17 results on '"Xiaoming Zhou"'
Search Results
2. Regulation of Hippo-YAP signaling by insulin-like growth factor-1 receptor in the tumorigenesis of diffuse large B-cell lymphoma
- Author
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Hongzhi Xu, Xiaoming Zhou, Ya Zhang, Xin Wang, Jianhong Wang, Xiangxiang Zhou, Juan Yang, Xiaosheng Fang, Na Chen, and Ying Li
- Subjects
0301 basic medicine ,Male ,Cancer Research ,Cell cycle checkpoint ,Apoptosis ,Mice, SCID ,medicine.disease_cause ,Receptor, IGF Type 1 ,Mice ,Random Allocation ,0302 clinical medicine ,Pseudolymphoma ,Hippo-YAP ,Insulin-Like Growth Factor I ,RNA, Small Interfering ,Podophyllotoxin ,Gene knockdown ,B-Lymphocytes ,Hematology ,lcsh:Diseases of the blood and blood-forming organs ,Cell cycle ,Middle Aged ,Tyrphostins ,lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens ,Neoplasm Proteins ,Gene Expression Regulation, Neoplastic ,Cell Transformation, Neoplastic ,Oncology ,030220 oncology & carcinogenesis ,CYR61 ,Female ,RNA Interference ,Lymphoma, Large B-Cell, Diffuse ,Signal Transduction ,Biology ,Protein Serine-Threonine Kinases ,lcsh:RC254-282 ,03 medical and health sciences ,Downregulation and upregulation ,Cell Line, Tumor ,medicine ,Animals ,Humans ,Hippo Signaling Pathway ,Molecular Biology ,CRISPR/Cas9 ,Adaptor Proteins, Signal Transducing ,Aged ,Cell growth ,lcsh:RC633-647.5 ,Research ,Verteporfin ,YAP-Signaling Proteins ,Cell Cycle Checkpoints ,Xenograft Model Antitumor Assays ,CTGF ,030104 developmental biology ,DLBCL ,Cancer research ,CRISPR-Cas Systems ,Carcinogenesis ,IGF-1R ,Transcription Factors - Abstract
Background Hippo-Yes-associated protein (YAP) signaling is a key regulator of organ size and tumorigenesis, yet the underlying molecular mechanism is still poorly understood. At present, the significance of the Hippo-YAP pathway in diffuse large B-cell lymphoma (DLBCL) is ill-defined. Methods The expression of YAP in DLBCL was determined in public database and clinical specimens. The effects of YAP knockdown, CRISPR/Cas9-mediated YAP deletion, and YAP inhibitor treatment on cell proliferation and the cell cycle were evaluated both in vitro and in vivo. RNA sequencing was conducted to detect dysregulated RNAs in YAP-knockout DLBCL cells. The regulatory effects of insulin-like growth factor-1 receptor (IGF-1R) on Hippo-YAP signaling were explored by targeted inhibition and rescue experiments. Results High expression of YAP was significantly correlated with disease progression and poor prognosis. Knockdown of YAP expression suppressed cell proliferation and induced cell cycle arrest in DLBCL cells. Verteporfin (VP), a benzoporphyrin derivative, exerted an anti-tumor effect by regulating the expression of YAP and the downstream target genes, CTGF and CYR61. In vitro and in vivo studies revealed that deletion of YAP expression with a CRISPR/Cas9 genome editing system significantly restrained tumor growth. Moreover, downregulation of IGF-1R expression led to a remarkable decrease in YAP expression. In contrast, exposure to IGF-1 promoted YAP expression and reversed the inhibition of YAP expression induced by IGF-1R inhibitors. Conclusions Our study highlights the critical role of YAP in the pathogenesis of DLBCL and uncovers the regulatory effect of IGF-1R on Hippo-YAP signaling, suggesting a novel therapeutic strategy for DLBCL.
- Published
- 2020
3. Leishmania infantum-chagasi activates SHP-1 and reduces NFAT5/TonEBP activity in the mouse kidney inner medulla
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Aihong Zhang, Nancy L. Koles, Naomi E. Aronson, Xiaoming Zhou, and Hong Wang
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STAT3 Transcription Factor ,GABA Plasma Membrane Transport Proteins ,Kidney Cortex ,Physiology ,Urine ,Biology ,Cell Line ,Kidney Concentrating Ability ,Mice ,NFAT5 ,medicine ,Animals ,Leishmania infantum ,Inner medulla ,Transcription factor ,Kidney Medulla ,Mice, Inbred BALB C ,Aquaporin 2 ,Symporters ,Protein Tyrosine Phosphatase, Non-Receptor Type 6 ,Leishmaniasis ,medicine.disease ,Molecular biology ,Disease Models, Animal ,Visceral leishmaniasis ,Gene Expression Regulation ,Immunology ,Mouse Kidney ,Leishmaniasis, Visceral ,Female ,Transcription Factors - Abstract
Visceral leishmaniasis patients have been reported to have a urine concentration defect. Concentration of urine by the renal inner medulla is essentially dependent on a transcription factor, NFAT5/TonEBP, because it activates expression of osmoprotective genes betaine/glycine transporter 1 (BGT1) and sodium/myo-inositol transporter (SMIT), and water channel aquaporin-2, all of which are imperative for concentrating urine. Leishmania parasites evade macrophage immune defenses by activating protein tyrosine phosphatases, among which SHP-1 is critical. We previously demonstrated that SHP-1 inhibits tonicity-dependent activation of NFAT5/TonEBP in HEK293 cells through screening a genome-wide small interfering (si) RNA library against phosphatases (Zhou X, Gallazzini M, Burg MB, Ferraris JD. Proc Natl Acad Sci USA 107: 7072–7077, 2010). We sought to examine whether Leishmania can activate SHP-1 and inhibit NFAT5/TonEBP activity in the renal inner medulla in a murine model of visceral leishmaniasis by injection of female BALB/c mice with a single intravenous dose of 5 × 105 L. chagasi metacyclic promastigotes. We found that SHP-1 is expressed in the kidney inner medulla. L. chagasi activates SHP-1 with an increase in stimulatory phosphorylation of SHP-1-Y536 in the region. L. chagasi reduces expression of NFAT5/TonEBP mRNA and protein as well as expression of its targeted genes: BGT1, SMIT, and aquaporin-2. The culture supernatant from L. chagasi metacyclic promastigotes increases SHP-1 protein abundance and potently inhibits NFAT5 transcriptional activity in mIMCD3 cells. However, L. chagasi in our animal model has no significant effect on urinary concentration. We conclude that L. chagasi, most likely through its secreted virulence factors, activates SHP-1 and reduces NFAT5/TonEBP gene expression, which leads to reduced NFAT5/TonEBP transcriptional activity in the kidney inner medulla.
