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1.
Development ; 146(14)2019 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-31320327

RESUMO

Cucumber (Cucumis sativus L.) is an important vegetable crop that carries on vegetative growth and reproductive growth simultaneously. Indeterminate growth is favourable for fresh market under protected environments, whereas determinate growth is preferred for pickling cucumber in the once-over mechanical harvest system. The genetic basis of determinacy is largely unknown in cucumber. In this study, map-based cloning of the de locus showed that the determinate growth habit is caused by a non-synonymous SNP in CsTFL1 CsTFL1 is expressed in the subapical regions of the shoot apical meristem, lateral meristem and young stems. Ectopic expression of CsTFL1 rescued the terminal flower phenotype in the Arabidopsis tfl1-11 mutant and delayed flowering in wild-type Arabidopsis Knockdown of CsTFL1 resulted in determinate growth and formation of terminal flowers in cucumber. Biochemical analyses indicated that CsTFL1 interacts with a homolog of the miRNA biogenesis gene CsNOT2a; CsNOT2a interacts with FDP. Cucumber CsFT directly interacts with CsNOT2a and CsFD, and CsFD interacts with two 14-3-3 proteins. These data suggest that CsTFL1 competes with CsFT for interaction with CsNOT2a-CsFDP to inhibit determinate growth and terminal flower formation in cucumber.

2.
Proc Natl Acad Sci U S A ; 116(28): 14349-14357, 2019 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-31239335

RESUMO

Endocytosis is essential to all eukaryotes, but how cargoes are selected for internalization remains poorly characterized. Extracellular cargoes are thought to be selected by transmembrane receptors that bind intracellular adaptors proteins to initiate endocytosis. Here, we report a mechanism for clathrin-mediated endocytosis (CME) of extracellular lanthanum [La(III)] cargoes, which requires extracellular arabinogalactan proteins (AGPs) that are anchored on the outer face of the plasma membrane. AGPs were colocalized with La(III) on the cell surface and in La(III)-induced endocytic vesicles in Arabidopsis leaf cells. Superresolution imaging showed that La(III) triggered AGP movement across the plasma membrane. AGPs were then colocalized and physically associated with the µ subunit of the intracellular adaptor protein 2 (AP2) complexes. The AGP-AP2 interaction was independent of CME, whereas AGP's internalization required CME and AP2. Moreover, we show that AGP-dependent endocytosis in the presence of La(III) also occurred in human cells. These findings indicate that extracellular AGPs act as conserved CME cargo receptors, thus challenging the current paradigm about endocytosis of extracellular cargoes.

3.
New Phytol ; 223(4): 1918-1936, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31091337

RESUMO

CCCH-type zinc-finger proteins play essential roles in regulating plant development and stress responses. However, the molecular and functional properties of non-tandem CCCH-type zinc-finger (non-TZF) proteins have been rarely characterized in plants. Here, we report the biological and molecular characterization of a sweet potato non-TZF gene, IbC3H18. We show that IbC3H18 exhibits tissue- and abiotic stress-specific expression, and could be effectively induced by abiotic stresses, including NaCl, polyethylene glycol (PEG) 6000, H2 O2 and abscisic acid (ABA) in sweet potato. Accordingly, overexpression of IbC3H18 led to increased, whereas knock-down of IbC3H18 resulted in decreased tolerance of sweet potato to salt, drought and oxidation stresses. In addition, IbC3H18 functions as a nuclear transcriptional activator and regulates the expression of a range of abiotic stress-responsive genes involved in reactive oxygen species (ROS) scavenging, ABA signaling, photosynthesis and ion transport pathways. Moreover, our data demonstrate that IbC3H18 physically interacts with IbPR5, and that overexpression of IbPR5 enhances salt and drought tolerance in transgenic tobacco plants. Collectively, our data indicate that IbC3H18 functions in enhancing abiotic stress tolerance in sweet potato, which may serve as a candidate gene for use in improving abiotic stress resistance in crops.

