59 results on '"Näär AM"'
Search Results
2. Functional antagonism between histone H3K4 demethylases in vivo
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Anders M. Näär, Peter Mulligan, James A. Walker, Davide Corona, Giosalba Burgio, Nicholas J. Dyson, Luisa Di Stefano, Di Stefano, L, Walker, JA, Burgio, G, Corona, D, Mulligan, P, Näär, AM, and Dyson NJ
- Subjects
Settore BIO/11 - Biologia Molecolare ,Biology ,Methylation ,Histones ,histone demethylases ,Histone H3 ,Heterochromatin ,Histone H2A ,Histone methylation ,Genetics ,Animals ,Drosophila Proteins ,Histone code ,Receptors, Notch ,EZH2 ,Oxidoreductases, N-Demethylating ,Histone-Lysine N-Methyltransferase ,Settore BIO/18 - Genetica ,Drosophila melanogaster ,Phenotype ,Gene Expression Regulation ,Histone methyltransferase ,Mutation ,Heterochromatin protein 1 ,Histone Demethylases ,Signal Transduction ,Research Paper ,Developmental Biology - Abstract
Dynamic regulation of histone modifications is critical during development, and aberrant activity of chromatin-modifying enzymes has been associated with diseases such as cancer. Histone demethylases have been shown to play a key role in eukaryotic gene transcription; however, little is known about how their activities are coordinated in vivo to regulate specific biological processes. In Drosophila, two enzymes, dLsd1 (Drosophila ortholog of lysine-specific demethylase 1) and Lid (little imaginal discs), demethylate histone H3 at Lys 4 (H3K4), a residue whose methylation is associated with actively transcribed genes. Our studies show that compound mutation of Lid and dLsd1 results in increased H3K4 methylation levels. However, unexpectedly, Lid mutations strongly suppress dLsd1 mutant phenotypes. Investigation of the basis for this antagonism revealed that Lid opposes the functions of dLsd1 and the histone methyltransferase Su(var)3–9 in promoting heterochromatin spreading at heterochromatin–euchromatin boundaries. Moreover, our data reveal a novel role for dLsd1 in Notch signaling in Drosophila, and a complex network of interactions between dLsd1, Lid, and Notch signaling at euchromatic genes. These findings illustrate the complexity of functional interplay between histone demethylases in vivo, providing insights into the epigenetic regulation of heterochromatin/euchromatin boundaries by Lid and dLsd1 and showing their involvement in Notch pathway-specific control of gene expression in euchromatin.
- Published
- 2011
3. MicroRNA-22 Is a Key Regulator of Lipid and Metabolic Homeostasis.
- Author
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Panella R, Petri A, Desai BN, Fagoonee S, Cotton CA, Nguyen PK, Lundin EM, Wagshal A, Wang DZ, Näär AM, Vlachos IS, Maratos-Flier E, Altruda F, Kauppinen S, and Pandolfi PP
- Subjects
- Animals, Mice, Homeostasis, Mice, Transgenic, Obesity genetics, Lipids, Diabetes Mellitus, Type 2, Non-alcoholic Fatty Liver Disease genetics, MicroRNAs genetics
- Abstract
Obesity is a growing public health problem associated with increased risk of type 2 diabetes, cardiovascular disease, nonalcoholic fatty liver disease (NAFLD) and cancer. Here, we identify microRNA-22 (miR-22) as an essential rheostat involved in the control of lipid and energy homeostasis as well as the onset and maintenance of obesity. We demonstrate through knockout and transgenic mouse models that miR-22 loss-of-function protects against obesity and hepatic steatosis, while its overexpression promotes both phenotypes even when mice are fed a regular chow diet. Mechanistically, we show that miR-22 controls multiple pathways related to lipid biogenesis and differentiation. Importantly, genetic ablation of miR-22 favors metabolic rewiring towards higher energy expenditure and browning of white adipose tissue, suggesting that modulation of miR-22 could represent a viable therapeutic strategy for treatment of obesity and other metabolic disorders.
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- 2023
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4. SARS-CoV-2 Spike triggers barrier dysfunction and vascular leak via integrins and TGF-β signaling.
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Biering SB, Gomes de Sousa FT, Tjang LV, Pahmeier F, Zhu C, Ruan R, Blanc SF, Patel TS, Worthington CM, Glasner DR, Castillo-Rojas B, Servellita V, Lo NTN, Wong MP, Warnes CM, Sandoval DR, Clausen TM, Santos YA, Fox DM, Ortega V, Näär AM, Baric RS, Stanley SA, Aguilar HC, Esko JD, Chiu CY, Pak JE, Beatty PR, and Harris E
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- Humans, Angiotensin-Converting Enzyme 2, Spike Glycoprotein, Coronavirus genetics, Endothelial Cells, Integrins, Peptidyl-Dipeptidase A genetics, Transforming Growth Factor beta, SARS-CoV-2, COVID-19
- Abstract
Severe COVID-19 is associated with epithelial and endothelial barrier dysfunction within the lung as well as in distal organs. While it is appreciated that an exaggerated inflammatory response is associated with barrier dysfunction, the triggers of vascular leak are unclear. Here, we report that cell-intrinsic interactions between the Spike (S) glycoprotein of SARS-CoV-2 and epithelial/endothelial cells are sufficient to induce barrier dysfunction in vitro and vascular leak in vivo, independently of viral replication and the ACE2 receptor. We identify an S-triggered transcriptional response associated with extracellular matrix reorganization and TGF-β signaling. Using genetic knockouts and specific inhibitors, we demonstrate that glycosaminoglycans, integrins, and the TGF-β signaling axis are required for S-mediated barrier dysfunction. Notably, we show that SARS-CoV-2 infection caused leak in vivo, which was reduced by inhibiting integrins. Our findings offer mechanistic insight into SARS-CoV-2-triggered vascular leak, providing a starting point for development of therapies targeting COVID-19., (© 2022. The Author(s).)
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- 2022
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5. Discovery of SARS-CoV-2 antiviral synergy between remdesivir and approved drugs in human lung cells.
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Nguyenla X, Wehri E, Van Dis E, Biering SB, Yamashiro LH, Zhu C, Stroumza J, Dugast-Darzacq C, Graham TGW, Wang X, Jockusch S, Tao C, Chien M, Xie W, Patel DJ, Meyer C, Garzia A, Tuschl T, Russo JJ, Ju J, Näär AM, Stanley S, and Schaletzky J
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- Humans, SARS-CoV-2, Antiviral Agents therapeutic use, Sofosbuvir pharmacology, Nucleosides pharmacology, Adenosine Monophosphate, Alanine, Hepacivirus, Lung, Hepatitis C drug therapy, COVID-19 Drug Treatment
- Abstract
SARS coronavirus 2 (SARS-CoV-2) has caused an ongoing global pandemic with significant mortality and morbidity. At this time, the only FDA-approved therapeutic for COVID-19 is remdesivir, a broad-spectrum antiviral nucleoside analog. Efficacy is only moderate, and improved treatment strategies are urgently needed. To accomplish this goal, we devised a strategy to identify compounds that act synergistically with remdesivir in preventing SARS-CoV-2 replication. We conducted combinatorial high-throughput screening in the presence of submaximal remdesivir concentrations, using a human lung epithelial cell line infected with a clinical isolate of SARS-CoV-2. This identified 20 approved drugs that act synergistically with remdesivir, many with favorable pharmacokinetic and safety profiles. Strongest effects were observed with established antivirals, Hepatitis C virus nonstructural protein 5A (HCV NS5A) inhibitors velpatasvir and elbasvir. Combination with their partner drugs sofosbuvir and grazoprevir further increased efficacy, increasing remdesivir's apparent potency > 25-fold. We report that HCV NS5A inhibitors act on the SARS-CoV-2 exonuclease proofreader, providing a possible explanation for the synergy observed with nucleoside analog remdesivir. FDA-approved Hepatitis C therapeutics Epclusa® (velpatasvir/sofosbuvir) and Zepatier® (elbasvir/grazoprevir) could be further optimized to achieve potency and pharmacokinetic properties that support clinical evaluation in combination with remdesivir., (© 2022. The Author(s).)
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- 2022
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6. An intranasal ASO therapeutic targeting SARS-CoV-2.
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Zhu C, Lee JY, Woo JZ, Xu L, Nguyenla X, Yamashiro LH, Ji F, Biering SB, Van Dis E, Gonzalez F, Fox D, Wehri E, Rustagi A, Pinsky BA, Schaletzky J, Blish CA, Chiu C, Harris E, Sadreyev RI, Stanley S, Kauppinen S, Rouskin S, and Näär AM
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- Administration, Intranasal, Animals, Humans, Mice, Oligonucleotides, Antisense pharmacology, Oligonucleotides, Antisense therapeutic use, Pandemics prevention & control, RNA, Viral genetics, COVID-19, SARS-CoV-2
- Abstract
The COVID-19 pandemic is exacting an increasing toll worldwide, with new SARS-CoV-2 variants emerging that exhibit higher infectivity rates and that may partially evade vaccine and antibody immunity. Rapid deployment of non-invasive therapeutic avenues capable of preventing infection by all SARS-CoV-2 variants could complement current vaccination efforts and help turn the tide on the COVID-19 pandemic. Here, we describe a novel therapeutic strategy targeting the SARS-CoV-2 RNA using locked nucleic acid antisense oligonucleotides (LNA ASOs). We identify an LNA ASO binding to the 5' leader sequence of SARS-CoV-2 that disrupts a highly conserved stem-loop structure with nanomolar efficacy in preventing viral replication in human cells. Daily intranasal administration of this LNA ASO in the COVID-19 mouse model potently suppresses viral replication (>80-fold) in the lungs of infected mice. We find that the LNA ASO is efficacious in countering all SARS-CoV-2 "variants of concern" tested both in vitro and in vivo. Hence, inhaled LNA ASOs targeting SARS-CoV-2 represents a promising therapeutic approach to reduce or prevent transmission and decrease severity of COVID-19 in infected individuals. LNA ASOs are chemically stable and can be flexibly modified to target different viral RNA sequences and could be stockpiled for future coronavirus pandemics., (© 2022. The Author(s).)
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- 2022
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7. Targeting of miR-33 ameliorates phenotypes linked to age-related macular degeneration.
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Gnanaguru G, Wagschal A, Oh J, Saez-Torres KL, Li T, Temel RE, Kleinman ME, Näär AM, and D'Amore PA
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- Animals, Inflammation etiology, Inflammation pathology, Macaca fascicularis, Macular Degeneration etiology, Macular Degeneration pathology, Male, Mice, Mice, Inbred C57BL, MicroRNAs genetics, Oligonucleotides, Antisense genetics, ATP-Binding Cassette Transporters metabolism, Cholesterol metabolism, Inflammation therapy, Macular Degeneration therapy, MicroRNAs antagonists & inhibitors, Phenotype, Retinal Pigment Epithelium metabolism
- Abstract
Abnormal cholesterol/lipid homeostasis is linked to neurodegenerative conditions such as age-related macular degeneration (AMD), which is a leading cause of blindness in the elderly. The most prevalent form, termed "dry" AMD, is characterized by pathological cholesterol accumulation beneath the retinal pigment epithelial (RPE) cell layer and inflammation-linked degeneration in the retina. We show here that the cholesterol-regulating microRNA miR-33 was elevated in the RPE of aging mice. Expression of the miR-33 target ATP-binding cassette transporter (ABCA1), a cholesterol efflux pump genetically linked to AMD, declined reciprocally in the RPE with age. In accord, miR-33 modulated ABCA1 expression and cholesterol efflux in human RPE cells. Subcutaneous delivery of miR-33 antisense oligonucleotides (ASO) to aging mice and non-human primates fed a Western-type high fat/cholesterol diet resulted in increased ABCA1 expression, decreased cholesterol accumulation, and reduced immune cell infiltration in the RPE cell layer, accompanied by decreased pathological changes to RPE morphology. These findings suggest that miR-33 targeting may decrease cholesterol deposition and ameliorate AMD initiation and progression., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)
- Published
- 2021
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8. The Lipogenic Regulator SREBP2 Induces Transferrin in Circulating Melanoma Cells and Suppresses Ferroptosis.