- Published
- 2014
4. Sickle cell disease up‐regulates vasopressin, aquaporin 2, urea transporter A1, Na‐K‐Cl cotransporter 2, and epithelial Na channels in the mouse kidney medulla despite compromising urinary concentration ability
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Xiaoming Zhou, Ryan G. Morris, Mark A. Knepper, and Hong Wang
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Male ,Epithelial sodium channel ,Vasopressin ,Physiology ,Urea transporter ,Countercurrent multiplication ,030204 cardiovascular system & hematology ,Kidney ,lcsh:Physiology ,Mice ,0302 clinical medicine ,hemic and lymphatic diseases ,Solute Carrier Family 12, Member 1 ,Original Research ,lcsh:QP1-981 ,biology ,Chemistry ,Protein Tyrosine Phosphatase, Non-Receptor Type 6 ,Water-Electrolyte Balance ,Up-Regulation ,medicine.anatomical_structure ,Aquaporin 2 ,Regulatory Pathways ,Vasa recta ,SHP‐1 ,Female ,Cardiovascular Conditions, Disorders and Treatments ,medicine.medical_specialty ,Vasopressins ,Genetic Conditions Disorders and Treatments ,Urinary system ,Anemia, Sickle Cell ,03 medical and health sciences ,Physiology (medical) ,Internal medicine ,medicine ,Na-K-Cl cotransporter ,Animals ,NFAT5 ,Epithelial Sodium Channels ,Renal Physiology ,Aquaporin 1 ,urogenital system ,Membrane Transport Proteins ,Endocrinology ,TonEBP ,biology.protein ,030217 neurology & neurosurgery ,Transcription Factors - Abstract
Sickle cell disease (SCD)‐induced urinary concentration defect has been proposed as caused by impaired ability of the occluded vasa recta due to red blood cell sickling to serve as countercurrent exchangers and renal tubules to absorb water and solutes. However, the exact molecular mechanisms remain largely unknown. The present studies were undertaken to determine the effects of SCD on vasopressin, aquaporin2 (AQP2), urea transporter A1 (UTA1), Na‐K‐Cl co‐transporter 2 (NKCC2), epithelial Na channels (ENaC), aquaporin1 (AQP1), nuclear factor of activated T cells 5 (NFAT5) and Src homology region‐2 domain‐containing phosphatase‐1 (SHP‐1), an important regulator of NFAT5, in the Berkeley SCD mouse kidney medulla. Under water repletion, SCD only induced a minor urinary concentration defect associated with increased urinary vasopressin level alone with the well‐known effects of vasopressin: protein abundance of AQP2, UTA1 and ENaC‐β and apical targeting of AQP2 as compared with non‐SCD. SCD did not significantly affect AQP1 protein level. Water restriction had no further significant effect on SCD urinary vasopressin. NFAT5 is also critical to urinary concentration. Instead, water restriction‐activated NFAT5 associated with inhibition of SHP‐1 in the SCD mice. Yet, water restriction only elevated urinary osmolality by 28% in these mice as opposed to 104% in non‐SCD mice despite similar degree increases of protein abundance of AQP2, NKCC2 and AQP2‐S256‐P. Water‐restriction had no significant effect on protein abundance of ENaC or AQP1 in either strain. In conclusion, under water repletion SCD, only induces a minor defect in urinary concentration because of compensation from the up‐regulated vasopressin system. However, under water restriction, SCD mice struggle to concentrate urine despite activating NFAT5. SCD‐induced urinary concentration defect appears to be resulted from the poor blood flow in vasa recta rather than the renal tubules’ ability to absorb water and solutes.