4.
Plant Physiol ; 179(4): 1723-1738, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30718347

RESUMO

Plastid isoprenoids, a diverse group of compounds that includes carotenoids, chlorophylls, tocopherols, and multiple hormones, are essential for plant growth and development. Here, we identified and characterized SEED CAROTENOID DEFICIENT (SCD), which encodes an enzyme that functions in the biosynthesis of plastid isoprenoids in maize (Zea mays). SCD converts 2C-methyl-d-erytrithol 2,4-cyclodiphosphate to 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate in the penultimate step of the methylerythritol phosphate (MEP) pathway. In scd mutants, plant growth and development are impaired and the levels of MEP-derived isoprenoids, such as carotenoids, chlorophylls, and tocopherols, as well as abscisic and gibberellic acids, are reduced in leaves and seeds. This scd metabolic alteration varies among plant tissues and under different light conditions. RNA-sequencing of the scd mutant and wild type identified a limited number of differentially expressed genes in the MEP pathway, although isoprenoid levels were significantly reduced in scd seeds and dark-grown leaves. Furthermore, SCD-overexpressing transgenic lines showed little or no differences in isoprenoid levels, indicating that SCD may be subject to posttranslational regulation or not represent a rate-limiting step in the MEP pathway. These results enhance our understanding of the transcriptomic and metabolic regulatory roles of enzymes in the MEP pathway and of their effects on downstream isoprenoid pathways in various plant tissues and under different light conditions.


Assuntos
Proteínas de Plantas/fisiologia , Zea mays/metabolismo , Carotenoides/metabolismo , Cloroplastos/genética , Cloroplastos/fisiologia , Mapeamento Cromossômico , Clonagem Molecular , Eritritol/análogos & derivados , Eritritol/metabolismo , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plastídeos/metabolismo , Fosfatos Açúcares/genética , Fosfatos Açúcares/metabolismo , Terpenos/metabolismo , Zea mays/genética , Zea mays/crescimento & desenvolvimento
5.
Plant Cell ; 31(3): 699-714, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30760559

RESUMO

Potassium and nitrogen are essential nutrients for plant growth and development. Plants can sense potassium nitrate (K+/NO3 -) levels in soils, and accordingly they adjust root-to-shoot K+/NO3 - transport to balance the distribution of these ions between roots and shoots. In this study, we show that the transcription factorMYB59 maintains this balance by regulating the transcription of the Arabidopsis (Arabidopsis thaliana) Nitrate Transporter1.5 (NRT1.5)/ Nitrate Transporter/Peptide Transporter Family7.3 (NPF7.3) in response to low K+ (LK) stress. The myb59 mutant showed a yellow-shoot sensitive phenotype when grown on LK medium. Both the transcript and protein levels of NPF7.3 were remarkably reduced in the myb59 mutant. LK stress repressed transcript levels of both MYB59 and NPF7.3 The npf7.3 and myb59 mutants, as well as the npf7.3 myb59 double mutant, showed similar LK-sensitive phenotypes. Ion content analyses indicated that root-to-shoot K+/NO3 - transport was significantly reduced in these mutants under LK conditions. Moreover, chromatin immunoprecipitation and electrophoresis mobility shift assay assays confirmed that MYB59 bound directly to the NPF7.3 promoter. Expression of NPF7.3 in root vasculature driven by the PHOSPHATE 1 promoter rescued the sensitive phenotype of both npf7.3 and myb59 mutants. Together, these data demonstrate that MYB59 responds to LK stress and directs root-to-shoot K+/NO3 - transport by regulating the expression of NPF7.3 in Arabidopsis roots.