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Hong X, Roh W, Sullivan RJ, Wong KHK, Wittner BS, Guo H, Dubash TD, Sade-Feldman M, Wesley B, Horwitz E, Boland GM, Marvin DL, Bonesteel T, Lu C, Aguet F, Burr R, Freeman SS, Parida L, Calhoun K, Jewett MK, Nieman LT, Hacohen N, Näär AM, Ting DT, Toner M, Stott SL, Getz G, Maheswaran S, and Haber DA
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- Biomarkers, Tumor, Cells, Cultured, Disease Susceptibility, Gene Expression Regulation, Neoplastic, Gene Knockdown Techniques, Humans, Melanoma pathology, Mutation, Neoplastic Cells, Circulating pathology, Signal Transduction, Single-Cell Analysis, Sterol Regulatory Element Binding Protein 2 metabolism, Ferroptosis genetics, Lipogenesis genetics, Melanoma genetics, Melanoma metabolism, Neoplastic Cells, Circulating metabolism, Sterol Regulatory Element Binding Protein 2 genetics, Transferrin metabolism
- Abstract
Circulating tumor cells (CTC) are shed by cancer into the bloodstream, where a viable subset overcomes oxidative stress to initiate metastasis. We show that single CTCs from patients with melanoma coordinately upregulate lipogenesis and iron homeostasis pathways. These are correlated with both intrinsic and acquired resistance to BRAF inhibitors across clonal cultures of BRAF -mutant CTCs. The lipogenesis regulator SREBP2 directly induces transcription of the iron carrier Transferrin ( TF ), reducing intracellular iron pools, reactive oxygen species, and lipid peroxidation, thereby conferring resistance to inducers of ferroptosis. Knockdown of endogenous TF impairs tumor formation by melanoma CTCs, and their tumorigenic defects are partially rescued by the lipophilic antioxidants ferrostatin-1 and vitamin E. In a prospective melanoma cohort, presence of CTCs with high lipogenic and iron metabolic RNA signatures is correlated with adverse clinical outcome, irrespective of treatment regimen. Thus, SREBP2-driven iron homeostatic pathways contribute to cancer progression, drug resistance, and metastasis. SIGNIFICANCE: Through single-cell analysis of primary and cultured melanoma CTCs, we have uncovered intrinsic cancer cell heterogeneity within lipogenic and iron homeostatic pathways that modulates resistance to BRAF inhibitors and to ferroptosis inducers. Activation of these pathways within CTCs is correlated with adverse clinical outcome, pointing to therapeutic opportunities. This article is highlighted in the In This Issue feature, p. 521 ., (©2020 American Association for Cancer Research.)
- Published
- 2021
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9. A multi-pronged approach targeting SARS-CoV-2 proteins using ultra-large virtual screening.
- Author
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Gorgulla C, Padmanabha Das KM, Leigh KE, Cespugli M, Fischer PD, Wang ZF, Tesseyre G, Pandita S, Shnapir A, Calderaio A, Gechev M, Rose A, Lewis N, Hutcheson C, Yaffe E, Luxenburg R, Herce HD, Durmaz V, Halazonetis TD, Fackeldey K, Patten JJ, Chuprina A, Dziuba I, Plekhova A, Moroz Y, Radchenko D, Tarkhanova O, Yavnyuk I, Gruber C, Yust R, Payne D, Näär AM, Namchuk MN, Davey RA, Wagner G, Kinney J, and Arthanari H
- Abstract
The unparalleled global effort to combat the continuing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic over the last year has resulted in promising prophylactic measures. However, a need still exists for cheap, effective therapeutics, and targeting multiple points in the viral life cycle could help tackle the current, as well as future, coronaviruses. Here, we leverage our recently developed, ultra-large-scale in silico screening platform, VirtualFlow, to search for inhibitors that target SARS-CoV-2. In this unprecedented structure-based virtual campaign, we screened roughly 1 billion molecules against each of 40 different target sites on 17 different potential viral and host targets. In addition to targeting the active sites of viral enzymes, we also targeted critical auxiliary sites such as functionally important protein-protein interactions., Competing Interests: M.C. and V.D. report working for Innophore. C.C.G. reports being a shareholder and CEO of Innophore, an enzyme and drug discovery company. G.W. and C.G. report being co-founders of the company Virtual Discovery, Inc., which provides virtual screening services. G.W. reports serving as the director of this company. G.W. reports being a co-founder of PIC Therapeutics, Cellmig Biolabs, and Skinap Therapeutics. H.A. reports being an equity holder in PIC Therapeutics. I.I. and D.R. report working for Enamine, a company that is involved in the synthesis and distribution of chemical building blocks, fragments, and screening compounds. Y.M. reports being a scientific advisor for Enamine. Y.M., O.T., and A.P. report working for Chemspace, a company that is involved in the distribution of chemical building blocks, fragments, and screening compounds. I.D. reports working for UkrOrgSyntez Ltd. (UORSY), a company that is involved in the synthesis of chemical building blocks, fragments, and screening compounds. G.T., S.P., A.S., M.G., N.L., C.H., E.Y., R.L., R.Y., D.P., and J.K. report working for Google, a company that also provides cloud computing services. The research described here is scientifically and financially independent of the efforts in any of the above mentioned companies., (© 2020 The Author(s).)
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- 2021
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10. A MicroRNA Linking Human Positive Selection and Metabolic Disorders.
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Wang L, Sinnott-Armstrong N, Wagschal A, Wark AR, Camporez JP, Perry RJ, Ji F, Sohn Y, Oh J, Wu S, Chery J, Moud BN, Saadat A, Dankel SN, Mellgren G, Tallapragada DSP, Strobel SM, Lee MJ, Tewhey R, Sabeti PC, Schaefer A, Petri A, Kauppinen S, Chung RT, Soukas A, Avruch J, Fried SK, Hauner H, Sadreyev RI, Shulman GI, Claussnitzer M, and Näär AM
- Subjects
- Adipocytes, Brown pathology, Adiposity, Alleles, Animals, Cell Differentiation, Cell Line, Cells, Cultured, Diet, High-Fat, Energy Metabolism, Epigenesis, Genetic, Genetic Loci, Glucose metabolism, Homeostasis, Humans, Hypertrophy, Insulin Resistance, Leptin deficiency, Leptin metabolism, Male, Mammals genetics, Mice, Inbred C57BL, Mice, Obese, MicroRNAs metabolism, Obesity genetics, Oligonucleotides metabolism, Species Specificity, Metabolic Diseases genetics, MicroRNAs genetics
- Abstract
Positive selection in Europeans at the 2q21.3 locus harboring the lactase gene has been attributed to selection for the ability of adults to digest milk to survive famine in ancient times. However, the 2q21.3 locus is also associated with obesity and type 2 diabetes in humans, raising the possibility that additional genetic elements in the locus may have contributed to evolutionary adaptation to famine by promoting energy storage, but which now confer susceptibility to metabolic diseases. We show here that the miR-128-1 microRNA, located at the center of the positively selected locus, represents a crucial metabolic regulator in mammals. Antisense targeting and genetic ablation of miR-128-1 in mouse metabolic disease models result in increased energy expenditure and amelioration of high-fat-diet-induced obesity and markedly improved glucose tolerance. A thrifty phenotype connected to miR-128-1-dependent energy storage may link ancient adaptation to famine and modern metabolic maladaptation associated with nutritional overabundance., Competing Interests: Declaration of Interests A.M.N. has issued patents on miR-128-1 (U.S. Pat. Nos. 9,045,749; 9,476,046; 9,789,132)., (Copyright © 2020 Elsevier Inc. All rights reserved.)
- Published
- 2020
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11. A Multi-Pronged Approach Targeting SARS-CoV-2 Proteins Using Ultra-Large Virtual Screening.
- Author
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Gorgulla C, Padmanabha Das KM, Leigh KE, Cespugli M, Fischer PD, Wang ZF, Tesseyre G, Pandita S, Shnapir A, Calderaio A, Gechev M, Rose A, Lewis N, Hutcheson C, Yaffe E, Luxenburg R, Herce HD, Durmaz V, Halazonetis TD, Fackeldey K, Patten JJ, Chuprina A, Dziuba I, Plekhova A, Moroz Y, Radchenko D, Tarkhanova O, Yavnyuk I, Gruber C, Yust R, Payne D, Näär AM, Namchuk MN, Davey RA, Wagner G, Kinney J, and Arthanari H
- Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), previously known as 2019 novel coronavirus (2019-nCoV), has spread rapidly across the globe, creating an unparalleled global health burden and spurring a deepening economic crisis. As of July 7th, 2020, almost seven months into the outbreak, there are no approved vaccines and few treatments available. Developing drugs that target multiple points in the viral life cycle could serve as a strategy to tackle the current as well as future coronavirus pandemics. Here we leverage the power of our recently developed in silico screening platform, VirtualFlow, to identify inhibitors that target SARS-CoV-2. VirtualFlow is able to efficiently harness the power of computing clusters and cloud-based computing platforms to carry out ultra-large scale virtual screens. In this unprecedented structure-based multi-target virtual screening campaign, we have used VirtualFlow to screen an average of approximately 1 billion molecules against each of 40 different target sites on 17 different potential viral and host targets in the cloud. In addition to targeting the active sites of viral enzymes, we also target critical auxiliary sites such as functionally important protein-protein interaction interfaces. This multi-target approach not only increases the likelihood of finding a potent inhibitor, but could also help identify a collection of anti-coronavirus drugs that would retain efficacy in the face of viral mutation. Drugs belonging to different regimen classes could be combined to develop possible combination therapies, and top hits that bind at highly conserved sites would be potential candidates for further development as coronavirus drugs. Here, we present the top 200 in silico hits for each target site. While in-house experimental validation of some of these compounds is currently underway, we want to make this array of potential inhibitor candidates available to researchers worldwide in consideration of the pressing need for fast-tracked drug development., Competing Interests: Alexander Chuprina, Dmytro Radchenko, and Iryna Iavniuk work for Enamine, Kyiv Ukraine. Igor Dziuba works for UkrOrgSyntez Ltd, Kyiv Ukraine. Olga Tarkhanova, Alla Plekhova, and Yurii Moroz work for Chemspace Kyiv, Ukraine. Enamine, UkrOrgSyntez, and Chemspace are companies that are involved in the synthesis and distribution of drug-like compounds. Yurii Moroz is a scientific advisor for Enamine.