- Published
- 2019
5. High NaCl-induced inhibition of PTG contributes to activation of NFAT5 through attenuation of the negative effect of SHP-1
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Xiaoming Zhou, Joan D. Ferraris, Hong Wang, and Maurice B. Burg
- Subjects
MAPK/ERK pathway ,animal structures ,MAP Kinase Signaling System ,Physiology ,p38 mitogen-activated protein kinases ,Protein subunit ,Blotting, Western ,Nuclear Localization Signals ,Sodium Chloride ,Biology ,Transfection ,Polymerase Chain Reaction ,p38 Mitogen-Activated Protein Kinases ,NFAT5 ,Protein Phosphatase 1 ,Humans ,Immunoprecipitation ,RNA, Messenger ,RNA, Small Interfering ,Transcription factor ,Gene knockdown ,Protein Tyrosine Phosphatase, Non-Receptor Type 6 ,Protein phosphatase 1 ,Articles ,Molecular biology ,HEK293 Cells ,embryonic structures ,Phosphorylation ,HeLa Cells ,Plasmids ,Transcription Factors - Abstract
Activation of the transcription factor NFAT5 by high NaCl involves changes in phosphorylation. By siRNA screening, we previously found that protein targeting to glycogen (PTG), a regulatory subunit of protein phosphatase1 (PP1), contributes to regulation of high NaCl-induced NFAT5 transcriptional activity. The present study addresses the mechanism involved. We find that high NaCl-induced inhibition of PTG elevates NFAT5 activity by increasing NFAT5 transactivating activity, protein abundance, and nuclear localization. PTG acts via a catalytic subunit PP1γ. PTG associates physically with PP1γ, and NaCl reduces both this association and remaining PTG-associated PP1γ activity. High NaCl-induced phosphorylation of p38, ERK, and SHP-1 contributes to activation of NFAT5. Knockdown of PTG does not affect phosphorylation of p38 or ERK. However, PTG and PP1γ bind to SHP-1, and knockdown of either PTG or PP1γ increases high NaCl-induced phosphorylation of SHP-1-S591, which inhibits SHP-1. Mutation of SHP-1-S591 to alanine, which cannot be phosphorylated, increases inhibition of NFAT5 by SHP-1. Thus high NaCl reduces the stimulatory effect of PTG and PP1γ on SHP-1, which in turn reduces the inhibitory effect of SHP-1 on NFAT5. Our findings add to the known functions of PTG, which was previously recognized only for its glycogenic activity.
- Published
- 2013
6. Snail1 controls epithelial–mesenchymal lineage commitment in focal adhesion kinase–null embryonic cells
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Xiao Yan Li, Yuexian Hu, R. Grant Rowe, Jun-Lin Guan, Ann Park, Stephen J. Weiss, Xiaoming Zhou, David D. Schlaepfer, Dusko Ilic, and Gregory R. Dressler
- Subjects
Epithelial-Mesenchymal Transition ,Cellular differentiation ,Embryonic Development ,Biology ,Focal adhesion ,03 medical and health sciences ,Mice ,0302 clinical medicine ,Report ,Animals ,Epithelial–mesenchymal transition ,Mechanotransduction ,Research Articles ,030304 developmental biology ,0303 health sciences ,Cadherin ,digestive, oral, and skin physiology ,Cell Differentiation ,Cell Biology ,Embryo, Mammalian ,Embryonic stem cell ,Cell biology ,Mice, Inbred C57BL ,Phenotype ,030220 oncology & carcinogenesis ,Focal Adhesion Protein-Tyrosine Kinases ,Snail Family Transcription Factors ,Protein stabilization ,Signal transduction ,Signal Transduction ,Transcription Factors - Abstract
FAK promotes the epithelial–mesenchymal transition in mouse embryonic cells by regulating the transcription factor Snail1., Mouse embryonic cells isolated from focal adhesion kinase (FAK)–null animals at embryonic day 7.5 display multiple defects in focal adhesion remodeling, microtubule dynamics, mechanotransduction, proliferation, directional motility, and invasion. To date, the ability of FAK to modulate cell function has been ascribed largely to its control of posttranscriptional signaling cascades in this embryonic cell population. In this paper, we demonstrate that FAK unexpectedly exerts control over an epithelial–mesenchymal transition (EMT) program that commits embryonic FAK-null cells to an epithelial status highlighted by the expression of E-cadherin, desmoplakin, and cytokeratins. FAK rescue reestablished the mesenchymal characteristics of FAK-null embryonic cells to generate committed mouse embryonic fibroblasts via an extracellular signal–related kinase– and Akt-dependent signaling cascade that triggered Snail1 gene expression and Snail1 protein stabilization. These findings indentify FAK as a novel regulator of Snail1-dependent EMT in embryonic cells and suggest that multiple defects in FAK−/− cell behavior can be attributed to an inappropriate commitment of these cells to an epithelial, rather than fibroblastic, phenotype.
