6 research outputs found

    RNAi<b>-</b>Directed Downregulation of Vacuolar H<sup>+</sup><b>-</b>ATPase Subunit A Results in Enhanced Stomatal Aperture and Density in Rice

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    <div><p>Stomatal movement plays a key role in plant development and response to drought and salt stress by regulating gas exchange and water loss. A number of genes have been demonstrated to be involved in the regulation of this process. Using inverse genetics approach, we characterized the function of a rice (<i>Oryza sativa</i> L.) vacuolar H<sup>+</sup>-ATPase subunit A (<i>OsVHA-A</i>) gene in stomatal conductance regulation and physiological response to salt and osmotic stress. <i>OsVHA-A</i> was constitutively expressed in different rice tissues, and the fusion protein of GFP-OsVHA-A was exclusively targeted to tonoplast when transiently expressed in the onion epidermal cells. Heterologous expression of <i>OsVHA-A</i> was able to rescue the yeast mutant <i>vma1Δ</i> (lacking subunit A activity) phenotype, suggesting that it partially restores the activity of V-ATPase. Meanwhile, RNAi-directed knockdown of <i>OsVHA-A</i> led to a reduction of vacuolar H<sup>+</sup>-ATPase activity and an enhancement of plasma membrane H<sup>+</sup>-ATPase activity, thereby increasing the concentrations of extracellular H<sup>+</sup> and intracellular K<sup>+</sup> and Na<sup>+</sup> under stress conditions. Knockdown of <i>OsVHA-A</i> also resulted in the upregulation of <i>PAM3</i> (<i>plasma membrane H<sup>+</sup>-ATPase 3</i>) and downregulation of <i>CAM1</i> (<i>calmodulin 1</i>), <i>CAM3 (calmodulin 3</i>) and <i>YDA1</i> (<i>YODA</i>, a MAPKK gene). Altered level of the ion concentration and the gene expression by knockdown of <i>OsVHA-A</i> probably resulted in expanded aperture of stomatal pores and increased stomatal density. In addition, <i>OsVHA-A</i> RNAi plants displayed significant growth inhibition under salt and osmotic stress conditions. Taken together, our results suggest that <i>OsVHA-A</i> takes part in regulating stomatal density and opening via interfering with pH value and ionic equilibrium in guard cells and thereby affects the growth of rice plants.</p></div

    V-ATPase, PPase activity assays and vacuolar pH measurements in <i>OsVHA</i><i>-</i><i>A</i> RNAi transgenic lines.

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    <p>(A) Vacuolar H<sup>+</sup>-ATPase activity and (B) V-PPase activity were determined in wild type (WT) and three <i>OsVHA-A</i> RNA interference lines (<i>OsV-5</i>, <i>OsV-11</i>, and <i>OsV-18</i>). (C) The images showing emission intensities of vacuoles from epidermal root cells loaded with BCECF AM. Results shown are representative. Scale bars  = 100 μM. (D) The vacuolar pH values calculated from (C). (E) V-ATPase proton-pumping measured by the quenching of ACMA fluorescence. Ten micrograms of tonoplast vesicles were applied to detect fluorescence density. Each bar represents three replications. Asterisks (*) indicate significant differences from WT at <i>P</i><0.05.</p

    DataSheet1_Negatively Regulated by miR-29c-3p, MTFR1 Promotes the Progression and Glycolysis in Lung Adenocarcinoma via the AMPK/mTOR Signalling Pathway.PDF

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    Background: Lung adenocarcinoma (LUAD) is the major form of lung cancer that presents a major peril to public health. Owing to the high rates of morbidity, mortality and chemoresistance, it is necessary to develop more effective therapeutic targets of LUAD. Mitochondrial fission regulator 1 (MTFR1) affects the occurrence and development of some diseases by regulating mitochondrial dynamics and is dysregulated in LUAD. However, the functions and molecular mechanisms of MTFR1 in LUAD have not been investigated.Methods: Immunohistochemical (IHC) analysis, real-time quantitative polymerase chain reaction (RT-qPCR), bioinformatic analysis and western blot (WB) were performed to assess the expression of MTFR1 at both protein and mRNA levels. The biological functions of MTFR1 in LUAD cells were assessed based on various in vivo and in vitro experiments. The dual-luciferase reporter assay and some rescue experiments were performed to evaluate the underlying mechanism of MTFR1 in LUAD.Results: MTFR1 was upregulated in LUAD cells and tissues and correlated with dismal clinicopathologic features and a worse prognosis of patients with LUAD. Functionally, MTFR1 overexpression stimulated the proliferation, invasion, migration and glycolytic capacity and impeded the apoptosis of LUAD cells; however, opposite results were obtained when MTFR1 expression was knocked down. MTFR1, which was directly targeted by miR-29c-3p, may exert its biological functions through the AMPK/mTOR signalling pathway.Conclusion: MTFR1 promotes the progression of LUAD. Therefore, targeting MTFR1 can offer an effective therapeutic strategy for LUAD treatment.</p
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