- Published
- 2020
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12. SREBP1-dependent de novo fatty acid synthesis gene expression is elevated in malignant melanoma and represents a cellular survival trait.
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Wu S and Näär AM
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- Cell Line, Tumor, Cell Survival, Extracellular Signal-Regulated MAP Kinases antagonists & inhibitors, Humans, Indazoles pharmacology, Kaplan-Meier Estimate, Melanoma genetics, Melanoma mortality, Neoplasm Proteins antagonists & inhibitors, Neoplasms genetics, Piperazines pharmacology, Prognosis, Proto-Oncogene Proteins B-raf antagonists & inhibitors, RNA Interference, RNA Polymerase II metabolism, RNA, Small Interfering genetics, Skin Neoplasms genetics, Skin Neoplasms mortality, Transcription, Genetic, Vemurafenib pharmacology, Fatty Acids biosynthesis, Gene Expression Regulation, Neoplastic drug effects, Melanoma metabolism, Neoplasm Proteins physiology, Skin Neoplasms metabolism, Sterol Regulatory Element Binding Protein 1 physiology
- Abstract
de novo fatty acid biosynthesis (DNFA) is a hallmark adaptation of many cancers that supports survival, proliferation, and metastasis. Here we elucidate previously unexplored aspects of transcription regulation and clinical relevance of DNFA in cancers. We show that elevated expression of DNFA genes is characteristic of many tumor types and correlates with poor prognosis, especially in melanomas. Elevated DNFA gene expression depends on the SREBP1 transcription factor in multiple melanoma cell lines. SREBP1 predominantly binds to the transcription start sites of DNFA genes, regulating their expression by recruiting RNA polymerase II to promoters for productive transcription elongation. We find that SREBP1-regulated DNFA represents a survival trait in melanoma cells, regardless of proliferative state and oncogenic mutation status. Indeed, malignant melanoma cells exhibit elevated DNFA gene expression after the BRAF/MEK signaling pathway is blocked (e.g. by BRAF inhibitors), and DNFA expression remains higher in melanoma cells resistant to vemurafenib treatment than in untreated cells. Accordingly, DNFA pathway inhibition, whether by direct targeting of SREBP1 with antisense oligonucleotides, or through combinatorial effects of multiple DNFA enzyme inhibitors, exerts potent cytotoxic effects on both BRAFi-sensitive and -resistant melanoma cells. Altogether, these results implicate SREBP1 and DNFA enzymes as enticing therapeutic targets in melanomas.
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- 2019
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13. Insulin Receptor Associates with Promoters Genome-wide and Regulates Gene Expression.
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Hancock ML, Meyer RC, Mistry M, Khetani RS, Wagschal A, Shin T, Ho Sui SJ, Näär AM, and Flanagan JG
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- Animals, Cell Line, Tumor, Chromatin metabolism, Host Cell Factor C1 antagonists & inhibitors, Host Cell Factor C1 genetics, Host Cell Factor C1 metabolism, Humans, Insulin metabolism, Insulin pharmacology, Liver metabolism, Male, Mice, Mice, Inbred C57BL, Promoter Regions, Genetic, Protein Binding, Protein Subunits metabolism, RNA Interference, RNA Polymerase II metabolism, RNA, Small Interfering metabolism, Receptor, Insulin chemistry, Signal Transduction drug effects, Gene Expression Regulation drug effects, Genome-Wide Association Study, Receptor, Insulin metabolism
- Abstract
Insulin receptor (IR) signaling is central to normal metabolic control and dysregulated in prevalent chronic diseases. IR binds insulin at the cell surface and transduces rapid signaling via cytoplasmic kinases. However, mechanisms mediating long-term effects of insulin remain unclear. Here, we show that IR associates with RNA polymerase II in the nucleus, with striking enrichment at promoters genome-wide. The target genes were highly enriched for insulin-related functions including lipid metabolism and protein synthesis and diseases including diabetes, neurodegeneration, and cancer. IR chromatin binding was increased by insulin and impaired in an insulin-resistant disease model. Promoter binding by IR was mediated by coregulator host cell factor-1 (HCF-1) and transcription factors, revealing an HCF-1-dependent pathway for gene regulation by insulin. These results show that IR interacts with transcriptional machinery at promoters and identify a pathway regulating genes linked to insulin's effects in physiology and disease., (Copyright © 2019 Elsevier Inc. All rights reserved.)
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- 2019
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14. A lipid-free and insulin-supplemented medium supports De Novo fatty acid synthesis gene activation in melanoma cells.
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Wu S and Näär AM
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- Cell Line, Tumor, Cell Nucleus metabolism, Cell Proliferation drug effects, Cholesterol biosynthesis, Culture Media chemistry, Humans, Melanoma metabolism, Melanoma pathology, RNA Interference, RNA, Small Interfering metabolism, Selenium pharmacology, Sterol Regulatory Element Binding Protein 1 antagonists & inhibitors, Sterol Regulatory Element Binding Protein 1 metabolism, Sterol Regulatory Element Binding Protein 2 antagonists & inhibitors, Sterol Regulatory Element Binding Protein 2 metabolism, Transferrin pharmacology, Fatty Acids biosynthesis, Insulin pharmacology, Sterol Regulatory Element Binding Protein 1 genetics, Sterol Regulatory Element Binding Protein 2 genetics, Transcriptional Activation drug effects
- Abstract
While investigating the role played by de novo lipid (DNL) biosynthesis in cancer cells, we sought a medium condition that would support cell proliferation without providing any serum lipids. Here we report that a defined serum free cell culture medium condition containing insulin, transferrin and selenium (ITS) supports controlled study of transcriptional regulation of de novo fatty acid (DNFA) production and de novo cholesterol synthesis (DNCS) in melanoma cell lines. This lipid-free ITS medium is able to support continuous proliferation of several melanoma cell lines that utilize DNL to support their lipid requirements. We show that the ITS medium stimulates gene transcription in support of both DNFA and DNCS, specifically mediated by SREBP1/2 in melanoma cells. We further found that the ITS medium promoted SREBP1 nuclear localization and occupancy on DNFA gene promoters. Our data show clear utility of this serum and lipid-free medium for melanoma cancer cell culture and lipid-related areas of investigation., Competing Interests: The authors have declared that no competing interests exist.
- Published
- 2019
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15. Development of Locked Nucleic Acid Antisense Oligonucleotides Targeting Ebola Viral Proteins and Host Factor Niemann-Pick C1.
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Chery J, Petri A, Wagschal A, Lim SY, Cunningham J, Vasudevan S, Kauppinen S, and Näär AM
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- Animals, Disease Models, Animal, Ebolavirus genetics, Ebolavirus pathogenicity, Hemorrhagic Fever, Ebola therapy, Hemorrhagic Fever, Ebola virology, Humans, Immunity, Innate genetics, Mice, Niemann-Pick C1 Protein antagonists & inhibitors, Nucleoproteins antagonists & inhibitors, Nucleoproteins genetics, Oligonucleotides genetics, Oligonucleotides therapeutic use, Oligonucleotides, Antisense therapeutic use, Primates virology, Viral Proteins antagonists & inhibitors, Virus Replication genetics, Hemorrhagic Fever, Ebola genetics, Niemann-Pick C1 Protein genetics, Oligonucleotides, Antisense genetics, Viral Proteins genetics
- Abstract
The Ebola virus is a zoonotic pathogen that can cause severe hemorrhagic fever in humans, with up to 90% lethality. The deadly 2014 Ebola outbreak quickly made an unprecedented impact on human lives. While several vaccines and therapeutics are under development, current approaches contain several limitations, such as virus mutational escape, need for formulation or refrigeration, poor scalability, long lead-time, and high cost. To address these challenges, we developed locked nucleic acid (LNA)-modified antisense oligonucleotides (ASOs) to target critical Ebola viral proteins and the human intracellular host protein Niemann-Pick C1 (NPC1), required for viral entry into infected cells. We generated noninfectious viral luciferase reporter assays to identify LNA ASOs that inhibit translation of Ebola viral proteins in vitro and in human cells. We demonstrated specific inhibition of key Ebola genes VP24 and nucleoprotein, which inhibit a proper immune response and promote Ebola virus replication, respectively. We also identified LNA ASOs targeting human host factor NPC1 and demonstrated reduced infection by chimeric vesicular stomatitis virus harboring the Ebola glycoprotein, which directly binds to NPC1 for viral infection. These results support further in vivo testing of LNA ASOs in infectious Ebola virus disease animal models as potential therapeutic modalities for treatment of Ebola.
- Published
- 2018
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16. miR-33: A Metabolic Conundrum.
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Näär AM
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- Adiposity, Animals, Mice, Mice, Knockout, Obesity, Tissue Expansion, Insulin Resistance, MicroRNAs
- Abstract
The miR-33 microRNAs (miRNAs) are crucial regulators of cholesterol/lipids, and may represent therapeutic targets for the treatment of atherosclerosis. A recent report by Price et al. showed that miR-33 knockout (KO) mice exhibit obesity, insulin resistance, and increased food intake, suggesting that metabolic regulation by miR-33 is more complex than was previously known., (Copyright © 2018 Elsevier Ltd. All rights reserved.)
- Published
- 2018
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17. miRNA regulation of LDL-cholesterol metabolism.
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Goedeke L, Wagschal A, Fernández-Hernando C, and Näär AM
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- Animals, Cardiovascular Diseases genetics, Humans, Metabolic Diseases genetics, Receptors, LDL genetics, Triglycerides genetics, Cholesterol, LDL genetics, MicroRNAs genetics
- Abstract
In the past decade, microRNAs (miRNAs) have emerged as key regulators of circulating levels of lipoproteins. Specifically, recent work has uncovered the role of miRNAs in controlling the levels of atherogenic low-density lipoprotein LDL (LDL)-cholesterol by post-transcriptionally regulating genes involved in very low-density lipoprotein (VLDL) secretion, cholesterol biosynthesis, and hepatic LDL receptor (LDLR) expression. Interestingly, several of these miRNAs are located in genomic loci associated with abnormal levels of circulating lipids in humans. These findings reinforce the interest of targeting this subset of non-coding RNAs as potential therapeutic avenues for regulating plasma cholesterol and triglyceride (TAG) levels. In this review, we will discuss how these new miRNAs represent potential pre-disposition factors for cardiovascular disease (CVD), and putative therapeutic targets in patients with cardiometabolic disorders. This article is part of a Special Issue entitled: MicroRNAs and lipid/energy metabolism and related diseases edited by Carlos Fernández-Hernando and Yajaira Suárez., (Copyright © 2016 Elsevier B.V. All rights reserved.)
- Published
- 2016
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18. MicroRNA-33 Regulates the Innate Immune Response via ATP Binding Cassette Transporter-mediated Remodeling of Membrane Microdomains.