- Published
- 2011
7. Contribution of SHP-1 protein tyrosine phosphatase to osmotic regulation of the transcription factor TonEBP/OREBP
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Joan D. Ferraris, Morgan Gallazzini, Xiaoming Zhou, and Maurice B. Burg
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Cytoplasm ,Osmosis ,Small interfering RNA ,Phosphatase ,Protein tyrosine phosphatase ,Sodium Chloride ,Biology ,Biochemistry ,Models, Biological ,Cell Line ,Genetics ,Humans ,Phosphorylation ,RNA, Small Interfering ,Molecular Biology ,Transcription factor ,Heat-Shock Proteins ,Gene Library ,Cell Nucleus ,Gene knockdown ,Multidisciplinary ,Chemistry ,Kinase ,Protein Tyrosine Phosphatase, Non-Receptor Type 6 ,HEK 293 cells ,Biological Sciences ,Molecular biology ,Cell biology ,Gene Expression Regulation ,Biotechnology ,Transcription Factors - Abstract
Hypertonicity activates the transcription factor TonEBP/OREBP, resulting in increased expression of osmoprotective genes, including those responsible for accumulation of organic osmolytes and heat-shock proteins. Phosphorylation of TonEBP/OREBP contributes to its activation. Several of the kinases that are involved were previously identified, but the phosphatases were not. In the present studies we screened a genomewide human phosphatase siRNA library in human embryonic kidney (HEK)293 cells for effects on TonEBP/OREBP transcriptional activity. We found that siRNAs against 57 phosphatases significantly alter TonEBP/OREBP transcriptional activity during normotonicity (290 mosmol/kg) or hypertonicity (500 mosmol/kg, NaCl added) or both. Most siRNAs increase TonEBP/OREBP activity, implying that the targeted phosphatases normally reduce that activity. We further studied in detail SHP-1, whose knockdown by its specific siRNA increases TonEBP/OREBP transcriptional activity at 500 mosmol/kg. We confirmed that SHP-1 is inhibitory by overexpressing it, which reduces TonEBP/OREBP transcriptional activity at 500 mosmol/kg. SHP-1 dephosphorylates TonEBP/OREBP at a known regulatory site, Y143, both in vivo and in vitro. It inhibits TonEBP/OREBP by both reducing TonEBP/OREBP nuclear localization, which is Y143 dependent, and by lowering high NaCl-induced TonEBP/OREBP transactivating activity. SHP-1 coimmunoprecipitates with TonEBP/OREBP and vice versa, suggesting that they are physically associated in the cell. High NaCl inhibits the effect of SHP-1 on TonEBP/OREBP by increasing phosphorylation of SHP-1 on Ser591, which reduces its phosphatase activity and localization to the nucleus. Thus, TonEBP/OREBP is extensively regulated by phosphatases, including SHP-1, whose inhibition by high NaCl increases phosphorylation of TonEBP/OREBP at Y143, contributing to the nuclear localization and activation of TonEBP/OREBP.
- Published
- 2010
8. Inhibitory phosphorylation of GSK-3β by AKT, PKA, and PI3K contributes to high NaCl-induced activation of the transcription factor NFAT5 (TonEBP/OREBP)
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Xiaoming Zhou, Joan D. Ferraris, Maurice B. Burg, and Hong Wang
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Physiology ,Active Transport, Cell Nucleus ,AKT1 ,macromolecular substances ,Sodium Chloride ,Mitogen-Activated Protein Kinase 14 ,Glycogen Synthase Kinase 3 ,Mice ,Phosphatidylinositol 3-Kinases ,NFAT5 ,Animals ,Humans ,Phosphorylation ,Transcription factor ,GSK3B ,Protein kinase B ,PI3K/AKT/mTOR pathway ,Glycogen Synthase Kinase 3 beta ,Chemistry ,Kinase ,Articles ,Molecular biology ,Cyclic AMP-Dependent Protein Kinases ,HEK293 Cells ,Proto-Oncogene Proteins c-akt ,Transcription Factors - Abstract
High NaCl activates the transcription factor nuclear factor of activated T cells 5 (NFAT5), leading to increased transcription of osmoprotective target genes. Kinases PKA, PI3K, AKT1, and p38α were known to contribute to the high NaCl-induced increase of NFAT5 activity. We now identify another kinase, GSK-3β. siRNA-mediated knock-down of GSK-3β increases NFAT5 transcriptional and transactivating activities without affecting high NaCl-induced nuclear localization of NFAT5 or NFAT5 protein expression. High NaCl increases phosphorylation of GSK-3β-S9, which inhibits GSK-3β. In GSK-3β-null mouse embryonic fibroblasts transfection of GSK-3β, in which serine 9 is mutated to alanine, so that it cannot be inhibited by phosphorylation at that site, inhibits high NaCl-induced NFAT5 transcriptional activity more than transfection of wild-type GSK-3β. High NaCl-induced phosphorylation of GSK-3β-S9 depends on PKA, PI3K, and AKT, but not p38α. Overexpression of PKA catalytic subunit α or of catalytically active AKT1 reduces inhibition of NFAT5 by GSK-3β, but overexpression of p38α together with its catalytically active upstream kinase, MKK6, does not. Thus, GSK-3β normally inhibits NFAT5 by suppressing its transactivating activity. When activated by high NaCl, PKA, PI3K, and AKT1, but not p38α, increase phosphorylation of GSK-3β-S9, which reduces the inhibitory effect of GSK-3β on NFAT5, and thus contributes to activation of NFAT5.