6.
New Phytol ; 221(1): 341-355, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30019753

RESUMO

Group A protein phosphatase 2Cs (PP2Cs) are abscisic acid (ABA) co-receptors that negatively regulate the ABA signaling pathway by inhibiting the downstream SnRK2 protein kinases. It has long been observed that exogenous ABA treatments dramatically induce the expression of group A PP2C genes, but the underlying molecular mechanisms and the biological significance remain largely unknown. Here, by using GUS reporter transgenic lines in which various lengths of ABI1 and ABI2 promoters were used to drive GUS gene expression, we defined the promoter fragments that confer ABA inducibility to ABI1 and ABI2. We further showed that ABRE-binding factors (ABFs), the bZIP family transcription factors, directly bind to the promoters of group A PP2C genes, and mediate rapid induction of their expression on exogenous ABA treatments. Moreover, our data indicated that ABA dramatically induces the expression of ABF genes and the accumulation of endogenous ABF proteins, and that ABFs themselves are involved in this induction, thus providing another layer of ABA regulation towards ABF proteins in addition to the well-characterized ABA-induced phosphorylation by SnRK2 protein kinases. Together, our data demonstrate that ABFs mediate rapid ABA induction of group A PP2C genes, thus playing a role in the negative feedback regulation of ABA signaling.

7.
Artigo em Inglês | MEDLINE | ID: mdl-30478060

RESUMO

Phytochrome A (phyA) is the only plant photoreceptor that perceives far-red light and then mediates various responses to this signal. Phosphorylation and dephosphorylation of oat phyA have been extensively studied, and it was shown that phosphorylation of a serine residue in the hinge region of oat phyA could regulate the interaction of phyA with its signal transducers. However, little is known about the role of the hinge region of Arabidopsis phyA. Here, we report that three sites in the hinge region of Arabidopsis phyA (i.e., S590, T593, and S602) are essential in regulating phyA function. Mutating all three of these sites to either alanines or aspartic acids impaired phyA function, changed the interactions of mutant phyA with FHY1 and FHL, and delayed the degradation of mutant phyA upon light exposure. Moreover, the in vivo formation of a phosphorylated phyA form was greatly affected by these mutations, while our data indicated that the abundance of this phosphorylated phyA form correlated well with the extent of phyA function, thus suggesting a pivotal role of the phosphorylated phyA in inducing the far-red light response. Taking these data together, our study reveals the important role of the hinge region of Arabidopsis phyA in regulating phyA phosphorylation and function, thus linking specific residues in the hinge region to the regulatory mechanisms of phyA phosphorylation.

8.
Life Sci ; 2018 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-30496728

RESUMO

AIMS: The tumor necrosis factor (TNF)-alpha-induced protein 8-like 2 (TIPE2) participates in multiple inflammatory diseases. However, its underlying mechanism in osteoporosis has not been elucidated. The aim of current study is to preliminarily clarify the function of TIPE2 in the pathogenesis of osteoporosis. MAIN METHODS: TIPE2 expression in patients with osteoporosis was measured by Western blot and qRT-PCR methods. Proinflammatory cytokines including TNF-α, IL-1 and IL-6 were assessed via enzyme-linked immunosorbent assay. Serum fasting PINP and ß-CTX were measured by the chemiluminescence method. Simple logistic regression analysis was performed for the odds ratio (OR) for TIPE2. KEY FINDINGS: TIPE2 expression in patients with osteoporosis was dramatically decreased and negatively correlated with proinflammatory cytokines. Furthermore, TIPE2 level was negatively correlated with fasting ß-CTX, but not PINP, indicating that TIPE2 participates in the pathogenesis of osteoporosis dominantly by supression of bone resorption. Interestingly, TIPE2 expression level was positively correlated with bone mineral density (BMD), and its expression level can predict the risk of bone fracture using the simple logistic regression assay. SIGNIFICANCE: Our findings clarify that TIPE2 alleviates the pathogenesis of osteoporosis by suppressing the inflammatory status and the ability of TIPE2 for predicts bone fracture further demonstrated that TIPE2 might serve as a novel diagnostic marker and a therapeutic target for osteoporosis.