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Lai L, Azzam KM, Lin WC, Rai P, Lowe JM, Gabor KA, Madenspacher JH, Aloor JJ, Parks JS, Näär AM, and Fessler MB
- Subjects
- ATP Binding Cassette Transporter 1 genetics, ATP Binding Cassette Transporter, Subfamily G, Member 1 genetics, Animals, Membrane Microdomains genetics, Mice, Mice, Knockout, MicroRNAs genetics, RAW 264.7 Cells, Sterol Regulatory Element Binding Protein 2 genetics, Sterol Regulatory Element Binding Protein 2 immunology, ATP Binding Cassette Transporter 1 immunology, ATP Binding Cassette Transporter, Subfamily G, Member 1 immunology, Immunity, Innate, Macrophages immunology, Membrane Microdomains immunology, MicroRNAs immunology
- Abstract
MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene expression by promoting degradation and/or repressing translation of specific target mRNAs. Several miRNAs have been identified that regulate the amplitude of the innate immune response by directly targeting Toll-like receptor (TLR) pathway members and/or cytokines. miR-33a and miR-33b (the latter present in primates but absent in rodents and lower species) are located in introns of the sterol regulatory element-binding protein (SREBP)-encoding genes and control cholesterol/lipid homeostasis in concert with their host gene products. These miRNAs regulate macrophage cholesterol by targeting the lipid efflux transporters ATP binding cassette (ABC)A1 and ABCG1. We and others have previously reported that Abca1(-/-) and Abcg1(-/-) macrophages have increased TLR proinflammatory responses due to augmented lipid raft cholesterol. Given this, we hypothesized that miR-33 would augment TLR signaling in macrophages via a raft cholesterol-dependent mechanism. Herein, we report that multiple TLR ligands down-regulate miR-33 in murine macrophages. In the case of lipopolysaccharide, this is a delayed, Toll/interleukin-1 receptor (TIR) domain-containing adapter-inducing interferon-β-dependent response that also down-regulates Srebf-2, the host gene for miR-33. miR-33 augments macrophage lipid rafts and enhances proinflammatory cytokine induction and NF-κB activation by LPS. This occurs through an ABCA1- and ABCG1-dependent mechanism and is reversible by interventions upon raft cholesterol and by ABC transporter-inducing liver X receptor agonists. Taken together, these findings extend the purview of miR-33, identifying it as an indirect regulator of innate immunity that mediates bidirectional cross-talk between lipid homeostasis and inflammation., (© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.)
- Published
- 2016
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19. Inhibiting fungal multidrug resistance by disrupting an activator-Mediator interaction.
- Author
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Nishikawa JL, Boeszoermenyi A, Vale-Silva LA, Torelli R, Posteraro B, Sohn YJ, Ji F, Gelev V, Sanglard D, Sanguinetti M, Sadreyev RI, Mukherjee G, Bhyravabhotla J, Buhrlage SJ, Gray NS, Wagner G, Näär AM, and Arthanari H
- Subjects
- Animals, Binding Sites drug effects, Candida glabrata genetics, Candidiasis drug therapy, Candidiasis microbiology, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Drug Resistance, Multiple, Fungal drug effects, Fluconazole pharmacology, Gene Expression Regulation, Fungal drug effects, Hydrazines pharmacokinetics, Hydrazines pharmacology, Ketoconazole pharmacology, Mediator Complex chemistry, Mice, Models, Molecular, Nuclear Magnetic Resonance, Biomolecular, Protein Binding drug effects, Protein Structure, Tertiary, Saccharomyces cerevisiae Proteins chemistry, Saccharomyces cerevisiae Proteins genetics, Saccharomyces cerevisiae Proteins metabolism, Thiourea analogs & derivatives, Thiourea pharmacokinetics, Thiourea pharmacology, Trans-Activators chemistry, Transcription Factors genetics, Transcription Factors metabolism, Transcriptional Activation drug effects, Up-Regulation drug effects, Antifungal Agents pharmacology, Candida glabrata drug effects, Candida glabrata metabolism, Drug Resistance, Fungal drug effects, Fungal Proteins metabolism, Mediator Complex metabolism, Trans-Activators metabolism
- Abstract
Eukaryotic transcription activators stimulate the expression of specific sets of target genes through recruitment of co-activators such as the RNA polymerase II-interacting Mediator complex. Aberrant function of transcription activators has been implicated in several diseases. However, therapeutic targeting efforts have been hampered by a lack of detailed molecular knowledge of the mechanisms of gene activation by disease-associated transcription activators. We previously identified an activator-targeted three-helix bundle KIX domain in the human MED15 Mediator subunit that is structurally conserved in Gal11/Med15 Mediator subunits in fungi. The Gal11/Med15 KIX domain engages pleiotropic drug resistance transcription factor (Pdr1) orthologues, which are key regulators of the multidrug resistance pathway in Saccharomyces cerevisiae and in the clinically important human pathogen Candida glabrata. The prevalence of C. glabrata is rising, partly owing to its low intrinsic susceptibility to azoles, the most widely used antifungal agent. Drug-resistant clinical isolates of C. glabrata most commonly contain point mutations in Pdr1 that render it constitutively active, suggesting that this transcriptional activation pathway represents a linchpin in C. glabrata multidrug resistance. Here we perform sequential biochemical and in vivo high-throughput screens to identify small-molecule inhibitors of the interaction of the C. glabrata Pdr1 activation domain with the C. glabrata Gal11A KIX domain. The lead compound (iKIX1) inhibits Pdr1-dependent gene activation and re-sensitizes drug-resistant C. glabrata to azole antifungals in vitro and in animal models for disseminated and urinary tract C. glabrata infection. Determining the NMR structure of the C. glabrata Gal11A KIX domain provides a detailed understanding of the molecular mechanism of Pdr1 gene activation and multidrug resistance inhibition by iKIX1. We have demonstrated the feasibility of small-molecule targeting of a transcription factor-binding site in Mediator as a novel therapeutic strategy in fungal infectious disease.
- Published
- 2016
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20. Genome-wide identification of microRNAs regulating cholesterol and triglyceride homeostasis.
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Wagschal A, Najafi-Shoushtari SH, Wang L, Goedeke L, Sinha S, deLemos AS, Black JC, Ramírez CM, Li Y, Tewhey R, Hatoum I, Shah N, Lu Y, Kristo F, Psychogios N, Vrbanac V, Lu YC, Hla T, de Cabo R, Tsang JS, Schadt E, Sabeti PC, Kathiresan S, Cohen DE, Whetstine J, Chung RT, Fernández-Hernando C, Kaplan LM, Bernards A, Gerszten RE, and Näär AM
- Subjects
- Animals, Apolipoproteins E genetics, Cholesterol metabolism, Genome-Wide Association Study, Homeostasis genetics, Humans, Lipoproteins metabolism, Mice, Mice, Inbred C57BL, Mice, Knockout, Polymorphism, Single Nucleotide, ATP Binding Cassette Transporter 1 metabolism, Cholesterol, HDL metabolism, Cholesterol, LDL metabolism, Diet, High-Fat, Dyslipidemias genetics, MicroRNAs genetics, Receptors, LDL metabolism, Triglycerides metabolism
- Abstract
Genome-wide association studies (GWASs) have linked genes to various pathological traits. However, the potential contribution of regulatory noncoding RNAs, such as microRNAs (miRNAs), to a genetic predisposition to pathological conditions has remained unclear. We leveraged GWAS meta-analysis data from >188,000 individuals to identify 69 miRNAs in physical proximity to single-nucleotide polymorphisms (SNPs) associated with abnormal levels of circulating lipids. Several of these miRNAs (miR-128-1, miR-148a, miR-130b, and miR-301b) control the expression of key proteins involved in cholesterol-lipoprotein trafficking, such as the low-density lipoprotein (LDL) receptor (LDLR) and the ATP-binding cassette A1 (ABCA1) cholesterol transporter. Consistent with human liver expression data and genetic links to abnormal blood lipid levels, overexpression and antisense targeting of miR-128-1 or miR-148a in high-fat diet-fed C57BL/6J and Apoe-null mice resulted in altered hepatic expression of proteins involved in lipid trafficking and metabolism, and in modulated levels of circulating lipoprotein-cholesterol and triglycerides. Taken together, these findings support the notion that altered expression of miRNAs may contribute to abnormal blood lipid levels, predisposing individuals to human cardiometabolic disorders.
- Published
- 2015
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21. MicroRNA-148a regulates LDL receptor and ABCA1 expression to control circulating lipoprotein levels.
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Goedeke L, Rotllan N, Canfrán-Duque A, Aranda JF, Ramírez CM, Araldi E, Lin CS, Anderson NN, Wagschal A, de Cabo R, Horton JD, Lasunción MA, Näär AM, Suárez Y, and Fernández-Hernando C
- Subjects
- ATP Binding Cassette Transporter 1 metabolism, Animals, Gene Expression Regulation, Hep G2 Cells, High-Throughput Screening Assays, Humans, Mice, MicroRNAs metabolism, RNA Processing, Post-Transcriptional, Receptors, LDL metabolism, Signal Transduction, Sterol Regulatory Element Binding Protein 1 genetics, Sterol Regulatory Element Binding Protein 1 metabolism, ATP Binding Cassette Transporter 1 genetics, Cholesterol, HDL metabolism, Cholesterol, LDL metabolism, Hepatocytes metabolism, Liver metabolism, MicroRNAs genetics, Receptors, LDL genetics
- Abstract
The hepatic low-density lipoprotein receptor (LDLR) pathway is essential for clearing circulating LDL cholesterol (LDL-C). Whereas the transcriptional regulation of LDLR is well characterized, the post-transcriptional mechanisms that govern LDLR expression are just beginning to emerge. Here we develop a high-throughput genome-wide screening assay to systematically identify microRNAs (miRNAs) that regulate LDLR activity in human hepatic cells. From this screen we identified and characterized miR-148a as a negative regulator of LDLR expression and activity and defined a sterol regulatory element-binding protein 1 (SREBP1)-mediated pathway through which miR-148a regulates LDL-C uptake. In mice, inhibition of miR-148a increased hepatic LDLR expression and decreased plasma LDL-C. Moreover, we found that miR-148a regulates hepatic expression of ATP-binding cassette, subfamily A, member 1 (ABCA1) and circulating high-density lipoprotein cholesterol (HDL-C) levels in vivo. These studies uncover a role for miR-148a as a key regulator of hepatic LDL-C clearance through direct modulation of LDLR expression and demonstrate the therapeutic potential of inhibiting miR-148a to ameliorate an elevated LDL-C/HDL-C ratio, a prominent risk factor for cardiovascular disease.
- Published
- 2015
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22. Haploinsufficiency for BRCA1 leads to cell-type-specific genomic instability and premature senescence.