- Published
- 2013
9. Rac1/osmosensing scaffold for MEKK3 contributes via phospholipase C-γ1 to activation of the osmoprotective transcription factor NFAT5
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Xiaoming Zhou, Maurice B. Burg, Yuichiro Izumi, and Joan D. Ferraris
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rac1 GTP-Binding Protein ,p38 mitogen-activated protein kinases ,MAP Kinase Kinase 3 ,Blotting, Western ,RAC1 ,Biology ,Cell Line ,Mitogen-Activated Protein Kinase 14 ,Mice ,NFAT5 ,Gene Knockdown Techniques ,Animals ,Humans ,RNA, Small Interfering ,Luciferases ,Transcription factor ,Regulation of gene expression ,Multidisciplinary ,Phospholipase C gamma ,Microfilament Proteins ,Neuropeptides ,Transfection ,Biological Sciences ,rac GTP-Binding Proteins ,Rac GTP-Binding Proteins ,Gene Expression Regulation ,Cancer research ,Transcription Factors - Abstract
Separate reports that hypertonicity activates p38 via a Rac1–OSM–MEKK3–MKK3–p38 pathway and that p38α contributes to activation of TonEBP/OREBP led us to the hypothesis that Rac1 might activate TonEBP/OREBP via p38. The present studies examine that possibility. High NaCl is hypertonic. We find that siRNA knockdown of Rac1 reduces high NaCl-induced increase of TonEBP/OREBP transcriptional activity (by reducing its transactivating activity but not its nuclear localization). Similarly, siRNA knockdown of osmosensing scaffold for MEKK3 (OSM) also reduces high NaCl-dependent TonEBP/OREBP transcriptional and transactivating activities. Simultaneous siRNA knockdown of Rac1 and OSM is not additive in reduction of TonEBP/OREBP transcriptional activity, indicating a common pathway. However, siRNA knockdown of MKK3 does not reduce TonEBP/OREBP transcriptional activity, although siRNA knockdown of MKK6 does. Nevertheless, the effect of Rac1 on TonEBP/OREBP is also independent of MKK6 because it occurs in MKK6-null cells. Furthermore, we find that siRNA knockdown of Rac1 or OSM actually increases activity (phosphorylation) of p38, rather than decreasing it, as previously reported. Thus, the effect of Rac1 on TonEBP/OREBP is independent of p38. We find instead that phospholipase C-γ1 (PLC-γ1) is involved. When transfected into PLC-γ1–null mouse embryonic fibroblast cells, catalytically active Rac1 does not increase TonEBP/OREBP transcriptional activity unless PLC-γ1 is reconstituted. Similarly, dominant-negative Rac1 also does not inhibit TonEBP/OREBP in PLC-γ1–null cells unless PLC-γ1 is reconstituted. We conclude that Rac1/OSM supports TonEBP/OREBP activity and that this activity is mediated via PLC-γ1, not p38.
- Published
- 2011
10. Rheb controls misfolded protein metabolism by inhibiting aggresome formation and autophagy
- Author
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Tsuneo Ikenoue, Kun-Liang Guan, Xiaoming Zhou, Li Li, Ken Inoki, and Xiaowei Chen
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Programmed cell death ,Protein Folding ,Green Fluorescent Proteins ,Microtubules ,Tuberous Sclerosis Complex 1 Protein ,Mice ,JUNQ and IPOD ,medicine ,Autophagy ,Animals ,Transcription factor ,PI3K/AKT/mTOR pathway ,Monomeric GTP-Binding Proteins ,Mice, Knockout ,Multidisciplinary ,biology ,Tumor Suppressor Proteins ,Neuropeptides ,Dyneins ,Biological Sciences ,Cell biology ,ErbB Receptors ,medicine.anatomical_structure ,Aggresome ,Liver ,biology.protein ,Ras Homolog Enriched in Brain Protein ,TSC1 ,RHEB ,Transcription Factors - Abstract
Perinuclear aggresome formation is a key mechanism to dispose of misfolded proteins that exceed the degradative capacity of ubiquitin–proteasome and autophagy–lysosome systems. Functional blockade of either degradative system leads to an enhanced aggresome formation. The tuberous sclerosis complex–Ras homologue enriched in brain–mammalian target of rapamycin (TSC–Rheb–mTOR) pathway is known to play a central role in modulating protein synthesis and autophagy. However, in spite of the constitutive activation of mTOR and the abrogated autophagy activity in TSC1- or TSC2-deficient cells, the TSC mutant cells are defective in aggresome formation and undergo apoptosis upon misfolded protein accumulation both in vitro and in vivo. High Rheb activity in TSC mutant cells inhibits aggresome formation and sensitizes cell death in response to misfolded proteins. Surprisingly, this previously unrecognized function of Rheb is independent of TOR complex 1. Active Rheb disrupts the interaction between dynein and misfolded protein cargos, and therefore blocks aggresome formation by inhibiting dynein-dependent transportation of misfolded proteins. This study reveals a function of Rheb in controlling misfolded protein metabolism by modulating aggresome formation.