9.
Mol Immunol ; 101: 471-478, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30114619

RESUMO

TIPE2 participates in multiple types of cancer development. However, its mechanism underlying chemoresistance in osteosarcoma has not been elucidated. Herein, we observed the expression of TIPE2 and MDR1 in cis-platin-resistant osteosarcoma tissues and cell lines. Compared to their matched sensitive cell lines and tissues, TIPE2 was downregulated while MDR1 expression was increased. Further investigation showed that overexpression of TIPE2 effectively inhibited MDR1 expression and greatly sensitized osteosarcoma cells to cis-platin, both in vivo and in vitro. Mechanistically, TIPE2 inhibited the transcription of the MDR1 promoter by interfering with the TAK1-NF-κB and -AP-1 pathways. Overall, our results elucidated for the first time that TIPE2 sensitizes osteosarcoma cells to cis-platin through downregulation of MDR1 and may be a novel target in osteosarcoma therapy.

10.
J Integr Plant Biol ; 60(10): 924-937, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29786952

RESUMO

The small ubiquitin-related modifier (SUMO) modification plays an important role in the regulation of abscisic acid (ABA) signaling, but the function of the SUMO protease, in ABA signaling, remains largely unknown. Here, we show that the SUMO protease, ASP1 positively regulates ABA signaling. Mutations in ASP1 resulted in an ABA-insensitive phenotype, during early seedling development. Wild-type ASP1 successfully rescued, whereas an ASP1 mutant (C577S), defective in SUMO protease activity, failed to rescue, the ABA-insensitive phenotype of asp1-1. Expression of ABI5 and MYB30 target genes was attenuated in asp1-1 and our genetic analyses revealed that ASP1 may function upstream of ABI5 and MYB30. Interestingly, ASP1 accumulated upon ABA treatment, and ABA-induced accumulation of ABI5 (a positive regulator of ABA signaling) was abolished, whereas ABA-induced accumulation of MYB30 (a negative regulator of ABA signaling) was increased in asp1-1. These findings support the hypothesis that increased levels of ASP1, upon ABA treatment, tilt the balance between ABI5 and MYB30 towards ABI5-mediated ABA signaling.


Assuntos
Ácido Abscísico/farmacologia , Proteínas de Arabidopsis/metabolismo , Arabidopsis/efeitos dos fármacos , Arabidopsis/metabolismo , Cisteína Endopeptidases/metabolismo , Plântula/efeitos dos fármacos , Plântula/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Cisteína Endopeptidases/genética , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Plântula/genética , Transdução de Sinais/efeitos dos fármacos
11.
Plant Cell ; 30(4): 815-834, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29618630

RESUMO

The reversible phosphorylation of proteins by kinases and phosphatases is an antagonistic process that modulates many cellular functions. Protein phosphatases are usually negatively regulated by inhibitor proteins. During abscisic acid (ABA) signaling, these inhibitor proteins comprise PYR1/PYL/RCAR ABA receptors, which inhibit the core negative regulators, the clade A type 2C protein phosphatases (PP2Cs). However, it is not known whether these PP2Cs are positively regulated by other proteins. Here, we identified an Arabidopsis thaliana ear1 (enhancer of aba co-receptor1) mutant that exhibits pleiotropic ABA-hypersensitive phenotypes. EAR1 encodes an uncharacterized protein that is conserved in both monocots and dicots. EAR1 interacts with the N-terminal inhibition domains of all six PP2Cs, ABA INSENSITIVE1 (ABI1), ABI2, HYPERSENSITIVE TO ABA1 (HAB1), HAB2, ABA-HYPERSENSITIVE GERMINATION1 (AHG1), and AHG3, during ABA signaling and enhances the activity of PP2Cs both in vitro and in vivo. ABA treatment caused EAR1 to accumulate in the nucleus. These results indicate that EAR1 is a negative regulator of ABA signaling that enhances the activity of PP2Cs by interacting with and releasing the N-terminal autoinhibition of these proteins.