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Sedic M, Skibinski A, Brown N, Gallardo M, Mulligan P, Martinez P, Keller PJ, Glover E, Richardson AL, Cowan J, Toland AE, Ravichandran K, Riethman H, Naber SP, Näär AM, Blasco MA, Hinds PW, and Kuperwasser C
- Subjects
- DNA Damage, Epithelial Cells cytology, Heterozygote, Humans, Mammary Glands, Human cytology, Mutation, Retinoblastoma Protein genetics, Retinoblastoma Protein metabolism, Sirtuin 1 genetics, Sirtuin 1 metabolism, Tumor Suppressor Protein p53 genetics, Tumor Suppressor Protein p53 metabolism, Cellular Senescence genetics, Epithelial Cells metabolism, Genes, BRCA1, Genomic Instability genetics, Haploinsufficiency, Mammary Glands, Human metabolism, Telomere Shortening genetics
- Abstract
Although BRCA1 function is essential for maintaining genomic integrity in all cell types, it is unclear why increased risk of cancer in individuals harbouring deleterious mutations in BRCA1 is restricted to only a select few tissues. Here we show that human mammary epithelial cells (HMECs) from BRCA1-mutation carriers (BRCA1(mut/+)) exhibit increased genomic instability and rapid telomere erosion in the absence of tumour-suppressor loss. Furthermore, we uncover a novel form of haploinsufficiency-induced senescence (HIS) specific to epithelial cells, which is triggered by pRb pathway activation rather than p53 induction. HIS and telomere erosion in HMECs correlate with misregulation of SIRT1 leading to increased levels of acetylated pRb as well as acetylated H4K16 both globally and at telomeric regions. These results identify a novel form of cellular senescence and provide a potential molecular basis for the rapid cell- and tissue- specific predisposition of breast cancer development associated with BRCA1 haploinsufficiency.
- Published
- 2015
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23. A role for FACT in repopulation of nucleosomes at inducible genes.
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Voth WP, Takahata S, Nishikawa JL, Metcalfe BM, Näär AM, and Stillman DJ
- Subjects
- Antifungal Agents pharmacology, Chromatin Assembly and Disassembly, Drug Resistance, Fungal genetics, Fungal Proteins metabolism, Gene Expression Regulation, Fungal drug effects, Histones metabolism, Mutation, Protein Binding, Transcription Factors metabolism, Transcription, Genetic, Fungal Proteins genetics, Nucleosomes genetics, Nucleosomes metabolism, Transcription Factors genetics
- Abstract
Xenobiotic drugs induce Pleiotropic Drug Resistance (PDR) genes via the orthologous Pdr1/Pdr3 transcription activators. We previously identified the Mediator transcription co-activator complex as a key target of Pdr1 orthologs and demonstrated that Pdr1 interacts directly with the Gal11/Med15 subunit of the Mediator complex. Based on an interaction between Pdr1 and the FACT complex, we show that strains with spt16 or pob3 mutations are sensitive to xenobiotic drugs and display diminished PDR gene induction. Although FACT acts during the activation of some genes by assisting in the nucleosomes eviction at promoters, PDR promoters already contain nucleosome-depleted regions (NDRs) before induction. To determine the function of FACT at PDR genes, we examined the kinetics of RNA accumulation and changes in nucleosome occupancy following exposure to a xenobiotic drug in wild type and FACT mutant yeast strains. In the presence of normal FACT, PDR genes are transcribed within 5 minutes of xenobiotic stimulation and transcription returns to basal levels by 30-40 min. Nucleosomes are constitutively depleted in the promoter regions, are lost from the open reading frames during transcription, and the ORFs are wholly repopulated with nucleosomes as transcription ceases. While FACT mutations cause minor delays in activation of PDR genes, much more pronounced and significant defects in nucleosome repopulation in the ORFs are observed in FACT mutants upon transcription termination. FACT therefore has a major role in nucleosome redeposition following cessation of transcription at the PDR genes, the opposite of its better-known function in nucleosome disassembly.
- Published
- 2014
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24. Pharmacological inhibition of a microRNA family in nonhuman primates by a seed-targeting 8-mer antimiR.
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Rottiers V, Obad S, Petri A, McGarrah R, Lindholm MW, Black JC, Sinha S, Goody RJ, Lawrence MS, deLemos AS, Hansen HF, Whittaker S, Henry S, Brookes R, Najafi-Shoushtari SH, Chung RT, Whetstine JR, Gerszten RE, Kauppinen S, and Näär AM
- Subjects
- Animals, Cholesterol, HDL blood, Female, Hep G2 Cells, Humans, Mice, Mice, Inbred C57BL, MicroRNAs genetics, Primates, Gene Silencing, MicroRNAs antagonists & inhibitors
- Abstract
MicroRNAs (miRNAs) regulate many aspects of human biology. They target mRNAs for translational repression or degradation through base pairing with 3' untranslated regions, primarily via seed sequences (nucleotides 2 to 8 in the mature miRNA sequence). A number of individual miRNAs and miRNA families share seed sequences and targets, but differ in the sequences outside of the seed. miRNAs have been implicated in the etiology of a wide variety of human diseases and therefore represent promising therapeutic targets. However, potential redundancy of different miRNAs sharing the same seed sequence and the challenge of simultaneously targeting miRNAs that differ significantly in nonseed sequences complicate therapeutic targeting approaches. We recently demonstrated effective inhibition of entire miRNA families using seed-targeting 8-mer locked nucleic acid (LNA)-modified antimiRs in short-term experiments in mammalian cells and in mice. However, the long-term efficacy and safety of this approach in higher organisms, such as humans and nonhuman primates, have not been determined. We show that pharmacological inhibition of the miR-33 family, key regulators of cholesterol/lipid homeostasis, by a subcutaneously delivered 8-mer LNA-modified antimiR in obese and insulin-resistant nonhuman primates results in derepression of miR-33 targets, such as ABCA1, increases circulating high-density lipoprotein cholesterol, and is well tolerated over 108 days of treatment. These findings demonstrate the efficacy and safety of an 8-mer LNA-antimiR against an miRNA family in a nonhuman primate metabolic disease model, suggesting that this could be a feasible approach for therapeutic targeting of miRNA families sharing the same seed sequence in human diseases.
- Published
- 2013
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25. SIRT6 recruits SNF2H to DNA break sites, preventing genomic instability through chromatin remodeling.
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Toiber D, Erdel F, Bouazoune K, Silberman DM, Zhong L, Mulligan P, Sebastian C, Cosentino C, Martinez-Pastor B, Giacosa S, D'Urso A, Näär AM, Kingston R, Rippe K, and Mostoslavsky R
- Subjects
- Adenosine Triphosphatases genetics, Animals, Cells, Cultured, Cerebral Cortex cytology, Cerebral Cortex metabolism, Chromatin Immunoprecipitation, Chromosomal Proteins, Non-Histone genetics, Hippocampus cytology, Hippocampus metabolism, Histones metabolism, Humans, Immunoprecipitation, Mice, Mice, Knockout, Nucleosomes metabolism, Sirtuins genetics, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, Adenosine Triphosphatases metabolism, Chromatin genetics, Chromatin Assembly and Disassembly, Chromosomal Proteins, Non-Histone metabolism, DNA Damage genetics, DNA Repair genetics, Genomic Instability, Sirtuins metabolism, Sirtuins physiology
- Abstract
DNA damage is linked to multiple human diseases, such as cancer, neurodegeneration, and aging. Little is known about the role of chromatin accessibility in DNA repair. Here, we find that the deacetylase sirtuin 6 (SIRT6) is one of the earliest factors recruited to double-strand breaks (DSBs). SIRT6 recruits the chromatin remodeler SNF2H to DSBs and focally deacetylates histone H3K56. Lack of SIRT6 and SNF2H impairs chromatin remodeling, increasing sensitivity to genotoxic damage and recruitment of downstream factors such as 53BP1 and breast cancer 1 (BRCA1). Remarkably, SIRT6-deficient mice exhibit lower levels of chromatin-associated SNF2H in specific tissues, a phenotype accompanied by DNA damage. We demonstrate that SIRT6 is critical for recruitment of a chromatin remodeler as an early step in the DNA damage response, indicating that proper unfolding of chromatin plays a rate-limiting role. We present a unique crosstalk between a histone modifier and a chromatin remodeler, regulating a coordinated response to prevent DNA damage., (Copyright © 2013 Elsevier Inc. All rights reserved.)
- Published
- 2013
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26. Anti-atherosclerosis or No Anti-atherosclerosis: That is the miR-33 question.
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Näär AM
- Subjects
- Animals, Male, Aorta metabolism, Aortic Diseases therapy, Atherosclerosis therapy, Liver metabolism, MicroRNAs metabolism, Oligonucleotides, Antisense metabolism, Receptors, LDL deficiency
- Published
- 2013
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27. MicroRNAs in metabolism and metabolic disorders.
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Rottiers V and Näär AM
- Subjects
- Cholesterol metabolism, Endocrine System metabolism, Fatty Liver genetics, Fatty Liver metabolism, Glucose metabolism, Homeostasis, Humans, Insulin metabolism, Lipid Metabolism, Metabolic Syndrome genetics, Metabolic Syndrome metabolism, MicroRNAs genetics, Non-alcoholic Fatty Liver Disease, Obesity genetics, Obesity metabolism, Oligoribonucleotides, Antisense pharmacology, Signal Transduction, Sterol Regulatory Element Binding Proteins genetics, Sterol Regulatory Element Binding Proteins metabolism, Metabolic Diseases genetics, Metabolic Diseases metabolism, MicroRNAs metabolism
- Abstract
MicroRNAs (miRNAs) have recently emerged as key regulators of metabolism. For example, miR-33a and miR-33b have a crucial role in controlling cholesterol and lipid metabolism in concert with their host genes, the sterol-regulatory element-binding protein (SREBP) transcription factors. Other metabolic miRNAs, such as miR-103 and miR-107, regulate insulin and glucose homeostasis, whereas miRNAs such as miR-34a are emerging as key regulators of hepatic lipid homeostasis. The discovery of circulating miRNAs has highlighted their potential as both endocrine signalling molecules and disease markers. Dysregulation of miRNAs may contribute to metabolic abnormalities, suggesting that miRNAs may potentially serve as therapeutic targets for ameliorating cardiometabolic disorders.
- Published
- 2012
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28. A conserved SREBP-1/phosphatidylcholine feedback circuit regulates lipogenesis in metazoans.
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Walker AK, Jacobs RL, Watts JL, Rottiers V, Jiang K, Finnegan DM, Shioda T, Hansen M, Yang F, Niebergall LJ, Vance DE, Tzoneva M, Hart AC, and Näär AM
- Subjects
- Animals, Cell Line, Tumor, Endoplasmic Reticulum metabolism, Humans, Lipogenesis, Mice, Models, Animal, Phosphatidylcholines biosynthesis, RNA Interference, S-Adenosylmethionine biosynthesis, Caenorhabditis elegans metabolism, Caenorhabditis elegans Proteins metabolism, Sterol Regulatory Element Binding Protein 1 metabolism, Transcription Factors metabolism
- Abstract
Sterol regulatory element-binding proteins (SREBPs) activate genes involved in the synthesis and trafficking of cholesterol and other lipids and are critical for maintaining lipid homeostasis. Aberrant SREBP activity, however, can contribute to obesity, fatty liver disease, and insulin resistance, hallmarks of metabolic syndrome. Our studies identify a conserved regulatory circuit in which SREBP-1 controls genes in the one-carbon cycle, which produces the methyl donor S-adenosylmethionine (SAMe). Methylation is critical for the synthesis of phosphatidylcholine (PC), a major membrane component, and we find that blocking SAMe or PC synthesis in C. elegans, mouse liver, and human cells causes elevated SREBP-1-dependent transcription and lipid droplet accumulation. Distinct from negative regulation of SREBP-2 by cholesterol, our data suggest a feedback mechanism whereby maturation of nuclear, transcriptionally active SREBP-1 is controlled by levels of PC. Thus, nutritional or genetic conditions limiting SAMe or PC production may activate SREBP-1, contributing to human metabolic disorders., (Copyright © 2011 Elsevier Inc. All rights reserved.)