- Published
- 2009
11. MKP-1 inhibits high NaCl-induced activation of p38 but does not inhibit the activation of TonEBP/OREBP: opposite roles of p38alpha and p38delta
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Maurice B. Burg, Yusen Liu, Xiaoming Zhou, Natalia I. Dmitrieva, and Joan D. Ferraris
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MAPK/ERK pathway ,Multidisciplinary ,NFATC Transcription Factors ,Kinase ,p38 mitogen-activated protein kinases ,Phosphatase ,Dual Specificity Phosphatase 1 ,Biology ,Sodium Chloride ,Biological Sciences ,Molecular biology ,Cell biology ,Cell Line ,Enzyme Activation ,Mitogen-Activated Protein Kinase 14 ,Enzyme activator ,Mitogen-Activated Protein Kinase 13 ,Phosphorylation ,Humans ,Transcription factor ,Transcription Factors - Abstract
High NaCl rapidly activates p38 MAPK by phosphorylating it, the phosphorylation presumably being regulated by a balance of kinases and phosphatases. Kinases are known, but the phosphatases are uncertain. Our initial purpose was to identify the phosphatases. We find that in HEK293 cells transient overexpression of MAPK phosphatase-1 (MKP-1), a dual-specificity phosphatase, inhibits high NaCl-induced phosphorylation of p38, and that overexpression of a dominant negative mutant of MKP-1 does the opposite. High NaCl lowers MKP-1 activity by increasing reactive oxygen species, which directly inhibit MKP-1, and by reducing binding of MKP-1 to p38. Because inhibition of p38 is reported to reduce hypertonicity-induced activation of the osmoprotective transcription factor, TonEBP/OREBP, we anticipated that MKP-1 expression might also. However, overexpression of MKP-1 has no significant effect on Ton EBP/OREBP activity. This paradox is explained by opposing effects of p38alpha and p38delta, both of which are activated by high NaCl and inhibited by MKP-1. Thus, we find that overexpression of p38alpha increases high NaCl-induced TonEBP/OREBP activity, but overexpression of p38delta reduces it. Also, siRNA-mediated knockdown of p38delta enhances the activation of TonEBP/OREBP. We conclude that high NaCl inhibits MKP-1, which contributes to the activation of p38. However, opposing actions of p38alpha and p38delta negate any effect on TonEBP/OREBP activity. Thus, activation of p38 isoforms by hypertonicity does not contribute to activation of TonEBP/OREBP because of opposing effects of p38alpha and p38delta, and effects of inhibitors of p38 depend on which isoform is affected, which can be misleading.
- Published
- 2008
12. Increased reactive oxygen species contribute to high NaCl-induced activation of the osmoregulatory transcription factor TonEBP/OREBP
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Maurice B. Burg, Xiaoming Zhou, Joan D. Ferraris, Qi Cai, and Anupam Agarwal
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Transcriptional Activation ,GABA Plasma Membrane Transport Proteins ,Physiology ,Biology ,Sodium Chloride ,Ouabain ,Nitric oxide ,Cell Line ,chemistry.chemical_compound ,Transactivation ,Superoxides ,Enhancer binding ,medicine ,Humans ,RNA, Messenger ,Enzyme Inhibitors ,Transcription factor ,Cells, Cultured ,chemistry.chemical_classification ,Reactive oxygen species ,Superoxide ,Water-Electrolyte Balance ,Catalase ,Oxidants ,Cell biology ,NG-Nitroarginine Methyl Ester ,chemistry ,Biochemistry ,Signal transduction ,Nitric Oxide Synthase ,Carrier Proteins ,Reactive Oxygen Species ,medicine.drug ,Transcription Factors - Abstract
The signaling pathways leading to high NaCl-induced activation of the transcription factor tonicity-responsive enhancer binding protein/osmotic response element binding protein (TonEBP/OREBP) remain incompletely understood. High NaCl has been reported to produce oxidative stress. Reactive oxygen species (ROS), which are a component of oxidative stress, contribute to regulation of transcription factors. The present study was undertaken to test whether the high NaCl-induced increase in ROS contributes to tonicity-dependent activation of TonEBP/OREBP. Human embryonic kidney 293 cells were used as a model. We find that raising NaCl increases ROS, including superoxide. N-acetylcysteine (NAC), an antioxidant, and MnTBAP, an inhibitor of superoxide, reduce high NaCl-induced superoxide activity and suppress both high NaCl-induced increase in TonEBP/OREBP transcriptional activity and high NaCl-induced increase in expression of BGT1mRNA, a transcriptional target of TonEBP/OREBP. Catalase, which decomposes hydrogen peroxide, does not have these effects, whether applied exogenously or overexpressed within the cells. Furthermore, NAC and MnTBAP, but not catalase, blunt high NaCl-induced increase in TonEBP/OREBP transactivation. NG-monomethyl-l-arginine, a general inhibitor of nitric oxide synthase, has no significant effect on either high NaCl-induced increase in superoxide or TonEBP/OREBP transcriptional activity, suggesting that the effects of ROS do not involve nitric oxide. Ouabain, an inhibitor of Na-K-ATPase, attenuates high NaCl-induced superoxide activity and inhibits TonEBP/OREBP transcriptional activity. We conclude that the high NaCl-induced increase in ROS, including superoxide, contributes to activation of TonEBP/OREBP by increasing its transactivation.