12.
Plant Cell ; 30(4): 835-852, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29588390

RESUMO

Phytochrome A (phyA) is the primary plant photoreceptor responsible for perceiving and mediating various responses to far-red (FR) light and is essential for survival in canopy shade. In this study, we identified two Arabidopsis thaliana mutants that grew longer hypocotyls in FR light. Genetic analyses showed that they were allelic and their FR phenotypes were caused by mutations in the gene named TANDEM ZINC-FINGER/PLUS3 (TZP), previously shown to encode a nuclear protein involved in blue light signaling and phyB-dependent regulation of photoperiodic flowering. We show that the expression of TZP is dramatically induced by light and that TZP proteins are differentially modified in different light conditions. Furthermore, we show that TZP interacts with both phyA and FAR-RED ELONGATED HYPOCOTYL1 (FHY1) and regulates the abundance of phyA, FHY1, and ELONGATED HYPOCOTYL5 proteins in FR light. Moreover, our data indicate that TZP is required for the formation of a phosphorylated form of phyA in the nucleus in FR light. Together, our results identify TZP as a positive regulator of phyA signaling required for phosphorylation of the phyA photoreceptor, thus suggesting an important role of phosphorylated phyA in inducing the FR light response.

13.
PLoS Genet ; 14(2): e1007237, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29462139

RESUMO

The control of seed germination and seed dormancy are critical for the successful propagation of plant species, and are important agricultural traits. Seed germination is tightly controlled by the balance of gibberellin (GA) and abscisic acid (ABA), and is influenced by environmental factors. The COP9 Signalosome (CSN) is a conserved multi-subunit protein complex that is best known as a regulator of the Cullin-RING family of ubiquitin E3 ligases (CRLs). Multiple viable mutants of the CSN showed poor germination, except for csn5b-1. Detailed analyses showed that csn1-10 has a stronger seed dormancy, while csn5a-1 mutants exhibit retarded seed germination in addition to hyperdormancy. Both csn5a-1 and csn1-10 plants show defects in the timely removal of the germination inhibitors: RGL2, a repressor of GA signaling, and ABI5, an effector of ABA responses. We provide genetic evidence to demonstrate that the germination phenotype of csn1-10 is caused by over-accumulation of RGL2, a substrate of the SCF (CRL1) ubiquitin E3 ligase, while the csn5a-1 phenotype is caused by over-accumulation of RGL2 as well as ABI5. The genetic data are consistent with the hypothesis that CSN5A regulates ABI5 by a mechanism that may not involve CSN1. Transcriptome analyses suggest that CSN1 has a more prominent role than CSN5A during seed maturation, but CSN5A plays a more important role than CSN1 during seed germination, further supporting the functional distinction of these two CSN genes. Our study delineates the molecular targets of the CSN complex in seed germination, and reveals that CSN5 has additional functions in regulating ABI5, thus the ABA signaling pathway.


Assuntos
Proteínas de Arabidopsis/metabolismo , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Complexo do Signalossomo COP9/fisiologia , Germinação , Proteólise , Fatores de Transcrição/metabolismo , Complexo do Signalossomo COP9/genética , Germinação/genética , Fenótipo , Plantas Geneticamente Modificadas , Sementes/genética , Sementes/crescimento & desenvolvimento , Transdução de Sinais/genética
14.
Molecules ; 23(2)2018 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-29414844

RESUMO

To develop natural product resources to control cigarette beetles (Lasioderma serricorne), the essential oil from Artemisia lavandulaefolia (Compositae) was investigated. Oil was extracted by hydrodistillation of the above-ground portion of A. lavandulaefolia and analyzed using gas chromatography-mass spectrometer (GC-MS). Extracted essential oil and three compounds isolated from the oil were then evaluated in laboratory assays to determine the fumigant, contact, and repellent efficacy against the stored-products' pest, L. serricorne. The bioactive constituents from the oil extracts were identified as chamazulene (40.4%), 1,8-cineole (16.0%), and ß-caryophyllene (11.5%). In the insecticidal activity assay, the adults of L. serricorne were susceptible to fumigant action of the essential oil and 1,8-cineole, with LC50 values of 31.81 and 5.18 mg/L air. The essential oil, 1,8-cineole, chamazulene, and ß-caryophyllene exhibited contact toxicity with LD50 values of 13.51, 15.58, 15.18 and 35.52 µg/adult, respectively. During the repellency test, the essential oil and chamazulene had repellency approximating the positive control. The results indicated that chamazulene was abundant in A. lavandulaefolia essential oil and was toxic to cigarette beetles.