- Published
- 2011
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29. MiRs with a sweet tooth.
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Näär AM
- Abstract
MicroRNAs (miRNAs) have recently been found to be critical regulators of metabolic homeostasis. A study in Nature by Trajkovski et al. (2011) shows that the highly related miRNAs miR-103 and miR-107 modulate insulin sensitivity and glucose homeostasis in obese mice. These miRNAs might represent therapeutic targets to ameliorate obesity-associated insulin resistance., (Copyright © 2011 Elsevier Inc. All rights reserved.)
- Published
- 2011
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30. A SIRT1-LSD1 corepressor complex regulates Notch target gene expression and development.
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Mulligan P, Yang F, Di Stefano L, Ji JY, Ouyang J, Nishikawa JL, Toiber D, Kulkarni M, Wang Q, Najafi-Shoushtari SH, Mostoslavsky R, Gygi SP, Gill G, Dyson NJ, and Näär AM
- Subjects
- Animals, Drosophila Proteins genetics, Drosophila Proteins metabolism, Drosophila melanogaster growth & development, Epigenesis, Genetic, Gene Expression Regulation, Developmental, Histones metabolism, Immunoprecipitation, Mutation, Oxidoreductases, N-Demethylating genetics, Oxidoreductases, N-Demethylating metabolism, Phenotype, Receptors, Notch metabolism, Sirtuin 1 genetics, Sirtuin 1 metabolism, Drosophila Proteins physiology, Drosophila melanogaster genetics, Oxidoreductases, N-Demethylating physiology, Receptors, Notch genetics, Sirtuin 1 physiology
- Abstract
Epigenetic regulation of gene expression by histone-modifying corepressor complexes is central to normal animal development. The NAD(+)-dependent deacetylase and gene repressor SIRT1 removes histone H4K16 acetylation marks and facilitates heterochromatin formation. However, the mechanistic contribution of SIRT1 to epigenetic regulation at euchromatic loci and whether it acts in concert with other chromatin-modifying activities to control developmental gene expression programs remain unclear. We describe here a SIRT1 corepressor complex containing the histone H3K4 demethylase LSD1/KDM1A and several other LSD1-associated proteins. SIRT1 and LSD1 interact directly and play conserved and concerted roles in H4K16 deacetylation and H3K4 demethylation to repress genes regulated by the Notch signaling pathway. Mutations in Drosophila SIRT1 and LSD1 orthologs result in similar developmental phenotypes and genetically interact with the Notch pathway in Drosophila. These findings offer new insights into conserved mechanisms of epigenetic gene repression and regulation of development by SIRT1 in metazoans., (Copyright © 2011 Elsevier Inc. All rights reserved.)
- Published
- 2011
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31. Structure of the VP16 transactivator target in the Mediator.
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Milbradt AG, Kulkarni M, Yi T, Takeuchi K, Sun ZY, Luna RE, Selenko P, Näär AM, and Wagner G
- Subjects
- Herpes Simplex Virus Protein Vmw65 genetics, Humans, Mediator Complex genetics, Mutation, Nuclear Magnetic Resonance, Biomolecular, Point Mutation, Protein Conformation, Herpes Simplex Virus Protein Vmw65 chemistry, Mediator Complex chemistry
- Abstract
The human Mediator coactivator complex interacts with many transcriptional activators and facilitates recruitment of RNA polymerase II to promote target gene transcription. The MED25 subunit is a critical target of the potent herpes simplex 1 viral transcriptional activator VP16. Here we determine the solution structure of the MED25 VP16-binding domain (VBD) and define its binding site for the N-terminal portion of the VP16 transactivation domain (TADn). A hydrophobic furrow, formed by a β-barrel and two α-helices in MED25 VBD, interacts tightly with VP16 TADn. Mutations in this furrow prevent binding of VP16 TAD to MED25 VBD and interfere with the ability of overexpressed MED25 VBD to inhibit VP16-dependent transcriptional activation in vivo. This detailed molecular understanding of transactivation by the benchmark activator VP16 could provide important insights into viral and cellular gene activation mechanisms.
- Published
- 2011
- Full Text
- View/download PDF
32. MicroRNAs in metabolism and metabolic diseases.
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Rottiers V, Najafi-Shoushtari SH, Kristo F, Gurumurthy S, Zhong L, Li Y, Cohen DE, Gerszten RE, Bardeesy N, Mostoslavsky R, and Näär AM
- Subjects
- AMP-Activated Protein Kinases metabolism, ATP Binding Cassette Transporter 1, ATP-Binding Cassette Transporters metabolism, Animals, Base Sequence, Biological Transport genetics, Cholesterol metabolism, Conserved Sequence genetics, Energy Metabolism genetics, Fatty Acids metabolism, Glucose metabolism, Homeostasis genetics, Humans, Insulin Receptor Substrate Proteins metabolism, Introns genetics, Mice, MicroRNAs biosynthesis, MicroRNAs genetics, Models, Biological, Molecular Sequence Data, Oxidation-Reduction, Sirtuins metabolism, Sterol Regulatory Element Binding Proteins genetics, Sterol Regulatory Element Binding Proteins metabolism, Metabolic Diseases genetics, Metabolism genetics, MicroRNAs metabolism
- Abstract
Aberrant cholesterol/lipid homeostasis is linked to a number of diseases prevalent in the developed world, including metabolic syndrome, type II diabetes, and cardiovascular disease. We have previously uncovered gene regulatory mechanisms of the sterol regulatory element-binding protein (SREBP) family of transcription factors, which control the expression of genes involved in cholesterol and lipid biosynthesis and uptake. Intriguingly, we recently discovered conserved microRNAs (miR-33a/b) embedded within intronic sequences of the human SREBF genes that act in a concerted manner with their host gene products to regulate cholesterol/lipid homeostasis. Indeed, miR-33a/b control the levels of ATP-binding cassette (ABC) transporter ABCA1, a cholesterol efflux pump critical for high-density lipoprotein (HDL) synthesis and reverse cholesterol transport from peripheral tissues. Importantly, antisense inhibition of miR-33 in mice results in elevated HDL and decreased atherosclerosis. Interestingly, miR-33a/b also act in the fatty acid/lipid homeostasis pathway by controlling the fatty acid β-oxidation genes carnitine O-octanoyltransferase (CROT), hydroxyacyl-coenzyme A-dehydrogenase (HADHB), and carnitine palmitoyltransferase 1A (CPT1A), as well as the energy sensor AMP-activated protein kinase (AMPKα1), the NAD(+)-dependent sirtuin SIRT6, and the insulin signaling intermediate IRS2, key regulators of glucose and lipid metabolism. These results have revealed a highly integrated microRNA (miRNA)-host gene circuit governing cholesterol/lipid metabolism and energy homeostasis in mammals that may have important therapeutic implications for the treatment of cardiometabolic disorders.
- Published
- 2011
- Full Text
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33. Functional antagonism between histone H3K4 demethylases in vivo.
- Author
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Di Stefano L, Walker JA, Burgio G, Corona DF, Mulligan P, Näär AM, and Dyson NJ
- Subjects
- Animals, Drosophila Proteins genetics, Drosophila melanogaster genetics, Gene Expression Regulation, Heterochromatin metabolism, Histone-Lysine N-Methyltransferase genetics, Histones metabolism, Methylation, Mutation genetics, Oxidoreductases, N-Demethylating genetics, Phenotype, Receptors, Notch genetics, Signal Transduction, Drosophila Proteins metabolism, Drosophila melanogaster enzymology, Histone Demethylases metabolism, Histone-Lysine N-Methyltransferase metabolism, Oxidoreductases, N-Demethylating metabolism
- Abstract
Dynamic regulation of histone modifications is critical during development, and aberrant activity of chromatin-modifying enzymes has been associated with diseases such as cancer. Histone demethylases have been shown to play a key role in eukaryotic gene transcription; however, little is known about how their activities are coordinated in vivo to regulate specific biological processes. In Drosophila, two enzymes, dLsd1 (Drosophila ortholog of lysine-specific demethylase 1) and Lid (little imaginal discs), demethylate histone H3 at Lys 4 (H3K4), a residue whose methylation is associated with actively transcribed genes. Our studies show that compound mutation of Lid and dLsd1 results in increased H3K4 methylation levels. However, unexpectedly, Lid mutations strongly suppress dLsd1 mutant phenotypes. Investigation of the basis for this antagonism revealed that Lid opposes the functions of dLsd1 and the histone methyltransferase Su(var)3-9 in promoting heterochromatin spreading at heterochromatin-euchromatin boundaries. Moreover, our data reveal a novel role for dLsd1 in Notch signaling in Drosophila, and a complex network of interactions between dLsd1, Lid, and Notch signaling at euchromatic genes. These findings illustrate the complexity of functional interplay between histone demethylases in vivo, providing insights into the epigenetic regulation of heterochromatin/euchromatin boundaries by Lid and dLsd1 and showing their involvement in Notch pathway-specific control of gene expression in euchromatin.
- Published
- 2011
- Full Text
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34. Differential roles of transcriptional mediator subunits in regulation of multidrug resistance gene expression in Saccharomyces cerevisiae.
- Author
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Shahi P, Gulshan K, Näär AM, and Moye-Rowley WS
- Subjects
- Mediator Complex genetics, Mitochondria metabolism, Mutation genetics, Phenotype, Promoter Regions, Genetic genetics, Protein Binding, Protein Subunits genetics, Saccharomyces cerevisiae cytology, Saccharomyces cerevisiae Proteins metabolism, Signal Transduction genetics, Transcriptional Activation genetics, Drug Resistance, Multiple, Fungal genetics, Gene Expression Regulation, Fungal, Mediator Complex metabolism, Protein Subunits metabolism, Saccharomyces cerevisiae genetics
- Abstract
The multiprotein transcriptional Mediator complex provides a key link between RNA polymerase II and upstream transcriptional activator proteins. Previous work has established that the multidrug resistance transcription factors Pdr1 and Pdr3 interact with the Mediator component Med15/Gal11 to drive normal levels of expression of the ATP-binding cassette transporter-encoding gene PDR5 in Saccharomyces cerevisiae. PDR5 transcription is induced upon loss of the mitochondrial genome (rho(0) cells) and here we provide evidence that this rho(0) induction is Med15 independent. A search through other known Mediator components determined that Med12/Srb8, a member of the CDK8 Mediator submodule, is required for rho(0) activation of PDR5 transcription. The CDK8 submodule contains the cyclin C homologue (CycC/Srb11), cyclin-dependent kinase Cdk8/Srb10, and the large Med13/Srb9 protein. Loss of these other proteins did not lead to the same block in PDR5 induction. Chromatin immunoprecipitation analyses demonstrated that Med15 is associated with the PDR5 promoter in both rho(+) and rho(0), whereas Med12 recruitment to this target promoter is highly responsive to loss of the mitochondrial genome. Coimmunoprecipitation experiments revealed that association of Pdr3 with Med12 can only be detected in rho(0) cells. These experiments uncover the unique importance of Med12 in activated transcription of PDR5 seen in rho(0) cells.
- Published
- 2010
- Full Text
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35. Conserved role of SIRT1 orthologs in fasting-dependent inhibition of the lipid/cholesterol regulator SREBP.