- Published
- 2005
13. PKC-α contributes to high NaCl-induced activation of NFAT5 (TonEBP/OREBP) through MAPK ERK1/2.
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Hong Wang, Ferraris, Joan D., Klein, Janet D., Sands, Jeff M., Burg, Maurice B., and Xiaoming Zhou
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KIDNEY cortex ,TRANSCRIPTION factors ,EXTRACELLULAR fluid ,PHOSPHORYLATION ,T cells ,GENE expression - Abstract
High NaCl in the renal medullary interstitial fluid powers the concentration of urine but can damage cells. The transcription factor nuclear factor of activated T cells 5 (NFAT5) activates the expression of osmoprotective genes. We studied whether PKC-α contributes to the activation of NFAT5. PKC-α protein abundance was greater in the renal medulla than in the cortex. Knockout of PKC-α reduced NFAT5 protein abundance and expression of its target genes in the inner medulla. In human embryonic kidney (HEK)-293 cells, high NaCl increased PKC-α activity, and small interfering RNA-mediated knockdown of PKC-α attenuated high NaCl-induced NFAT5 transcriptional activity. Expression of ERK1/2 protein and phosphorylation of ERK1/2 were higher in the renal inner medulla than in the cortex. Knockout of PKC-α decreased ERK1/2 phosphorylation in the inner medulla, as did knockdown of PKC-α in HEK-293 cells. Also, knockdown of ERK2 reduced high NaCl-dependent NFAT5 transcriptional activity in HEK-293 cells. Combined knockdown of PKC-α and ERK2 had no greater effect than knockdown of either alone. Knockdown of either PKC-α or ERK2 reduced the high NaCl-induced increase of NFAT5 transactivating activity. We have previously found that the high NaCl-induced increase of phosphorylation of Ser591 on Src homology 2 domaincontaining phosphatase 1 (SHP-1-S591-P) contributes to the activation of NFAT5 in cell culture, and here we found high levels of SHP-1-S591-P in the inner medulla. PKC-α has been previously shown to increase SHP-1-S591-P, which raised the possibility that PKC-α might be acting through SHP-1. However, we did not find that knockout of PKC-α in the renal medulla or knockdown in HEK-293 cells affected SHP-1-S591-P. We conclude that PKC-α contributes to high NaCl-dependent activation of NFAT5 through ERK1/2 but not through SHP-1-S591. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
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14. Contribution of SHP-1 protein tyrosine phosphatase to osmotic regulation of the transcription factor TonEBP/OREBP.
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Xiaoming Zhou, Gallazzini, Morgan, Burg, Maurice B., and Ferraris, Joan D.
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TRANSCRIPTION factors , *HEAT shock proteins , *PHOSPHATASES , *CELLS , *PHOSPHORYLATION - Abstract
Hypertonicity activates the transcription factor TonEBP/OREBP, resulting in increased expression of osmoprotective genes, including those responsible for accumulation of organic osmolytes and heat-shock proteins. Phosphorylation of TonEBP/OREBP contributes to its activation. Several of the kinases that are involved were previously identified, but the phosphatases were not. In the present studies we screened a genomewide human phosphatase siRNA library in human embryonic kidney (HEK)293 cells for effects on TonEBP/OREBP transcriptional activity. We found that siRNAs against 57 phosphatases significantly alter TonEBP/OREBP transcriptional activity during normotonicity (290 mosmol/kg) or hypertonicity (500 mosmol/kg, NaCl added) or both. Most siRNAs increase TonEBP/OREBP activity, implying that the targeted phosphatases normally reduce that activity. We further studied in detail SHP-1, whose knockdown by its specific siRNA increases TonEBP/OREBP transcriptional activity at 500 mosmol/kg. We confirmed that SHP-1 is inhibitory by overexpressing it, which reduces TonEBP/OREBP transcriptional activity at 500 mosmol/kg. SHP-1 dephosphorylates TonEBP/OREBP at a known regulatory site, Y143, both in vivo and in vitro. It inhibits TonEBP/OREBP by both reducing TonEBP/OREBP nuclear localization, which is Y143 dependent, and by lowering high NaCl-induced TonEBP/OREBP transactivating activity. SHP-1 coimmunoprecipitates with TonEBP/OREBP and vice versa, suggesting that they are physically associated in the cell. High NaCl inhibits the effect of SHP-1 on TonEBP/OREBP by increasing phosphorylation of SHP-1 on Ser591, which reduces its phosphatase activity and localization to the nucleus. Thus, TonEBP/OREBP is extensively regulated by phosphatases, including SHP-1, whose inhibition by high NaCl increases phosphorylation of TonEBP/OREBP at Y143, contributing to the nuclear localization and activation of TonEBP/OREBP. [ABSTRACT FROM AUTHOR]
- Published
- 2010
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15. MKP-1 inhibits high NaCI-induced activation of p38 but does not inhibit the activation of TonEBP/OREBP: Opposite roles of p38α and p38δ.
- Author
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Xiaoming Zhou, Ferraris, Joan D., Dmitrieva, Natalia I., Yusen Liu, and Burg, Maurice B.
- Subjects
- *
PHOSPHORYLATION , *REACTIVE oxygen species , *ESTERASES , *TRANSCRIPTION factors , *PHOTOSYNTHETIC oxygen evolution , *CHEMICAL reactions - Abstract
High NaCl rapidly activates p38 MAPK by phosphorylating it, the phosphorylation presumably being regulated by a balance of kinases and phosphatases. Kinases are known, but the phosphatases are uncertain. Our initial purpose was to identify the phosphatases. We find that in HEK293 cells transient overexpression of MAPK phosphatase-1 (MKP-1), a dual-specificity phosphatase, inhibits high NaCl-induced phosphorylation of p38, and that over-expression of a dominant negative mutant of MKP-1 does the opposite. High NaCl lowers MKP-1 activity by increasing reactive oxygen species, which directly inhibit MKP-1, and by reducing binding of MKP-1 to p38. Because inhibition of p38 is reported to reduce hypertonicity-induced activation of the osmoprotective transcription factor, TonEBP/OREBP, we anticipated that MKP-1 expression might also. However, overexpression of MKP-1 has no significant effect on Ton EBP/OREBP activity. This paradox is explained by opposing effects of p38α and p38δ, both of which are activated by high NaCl and inhibited by MKP-1. Thus, we find that overexpression of p38α increases high NaCI-induced TonEBP/OREBP activity, but overexpression of p38δ reduces it. Also, siRNA-mediated knockdown of p38δ enhances the activation of TonEBP/OREBP. We conclude that high NaCl inhibits MKP-1, which contributes to the activation of p38. However, opposing actions of p38α and p38δ negate any effect on TonEBP/OREBP activity. Thus, activation of p38 isoforms by hypertonicity does not contribute to activation of TonEBP/OREBP because of opposing effects of p38α and p38δ, and effects of inhibitors of p38 depend on which isoform is affected, which can be misleading. [ABSTRACT FROM AUTHOR]
- Published
- 2008
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16. Mitochondrial reactive oxygen species contribute to high NaC1-induced activation of the transcription factor TonEBP/OREBP.