Assuntos
Artemisia/química , Besouros/efeitos dos fármacos , Óleos Voláteis/química , Óleos Voláteis/farmacologia , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Animais , Repelentes de Insetos/química , Repelentes de Insetos/farmacologia , Inseticidas/química , Inseticidas/farmacologia , Estrutura Molecular , Óleos Voláteis/isolamento & purificação , Compostos Fitoquímicos/química , Extratos Vegetais/isolamento & purificação
15.
Mol Ecol ; 27(1): 216-232, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29134709

RESUMO

Understanding the genetic basis of the switch from asexual to sexual lifestyles in response to sometimes rapid environmental changes is one of the major challenges in fungal ecology. Light appears to play a critical role in the asexual-sexual switch-but fungal genomes harbour diverse light sensors. Fungal opsins are homologous to bacterial green-light-sensory rhodopsins, and their organismal functions in fungi have not been well understood. Three of these opsin-like proteins were widely distributed across fungal genomes, but homologs of the Fusarium opsin-like protein CarO were present only in plant-associated fungi. Key amino acids, including potential retinal binding sites, functionally diverged on the phylogeny of opsins. This diversification of opsin-like proteins could be correlated with life history-associated differences among fungi in their expression and function during morphological development. In Neurospora crassa and related species, knockout of the opsin NOP-1 led to a phenotype in the regulation of the asexual-sexual switch, modulating response to both light and oxygen conditions. Sexual development commenced early in ∆nop-1 strains cultured in unsealed plates under constant blue and white light. Furthermore, comparative transcriptomics showed that the expression of nop-1 is light-dependent and that the ∆nop-1 strain abundantly expresses genes involved in oxidative stress response, genes enriched in NAD/NADP binding sites, genes with functions in proton transmembrane movement and catalase activity, and genes involved in the homeostasis of protons. Based on these observations, we contend that light and oxidative stress regulate the switch via light-responsive and ROS pathways in model fungus N. crassa and other fungi.

16.
J Exp Bot ; 69(3): 423-439, 2018 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-29244171

RESUMO

Photomorphogenesis is an important process in which seedlings emerge from soil and begin autotrophic growth. Mechanisms of photomorphogenesis include light signal perception, signal transduction, and the modulation of expression of light-responsive genes, ultimately leading to cellular and developmental changes. Phytochrome-interacting factors (PIFs) play negative regulatory roles in photomorphogenesis. Light-induced activation of phytochromes triggers rapid phosphorylation and degradation of PIFs, but the kinases responsible for the phosphorylation of PIFs are largely unknown. Here, we show that Arabidopsis MPK6 is a kinase involved in phosphorylating PIF3 and regulating red light-induced cotyledon opening, a crucial process during seedling photomorphogenesis. MPK6 was activated by red light, and the cotyledon opening angle in red light was reduced in mpk6 seedlings. MKK10, a MAPKK whose function is currently unclear, appears to act as a kinase upstream of MPK6 in regulating cotyledon opening. Activation of MPK6 by MKK10 led to the phosphorylation of PIF3 and accelerated its turnover in transgenic seedlings. Accordingly, the overexpression of PIF3 suppressed MKK10-induced cotyledon opening. MKK10 and MPK6 function downstream of phyB in regulating seedling cotyledon opening in red light. Therefore, the MKK10-MPK6 cascade appears to mediate the regulation of red-light-controlled seedling photomorphogenesis via a mechanism that might involve the phosphorylation of PIF3.