- Author
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Walker AK, Yang F, Jiang K, Ji JY, Watts JL, Purushotham A, Boss O, Hirsch ML, Ribich S, Smith JJ, Israelian K, Westphal CH, Rodgers JT, Shioda T, Elson SL, Mulligan P, Najafi-Shoushtari H, Black JC, Thakur JK, Kadyk LC, Whetstine JR, Mostoslavsky R, Puigserver P, Li X, Dyson NJ, Hart AC, and Näär AM
- Subjects
- Acetylation, Animals, Benzamides pharmacology, Caenorhabditis elegans, Cell Line, Cholesterol biosynthesis, HeLa Cells, Heterocyclic Compounds, 4 or More Rings pharmacology, Humans, Lipids biosynthesis, Mice, Naphthols pharmacology, Niacinamide pharmacology, Protein Stability drug effects, Sirtuins antagonists & inhibitors, Down-Regulation drug effects, Fasting physiology, Sirtuin 1 metabolism, Sterol Regulatory Element Binding Protein 1 metabolism, Sterol Regulatory Element Binding Protein 2 metabolism
- Abstract
The sterol regulatory element-binding protein (SREBP) transcription factor family is a critical regulator of lipid and sterol homeostasis in eukaryotes. In mammals, SREBPs are highly active in the fed state to promote the expression of lipogenic and cholesterogenic genes and facilitate fat storage. During fasting, SREBP-dependent lipid/cholesterol synthesis is rapidly diminished in the mouse liver; however, the mechanism has remained incompletely understood. Moreover, the evolutionary conservation of fasting regulation of SREBP-dependent programs of gene expression and control of lipid homeostasis has been unclear. We demonstrate here a conserved role for orthologs of the NAD(+)-dependent deacetylase SIRT1 in metazoans in down-regulation of SREBP orthologs during fasting, resulting in inhibition of lipid synthesis and fat storage. Our data reveal that SIRT1 can directly deacetylate SREBP, and modulation of SIRT1 activity results in changes in SREBP ubiquitination, protein stability, and target gene expression. In addition, chemical activators of SIRT1 inhibit SREBP target gene expression in vitro and in vivo, correlating with decreased hepatic lipid and cholesterol levels and attenuated liver steatosis in diet-induced and genetically obese mice. We conclude that SIRT1 orthologs play a critical role in controlling SREBP-dependent gene regulation governing lipid/cholesterol homeostasis in metazoans in response to fasting cues. These findings may have important biomedical implications for the treatment of metabolic disorders associated with aberrant lipid/cholesterol homeostasis, including metabolic syndrome and atherosclerosis.
- Published
- 2010
- Full Text
- View/download PDF
36. MicroRNA-33 and the SREBP host genes cooperate to control cholesterol homeostasis.
- Author
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Najafi-Shoushtari SH, Kristo F, Li Y, Shioda T, Cohen DE, Gerszten RE, and Näär AM
- Subjects
- 3' Untranslated Regions, ATP Binding Cassette Transporter 1, ATP-Binding Cassette Transporters genetics, Animals, Cell Line, Diet, Gene Expression Regulation, Homeostasis, Humans, Introns, Liver metabolism, Macrophages metabolism, Mice, Mice, Inbred C57BL, MicroRNAs genetics, Oligonucleotides, Antisense pharmacology, RNA Interference, Sterol Regulatory Element Binding Protein 1 genetics, Sterol Regulatory Element Binding Protein 1 metabolism, Sterol Regulatory Element Binding Protein 2 genetics, Sterol Regulatory Element Binding Protein 2 metabolism, Sterol Regulatory Element Binding Proteins metabolism, Up-Regulation, ATP-Binding Cassette Transporters metabolism, Cholesterol metabolism, Cholesterol, HDL blood, MicroRNAs metabolism, Sterol Regulatory Element Binding Proteins genetics
- Abstract
Proper coordination of cholesterol biosynthesis and trafficking is essential to human health. The sterol regulatory element-binding proteins (SREBPs) are key transcription regulators of genes involved in cholesterol biosynthesis and uptake. We show here that microRNAs (miR-33a/b) embedded within introns of the SREBP genes target the adenosine triphosphate-binding cassette transporter A1 (ABCA1), an important regulator of high-density lipoprotein (HDL) synthesis and reverse cholesterol transport, for posttranscriptional repression. Antisense inhibition of miR-33 in mouse and human cell lines causes up-regulation of ABCA1 expression and increased cholesterol efflux, and injection of mice on a western-type diet with locked nucleic acid-antisense oligonucleotides results in elevated plasma HDL. Our findings indicate that miR-33 acts in concert with the SREBP host genes to control cholesterol homeostasis and suggest that miR-33 may represent a therapeutic target for ameliorating cardiometabolic diseases.
- Published
- 2010
- Full Text
- View/download PDF
37. Nuclear receptor-like transcription factors in fungi.
- Author
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Näär AM and Thakur JK
- Subjects
- Animals, DNA-Binding Proteins metabolism, Drug Resistance, Multiple, Fungal physiology, Fatty Acids metabolism, Humans, Oxidation-Reduction, Peroxisomes physiology, Transcriptional Activation physiology, Fungi genetics, Fungi metabolism, Receptors, Cytoplasmic and Nuclear metabolism, Transcription Factors metabolism
- Abstract
Members of the metazoan nuclear receptor superfamily regulate gene expression programs in response to binding of cognate lipophilic ligands. Evolutionary studies using bioinformatics tools have concluded that lower eukaryotes, such as fungi, lack nuclear receptor homologs. Here we review recent discoveries suggesting that members of the fungal zinc cluster family of transcription regulators represent functional analogs of metazoan nuclear receptors. These findings indicate that nuclear receptor-like ligand-dependent gene regulatory mechanisms emerged early during eukaryotic evolution, and provide the impetus for further detailed studies of the possible evolutionary and mechanistic relationships of fungal zinc cluster transcription factors and metazoan nuclear receptors. Clinical implications of the discovery of nuclear receptor-like transcription factors in pathogenic fungi will also be discussed.
- Published
- 2009
- Full Text
- View/download PDF
38. Mediator subunit Gal11p/MED15 is required for fatty acid-dependent gene activation by yeast transcription factor Oaf1p.
- Author
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Thakur JK, Arthanari H, Yang F, Chau KH, Wagner G, and Näär AM
- Subjects
- DNA-Binding Proteins genetics, Fatty Acids genetics, Mediator Complex, Nuclear Magnetic Resonance, Biomolecular, Protein Structure, Tertiary physiology, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins genetics, Trans-Activators genetics, Transcription Factors genetics, DNA-Binding Proteins metabolism, Fatty Acids metabolism, Lipid Metabolism physiology, Saccharomyces cerevisiae metabolism, Saccharomyces cerevisiae Proteins metabolism, Trans-Activators metabolism, Transcription Factors metabolism, Transcription, Genetic physiology
- Abstract
The yeast zinc cluster transcription factor Oaf1p activates transcription of target genes in response to direct binding of fatty acids in a manner analogous to the vertebrate nuclear receptor peroxisome proliferator-activated receptoralpha (PPARalpha). PPARs and other metazoan nuclear receptors productively engage several distinct LXXLL motif-containing co-activators, including p160 family members and the TRAP220/MED1 subunit of the Mediator co-activator, to promote ligand-dependent gene activation. Yeast, however, does not appear to harbor LXXLL motif co-activators, and the mechanism of fatty acid-dependent gene activation by the yeast PPARalpha analog Oaf1p is unknown. Here we show that the yeast Mediator subunit Gal11p/MED15 and its activator-targeted KIX domain plays a critical role in fatty acid-dependent transcriptional regulation of fatty acid beta-oxidation and peroxisomal genes by Oaf1p and for the ability of yeast to utilize fatty acids as a sole carbon source. Moreover, structural studies by NMR spectroscopy reveal that the Oaf1p activation domain interacts with the Gal11p/MED15 KIX domain in a manner similar to the yeast zinc cluster family member and xenobiotic receptor Pdr1p, revealing that the Gal11p/MED15 KIX domain is a key target of several ligand-dependent transcription factors in yeast. Together with previous work showing that the Caenorhabditis elegans Gal11p/MED15 homolog MDT-15 plays a critical role in regulation of fatty acid metabolism by the nematode PPAR-like nuclear receptor NHR-49, the findings presented here provide evidence for an ancient and essential role of a Mediator co-activator subunit in regulation of fatty acid metabolism by nuclear receptor-like transcription factors in eukaryotes.
- Published
- 2009
- Full Text
- View/download PDF
39. E2F1 represses beta-catenin transcription and is antagonized by both pRB and CDK8.
- Author
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Morris EJ, Ji JY, Yang F, Di Stefano L, Herr A, Moon NS, Kwon EJ, Haigis KM, Näär AM, and Dyson NJ
- Subjects
- Adenomatous Polyposis Coli Protein metabolism, Apoptosis, Cell Line, Cyclin-Dependent Kinase 8, Gene Expression Regulation, Genes, myc genetics, Glycogen Synthase Kinase 3 metabolism, Humans, Retinoblastoma Protein genetics, Signal Transduction, TCF Transcription Factors metabolism, Wnt Proteins metabolism, Cyclin-Dependent Kinases metabolism, E2F1 Transcription Factor antagonists & inhibitors, E2F1 Transcription Factor metabolism, Retinoblastoma Protein metabolism, Transcription, Genetic, beta Catenin antagonists & inhibitors, beta Catenin metabolism
- Abstract
The E2F1 transcription factor can promote proliferation or apoptosis when activated, and is a key downstream target of the retinoblastoma tumour suppressor protein (pRB). Here we show that E2F1 is a potent and specific inhibitor of beta-catenin/T-cell factor (TCF)-dependent transcription, and that this function contributes to E2F1-induced apoptosis. E2F1 deregulation suppresses beta-catenin activity in an adenomatous polyposis coli (APC)/glycogen synthase kinase-3 (GSK3)-independent manner, reducing the expression of key beta-catenin targets including c-MYC. This interaction explains why colorectal tumours, which depend on beta-catenin transcription for their abnormal proliferation, keep RB1 intact. Remarkably, E2F1 activity is also repressed by cyclin-dependent kinase-8 (CDK8), a colorectal oncoprotein. Elevated levels of CDK8 protect beta-catenin/TCF-dependent transcription from inhibition by E2F1. Thus, by retaining RB1 and amplifying CDK8, colorectal tumour cells select conditions that collectively suppress E2F1 and enhance the activity of beta-catenin.