- Author
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Xiaoming Zhou, Ferraris, Joan D., and Burg, Maurice B.
- Subjects
- *
REACTIVE oxygen species , *HYPERTONIC solutions , *MITOCHONDRIA , *TRANSCRIPTION factors , *CARRIER proteins , *HEAT shock proteins - Abstract
Hypertonicity activates the transcription factor tonicity-responsive enhancer/ osmotic response element binding protein (TonEBP/OREBP), resulting in increased expression of genes involved in osmoprotective accumulation of organic osmolytes, including glycine betaine, and in increased expression of osmoprotective heat shock proteins. Our previous studies showed that high NaCl increases reactive oxygen species (ROS), which contribute to activation of TonEBP/OREBP. Mitochondria are a major source of ROS. The purpose of the present study was to examine whether mitochondria produce the ROS that contribute to activation of TonEBP/OREBP. We inhibited mitochondrial ROS production in HEK293 cells with rotenone and myxothiazol, which inhibit mitochondrial complexes I and III, respectively. Rotenone (250 nM) and myxothiazol (12 nM) reduce high NaCl-induced ROS over 40%, whereas apocynin (100 μM), an inhibitor of NADPH oxidase, and allopurinol (100 μM), an inhibitor of xanthine oxidase, have no significant effect. Rotenone and myxothiazol reduce high NaCl-induced increases in TonEBP/OREBP transcriptional activity (ORE/TonE reporter assay) and BGT1 (betaine transporter) mRNA abundance ranging from 53 to 69%. They inhibit high NaCl-induced TonEBP/OREBP transactivating activity, but not its nuclear translocation. Release of ATP into the medium on hypertonic stress has been proposed to be a signal that triggers cellular osmotic responses. However, we do not detect release of ATP into the medium or inhibition of high NaCl-induced ORE/TonE reporter activity by an ATPase, apyrase (20 U/ml), indicating that high NaCl-induced activation of TonEBP/OREBP is not mediated by release of ATP. We conclude that high NaCl increases mitochondrial ROS production, which contributes to the activation of TonEBP/OREBP by increasing its transactivating activity. [ABSTRACT FROM AUTHOR]
- Published
- 2006
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17. Increased reactive oxygen species contribute to high NaCl-induced activation of the osmoregulatory transcription factor TonEBP/OREBP.
- Author
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Xiaoming Zhou, Ferraris, Joan D., Qi Cai, Agarwal, Anupam, and Burg, Maurice B.
- Subjects
- *
REACTIVE oxygen species , *SALT , *OSMOREGULATION , *TRANSCRIPTION factors , *NEPHROLOGY , *PHYSIOLOGY , *CARRIER proteins , *SUPEROXIDES , *CHROMOSOMAL translocation - Abstract
The signaling pathways leading to high NaCl-induced activation of the transcription factor tonicity-responsive enhancer binding protein/osmotic response element binding protein (TonEBP/OREBP) remain incompletely understood. High NaCl has been reported to produce oxidative stress. Reactive oxygen species (ROS), which are a component of oxidative stress, contribute to regulation of transcription factors. The present study was undertaken to test whether the high NaCl-induced increase in ROS contributes to tonicity-dependent activation of TonEBP/OREBP. Human embryonic kidney 293 cells were used as a model. We find that raising NaCl increases ROS, including superoxide. N-acetylcysteine (NAC), an antioxidant, and MnTBAP, an inhibitor of superoxide, reduce high NaCl-induced superoxide activity and suppress both high NaCl-induced increase in TonEBP/OREBP transcriptional activity and high NaCl-induced increase in expression of BGT1mRNA, a transcriptional target of TonEBP/OREBP. Catalase, which decomposes hydrogen peroxide, does not have these effects, whether applied exogenously or overexpressed within the cells. Furthermore, NAC and MnTBAP, but not catalase, blunt high NaCl-induced increase in TonEBP/OREBP transactivation. NG-monomethyl-L-arginine, a general inhibitor of nitric oxide synthase, has no significant effect on either high NaCl-induced increase in superoxide or TonEBP/OREBP transcriptional activity, suggesting that the effects of ROS do not involve nitric oxide. Ouabain, an inhibitor of Na-K-ATPase, attenuates high NaCl-induced superoxide activity and inhibits TonEBP/OREBP transcriptional activity. We conclude that the high NaCl-induced increase in ROS, including superoxide, contributes to activation of TonEBP/OREBP by increasing its transactivation. [ABSTRACT FROM AUTHOR]
- Published
- 2005
- Full Text
- View/download PDF
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