17.
Clin Cancer Res ; 24(2): 445-459, 2018 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-29084921

RESUMO

Purpose: Regulated in development and DNA damage response-1 (REDD1) is a stress-related protein and is involved in the progression of cancer. The role and regulatory mechanism of REDD1 in bladder urothelial carcinoma (BUC), however, is yet unidentified.Experimental Design: The expression of REDD1 in BUC was detected by Western blot analysis and immunohistochemistry (IHC). The correlation between REDD1 expression and clinical features in patients with BUC were assessed. The effects of REDD1 on cellular proliferation, apoptosis, autophagy, and paclitaxel sensitivity were determined both in vitro and in vivo Then the targeted-regulating mechanism of REDD1 by miRNAs was explored.Results: Here the significant increase of REDD1 expression is detected in BUC tissue, and REDD1 is first reported as an independent prognostic factor in patients with BUC. Silencing REDD1 expression in T24 and EJ cells decreased cell proliferation, increased apoptosis, and decreased autophagy, whereas the ectopic expression of REDD1 in RT4 and BIU87 cells had the opposite effect. In addition, the REDD1-mediated proliferation, apoptosis, and autophagy are found to be negatively regulated by miR-22 in vitro, which intensify the paclitaxel sensitivity via inhibition of the well-acknowledged REDD1-EEF2K-autophagy axis. AKT/mTOR signaling initially activated or inhibited in response to silencing or enhancing REDD1 expression and then recovered rapidly. Finally, the inhibited REDD1 expression by either RNAi or miR-22 sensitizes BUC tumor cells to paclitaxel in a subcutaneous transplant carcinoma model in vivoConclusions: REDD1 is confirmed as an oncogene in BUC, and antagonizing REDD1 could be a potential therapeutic strategy to sensitize BUC cells to paclitaxel. Clin Cancer Res; 24(2); 445-59. ©2017 AACR.

18.
Proc Natl Acad Sci U S A ; 114(46): 12321-12326, 2017 11 14.
Artigo em Inglês | MEDLINE | ID: mdl-29087315

RESUMO

CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1), a well-known E3 ubiquitin ligase, functions as a central regulator of plant growth and photomorphogenic development in plants, including hypocotyl elongation. It has been well-established that, in darkness, COP1 targets many photomorphogenesis-promoting factors for ubiquitination and degradation in the nucleus. However, increasing evidence has shown that a proportion of COP1 is also localized outside the nucleus in dark-grown seedlings, but the physiological function of this localization remains largely unclear. In this study, we demonstrate that COP1 directly targets and mediates the degradation of WAVE-DAMPENED 2-LIKE 3 (WDL3) protein, a member of the microtubule-associated protein (MAP) WVD2/WDL family involved in regulating hypocotyl cell elongation of Arabidopsis seedlings. We show that COP1 interacts with WDL3 in vivo in a dark-dependent manner at cortical microtubules. Moreover, our data indicate that COP1 directly ubiquitinates WDL3 in vitro and that WDL3 protein is degraded in WT seedlings but is abundant in the cop1 mutant in the dark. Consistently, introduction of the wdl3 mutation weakened, whereas overexpression of WDL3 enhanced, the short-hypocotyl phenotype of cop1 mutant in darkness. Together, this study reveals a function of COP1 in regulating the protein turnover of a cytosol-localized MAP in etiolated hypocotyls, thus providing insights into COP1-mediated degradation of downstream factors to control seedling photomorphogenesis.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Hipocótilo/genética , Proteínas Associadas aos Microtúbulos/genética , Microtúbulos/metabolismo , Ubiquitina-Proteína Ligases/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Arabidopsis/efeitos da radiação , Proteínas de Arabidopsis/metabolismo , Núcleo Celular/metabolismo , Núcleo Celular/ultraestrutura , Escuridão , Regulação da Expressão Gênica no Desenvolvimento , Hipocótilo/crescimento & desenvolvimento , Hipocótilo/metabolismo , Hipocótilo/efeitos da radiação , Luz , Transdução de Sinal Luminoso , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/ultraestrutura , Desenvolvimento Vegetal/genética , Proteólise , Plântula/genética , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Plântula/efeitos da radiação , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação
19.
Mol Med Rep ; 16(3): 3331-3338, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28765962

