated GA signaling in Aleurone Cells, thereby establishing a novel mechanism for ABA and GA signaling cross-talk.

Russell L. Jones – One of the best experts on this subject based on the ideXlab platform.

  • cprg hcl a potential h cl symporter prevents acidification of storage vacuoles in Aleurone Cells and inhibits ga dependent hydrolysis of storage protein and phytate
    Plant Journal, 2003
    Co-Authors: Yong-sic Hwang, Paul C. Bethke, Frank Gubler, Russell L. Jones
    Abstract:

    Summary The putative H+/Cl− symporter cycloprodigiosin-HCl (cPrG-HCl) was used to investigate the role of vacuole acidification in cereal Aleurone cell function. The protein storage vacuole (PSV) becomes acidified rapidly when Aleurone Cells are treated with gibberellic acid (GA) but not abscisic acid (ABA). We show that cPrG prevents PSV acidification in Aleurone layers and prevents synthesis of secretory proteins such as α-amylase. Our data support the hypothesis that decreased hydrolase synthesis is a consequence of decreased hydrolysis of storage proteins in PSV. Support for this hypothesis comes from experiments showing that breakdown of barley 7S globulins and phytate is inhibited by cPrG in GA-treated Aleurone layers. Decreased mobilization of PSV reserves is accompanied by reductions in the free amino acid pool size and in the amount of ions released from the Aleurone layer. Vacuolation of the Aleurone cell is a diagnostic feature of the response to GA, and vacuolation is also inhibited by cPrG. Evidence that cPrG acts as a potential H+/Cl− symporter in Aleurone is presented. We show that cPrG does not inhibit the synthesis and secretion of α-amylase when Cl− ions are omitted from the incubation medium. Although cPrG blocks many GA-induced responses of Aleurone layers, it does not affect early steps in GA signaling. The SLN1 protein, a negative regulator of GA signaling, is turned over in GA-treated Cells in the presence and absence of cPrG. Similarly, synthesis of the transcriptional activator GAMYB is unaffected by the presence of cPrG in GA-treated Cells.

  • cPrG-HCl a potential H+/Cl- symporter prevents acidification of storage vacuoles in Aleurone Cells and inhibits GA-dependent hydrolysis of storage protein and phytate.
    The Plant journal : for cell and molecular biology, 2003
    Co-Authors: Yong-sic Hwang, Paul C. Bethke, Frank Gubler, Russell L. Jones
    Abstract:

    Summary The putative H+/Cl− symporter cycloprodigiosin-HCl (cPrG-HCl) was used to investigate the role of vacuole acidification in cereal Aleurone cell function. The protein storage vacuole (PSV) becomes acidified rapidly when Aleurone Cells are treated with gibberellic acid (GA) but not abscisic acid (ABA). We show that cPrG prevents PSV acidification in Aleurone layers and prevents synthesis of secretory proteins such as α-amylase. Our data support the hypothesis that decreased hydrolase synthesis is a consequence of decreased hydrolysis of storage proteins in PSV. Support for this hypothesis comes from experiments showing that breakdown of barley 7S globulins and phytate is inhibited by cPrG in GA-treated Aleurone layers. Decreased mobilization of PSV reserves is accompanied by reductions in the free amino acid pool size and in the amount of ions released from the Aleurone layer. Vacuolation of the Aleurone cell is a diagnostic feature of the response to GA, and vacuolation is also inhibited by cPrG. Evidence that cPrG acts as a potential H+/Cl− symporter in Aleurone is presented. We show that cPrG does not inhibit the synthesis and secretion of α-amylase when Cl− ions are omitted from the incubation medium. Although cPrG blocks many GA-induced responses of Aleurone layers, it does not affect early steps in GA signaling. The SLN1 protein, a negative regulator of GA signaling, is turned over in GA-treated Cells in the presence and absence of cPrG. Similarly, synthesis of the transcriptional activator GAMYB is unaffected by the presence of cPrG in GA-treated Cells.

  • Active oxygen and cell death in cereal Aleurone Cells.
    Journal of experimental botany, 2002
    Co-Authors: Angelika Fath, Paul C. Bethke, Veronica Beligni, Russell L. Jones
    Abstract:

    The cereal Aleurone layer is a secretory tissue whose function is regulated by gibberellic acid (GA) and abscisic acid (ABA). Aleurone Cells lack functional chloroplasts, thus excluding photosynthesis as a source of active oxygen species (AOS) in cell death. Incubation of barley Aleurone layers or protoplasts in GA initiated the cell death programme, but incubation in ABA delays programmed cell death (PCD). Light, especially blue and UV-A light, and H 2 O 2 accelerate PCD of GA-treated Aleurone Cells, but ABA-treated Aleurone Cells are refractory to light and H 2 O 2 and are not killed. It was shown that light elevated intracellular H 2 O 2 , and that the rise in H 2 O 2 was greater in GA-treated Cells compared to Cells in ABA. Experiments with antioxidants show that PCD in Aleurone is probably regulated by AOS. The sensitivity of GA-treated Aleurone to light and H 2 O 2 is a result of lowered amounts of enzymes that metabolize AOS. mRNAs encoding catalase, ascorbate peroxidase and superoxide dismutase are all reduced during 6-18 h of incubation in GA, but these mRNAs were present in higher amounts in Cells incubated in ABA. The amounts of protein and enzyme activities encoded by these mRNAs were also dramatically reduced in GA-treated Cells. Aleurone Cells store and metabolize neutral lipids via the glyoxylate cycle in response to GA, and glyoxysomes are one potential source of AOS in the GA-treated Cells. Mitochondria are another potential source of AOS in GA-treated Cells. AOS generated by these organelles bring about membrane rupture and cell death.