RESUMO

Sustained hyperglycemic stimulation of vascular cells is involved in the pathogenesis of diabetes mellitus­induced cardiovascular complications. Silent information regulator T1 (SIRT1), a mammalian sirtuin, has been previously recognized to protect endothelial cells against hyperglycemia­induced oxidative stress. In the present study, human umbilical vein endothelial cells (HUV­EC­C) were treated with D­glucose, and the levels of oxidative stress, mitochondrial dysfunction, the rate of apoptosis and SIRT1 activity were measured. The effect of manipulated SIRT1 activity on hyperglycemia­induced oxidative stress, mitochondrial dysfunction and apoptosis was then assessed using the SIRT1 activator, resveratrol (RSV), and the SIRT1 inhibitor, sirtinol. The present study confirmed that hyperglycemia promotes oxidative stress and mitochondrial dysfunction in HUV­EC­C cells. The accumulation of reactive oxygen species, the swelling of mitochondria, the ratio of adenosine 5'­diphosphate to adenosine 5'­triphosphate and localized mitochondrial superoxide levels were all increased following D­glucose treatment, whereas the mitochondrial membrane potential was significantly reduced by >50 mg/ml D­glucose treatment. In addition, hyperglycemia was confirmed to induce apoptosis in HUV­EC­C cells. Furthermore, the results confirmed the prevention and aggravation of hyperglycemia­induced apoptosis by RSV treatment and sirtinol treatment, via the amelioration and enhancement of oxidative stress and mitochondrial dysfunction in HUV­EC­C cells, respectively. In conclusion, the present study revealed that hyperglycemia promotes oxidative stress, mitochondrial dysfunction and apoptosis in HUV­EC­C cells, and manipulation of SIRT1 activity regulated hyperglycemia­induced mitochondrial dysfunction and apoptosis in HUV­EC­C cells. The data revealed the protective effect of SIRT1 against hyperglycemia­induced apoptosis via the alleviation of mitochondrial dysfunction and oxidative stress.


Assuntos
Hiperglicemia/metabolismo , Mitocôndrias/metabolismo , Estresse Oxidativo , Sirtuína 1/metabolismo , Apoptose , Células Endoteliais da Veia Umbilical Humana , Humanos , Hiperglicemia/patologia , Mitocôndrias/patologia
20.
Proc Natl Acad Sci U S A ; 114(32): E6695-E6702, 2017 08 08.
Artigo em Inglês | MEDLINE | ID: mdl-28739888

RESUMO

Light and temperature are major environmental factors that coordinately control plant growth and survival. However, how plants integrate light and temperature signals to better adapt to environmental stresses is poorly understood. PHYTOCHROME-INTERACTING FACTOR 3 (PIF3), a key transcription factor repressing photomorphogenesis, has been shown to play a pivotal role in mediating plants' responses to various environmental signals. In this study, we found that PIF3 functions as a negative regulator of Arabidopsis freezing tolerance by directly binding to the promoters of C-REPEAT BINDING FACTOR (CBF) genes to down-regulate their expression. In addition, two F-box proteins, EIN3-BINDING F-BOX 1 (EBF1) and EBF2, directly target PIF3 for 26S proteasome-mediated degradation. Consistently, ebf1 and ebf2 mutants were more sensitive to freezing than were the wild type, and the pif3 mutation suppressed the freezing-sensitive phenotype of ebf1 Furthermore, cold treatment promoted the degradation of EBF1 and EBF2, leading to increased stability of the PIF3 protein and reduced expression of the CBF genes. Together, our study uncovers an important role of PIF3 in Arabidopsis freezing tolerance by negatively regulating the expression of genes in the CBF pathway.


Assuntos
Aclimatação/fisiologia , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Congelamento , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas F-Box/genética , Proteínas F-Box/metabolismo , Mutação
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