- Published
- 2008
- Full Text
- View/download PDF
40. A nuclear receptor-like pathway regulating multidrug resistance in fungi.
- Author
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Thakur JK, Arthanari H, Yang F, Pan SJ, Fan X, Breger J, Frueh DP, Gulshan K, Li DK, Mylonakis E, Struhl K, Moye-Rowley WS, Cormack BP, Wagner G, and Näär AM
- Subjects
- Animals, Antifungal Agents metabolism, Antifungal Agents pharmacology, Candida glabrata drug effects, Candida glabrata genetics, DNA-Binding Proteins chemistry, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Fungal Proteins chemistry, Fungal Proteins genetics, Genes, Fungal genetics, Mediator Complex, Multigene Family, Pregnane X Receptor, Protein Structure, Tertiary, Saccharomyces cerevisiae drug effects, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins chemistry, Saccharomyces cerevisiae Proteins genetics, Saccharomyces cerevisiae Proteins metabolism, Trans-Activators chemistry, Trans-Activators genetics, Trans-Activators metabolism, Transcription Factors metabolism, Transcription, Genetic genetics, Xenobiotics metabolism, Candida glabrata metabolism, Drug Resistance, Fungal genetics, Fungal Proteins metabolism, Gene Expression Regulation, Fungal genetics, Receptors, Steroid metabolism, Saccharomyces cerevisiae metabolism
- Abstract
Multidrug resistance (MDR) is a serious complication during treatment of opportunistic fungal infections that frequently afflict immunocompromised individuals, such as transplant recipients and cancer patients undergoing cytotoxic chemotherapy. Improved knowledge of the molecular pathways controlling MDR in pathogenic fungi should facilitate the development of novel therapies to combat these intransigent infections. MDR is often caused by upregulation of drug efflux pumps by members of the fungal zinc-cluster transcription-factor family (for example Pdr1p orthologues). However, the molecular mechanisms are poorly understood. Here we show that Pdr1p family members in Saccharomyces cerevisiae and the human pathogen Candida glabrata directly bind to structurally diverse drugs and xenobiotics, resulting in stimulated expression of drug efflux pumps and induction of MDR. Notably, this is mechanistically similar to regulation of MDR in vertebrates by the PXR nuclear receptor, revealing an unexpected functional analogy of fungal and metazoan regulators of MDR. We have also uncovered a critical and specific role of the Gal11p/MED15 subunit of the Mediator co-activator and its activator-targeted KIX domain in antifungal/xenobiotic-dependent regulation of MDR. This detailed mechanistic understanding of a fungal nuclear receptor-like gene regulatory pathway provides novel therapeutic targets for the treatment of multidrug-resistant fungal infections.
- Published
- 2008
- Full Text
- View/download PDF
41. Mediator subunit MED28 (Magicin) is a repressor of smooth muscle cell differentiation.
- Author
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Beyer KS, Beauchamp RL, Lee MF, Gusella JF, Näär AM, and Ramesh V
- Subjects
- Animals, Cytoskeletal Proteins genetics, Humans, Intracellular Signaling Peptides and Proteins genetics, Mediator Complex, Mice, NIH 3T3 Cells, RNA Interference, Up-Regulation, Cell Transdifferentiation, Cytoskeletal Proteins metabolism, Fibroblasts cytology, Intracellular Signaling Peptides and Proteins metabolism, Myocytes, Smooth Muscle cytology
- Abstract
Magicin, a protein that we isolated earlier as an interactor of the neurofibromatosis 2 protein merlin, was independently identified as MED28, a subunit of the mammalian Mediator complex. Mediator complex is an evolutionarily conserved transcriptional cofactor, which plays an essential role in positive and negative gene regulation. Distinct Mediator subunit composition is thought to contribute to gene regulation specificity based on the interaction of specific subunits with subsets of transcription factors. Here we report that down-regulation of Med28 expression in NIH3T3 cells results in a significant induction of several genes associated with smooth muscle cell (SMC) differentiation. Conversely, overexpression of MED28 represses expression of SMC genes, in concordance with our knockdown data. More importantly, multipotent mesenchymal-derived murine precursors can transdifferentiate into SMCs when Med28 is down-regulated. Our data also show that Med28 functions as a negative regulator of SMC differentiation in concert with other Mediator subunits including Med6, Med8, and Med18 within the Mediator head module. Our results provide strong evidence that MED28 may function as a scaffolding protein by maintaining the stability of a submodule within the head module and that components of this submodule act together in a gene regulatory program to suppress SMC differentiation. The results presented here demonstrate for the first time that the mammalian Mediator subunit MED28 functions as a repressor of SMC differentiation, which could have implications for disorders associated with abnormalities in SMC growth and differentiation, including atherosclerosis, asthma, hypertension, and smooth muscle tumors.
- Published
- 2007
- Full Text
- View/download PDF
42. Retinoblastoma protein and anaphase-promoting complex physically interact and functionally cooperate during cell-cycle exit.
- Author
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Binné UK, Classon MK, Dick FA, Wei W, Rape M, Kaelin WG Jr, Näär AM, and Dyson NJ
- Subjects
- Anaphase-Promoting Complex-Cyclosome, Cadherins metabolism, Cell Cycle, Cell Line, Tumor, Cyclin-Dependent Kinase Inhibitor p27 metabolism, G1 Phase, Humans, S Phase, S-Phase Kinase-Associated Proteins metabolism, Ubiquitin metabolism, Anaphase, Retinoblastoma Protein metabolism, Ubiquitin-Protein Ligase Complexes metabolism
- Abstract
The retinoblastoma protein (pRB) negatively regulates the progression from G1 to S phase of the cell cycle, in part, by repressing E2F-dependent transcription. pRB also possesses E2F-independent functions that contribute to cell-cycle control--for example, during pRB-mediated cell-cycle arrest pRB associates with Skp2, the F-box protein of the Skp1-Cullin-F-box protein (SCF) E3 ubiquitin ligase complex, and promotes the stability of the cyclin-dependent kinase-inhibitor p27(Kip1) through an unknown mechanism. Degradation of p27(Kip1) is mediated by ubiquitin-dependent targeting of p27(Kip1) by SCF -Skp2 (ref. 4). Here, we report a novel interaction between pRB and the anaphase-promoting complex/cyclosome (APC/C) that controls p27(Kip1) stability by targeting Skp2 for ubiquitin-mediated degradation. Cdh1, an activator of APC/C, not only interacts with pRB but is also required for a pRB-induced cell-cycle arrest. The results reveal an unexpected physical convergence between the pRB tumour-suppressor protein and E3 ligase complexes, and raise the possibility that pRB may direct APC/C to additional targets during pRB-mediated cell-cycle exit.
- Published
- 2007
- Full Text
- View/download PDF
43. An ARC/Mediator subunit required for SREBP control of cholesterol and lipid homeostasis.
- Author
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Yang F, Vought BW, Satterlee JS, Walker AK, Jim Sun ZY, Watts JL, DeBeaumont R, Saito RM, Hyberts SG, Yang S, Macol C, Iyer L, Tjian R, van den Heuvel S, Hart AC, Wagner G, and Näär AM
- Subjects
- Amino Acid Sequence, Animals, Caenorhabditis elegans, Humans, Mediator Complex, Mice, Models, Molecular, Molecular Sequence Data, Nuclear Magnetic Resonance, Biomolecular, Protein Binding, Protein Structure, Tertiary, Sterol Regulatory Element Binding Proteins chemistry, Sterol Regulatory Element Binding Proteins genetics, Transcriptional Activation, Caenorhabditis elegans Proteins metabolism, Cholesterol metabolism, Homeostasis, Lipid Metabolism, Sterol Regulatory Element Binding Proteins metabolism, Transcription Factors metabolism
- Abstract
The sterol regulatory element binding protein (SREBP) family of transcription activators are critical regulators of cholesterol and fatty acid homeostasis. We previously demonstrated that human SREBPs bind the CREB-binding protein (CBP)/p300 acetyltransferase KIX domain and recruit activator-recruited co-factor (ARC)/Mediator co-activator complexes through unknown mechanisms. Here we show that SREBPs use the evolutionarily conserved ARC105 (also called MED15) subunit to activate target genes. Structural analysis of the SREBP-binding domain in ARC105 by NMR revealed a three-helix bundle with marked similarity to the CBP/p300 KIX domain. In contrast to SREBPs, the CREB and c-Myb activators do not bind the ARC105 KIX domain, although they interact with the CBP KIX domain, revealing a surprising specificity among structurally related activator-binding domains. The Caenorhabditis elegans SREBP homologue SBP-1 promotes fatty acid homeostasis by regulating the expression of lipogenic enzymes. We found that, like SBP-1, the C. elegans ARC105 homologue MDT-15 is required for fatty acid homeostasis, and show that both SBP-1 and MDT-15 control transcription of genes governing desaturation of stearic acid to oleic acid. Notably, dietary addition of oleic acid significantly rescued various defects of nematodes targeted with RNA interference against sbp-1 and mdt-15, including impaired intestinal fat storage, infertility, decreased size and slow locomotion, suggesting that regulation of oleic acid levels represents a physiologically critical function of SBP-1 and MDT-15. Taken together, our findings demonstrate that ARC105 is a key effector of SREBP-dependent gene regulation and control of lipid homeostasis in metazoans.
- Published
- 2006
- Full Text
- View/download PDF
44. Integrator, a multiprotein mediator of small nuclear RNA processing, associates with the C-terminal repeat of RNA polymerase II.
- Author
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Baillat D, Hakimi MA, Näär AM, Shilatifard A, Cooch N, and Shiekhattar R
- Subjects
- Amino Acid Sequence, Amino Acid Substitution, Binding Sites, Blotting, Western, Carrier Proteins chemistry, Carrier Proteins metabolism, Cell Line, Chromatin Immunoprecipitation, Conserved Sequence, Endoribonucleases, Escherichia coli genetics, Evolution, Molecular, Glyceraldehyde-3-Phosphate Dehydrogenases analysis, Glyceraldehyde-3-Phosphate Dehydrogenases metabolism, HeLa Cells, Humans, Models, Biological, Molecular Sequence Data, Protein Structure, Tertiary, Protein Subunits chemistry, RNA biosynthesis, RNA Polymerase II chemistry, RNA, Messenger analysis, RNA, Messenger metabolism, RNA, Small Interfering metabolism, RNA, Small Nuclear genetics, Recombinant Fusion Proteins isolation & purification, Recombinant Fusion Proteins metabolism, Sequence Homology, Amino Acid, Transcription, Genetic, RNA Polymerase II metabolism, RNA Processing, Post-Transcriptional, RNA, Small Nuclear metabolism
- Abstract
The C-terminal domain (CTD) of RNA polymerase II (RNAPII) is an essential component of transcriptional regulation and RNA processing of protein-coding genes. A large body of data also implicates the CTD in the transcription and processing of RNAPII-mediated small nuclear RNAs (snRNAs). However, the identity of the complex (or complexes) that associates with the CTD and mediates the processing of snRNAs has remained elusive. Here, we describe an RNA polymerase II complex that contains at least 12 novel subunits, termed the Integrator, in addition to core RNAPII subunits. Two of the Integrator subunits display similarities to the subunits of the cleavage and polyadenylation specificity factor (CPSF) complex. We show that Integrator is recruited to the U1 and U2 snRNA genes and mediates the snRNAs' 3' end processing. The Integrator complex is evolutionarily conserved in metazoans and directly interacts with the C-terminal domain of the RNA polymerase II largest subunit.
- Published
- 2005
- Full Text
- View/download PDF
45. The activator-recruited cofactor/Mediator coactivator subunit ARC92 is a functionally important target of the VP16 transcriptional activator.
- Author
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Yang F, DeBeaumont R, Zhou S, and Näär AM
- Subjects
- Base Sequence, Cell Nucleus metabolism, Cloning, Molecular, DNA Primers, DNA, Complementary genetics, HeLa Cells, Humans, Mediator Complex, Molecular Sequence Data, Polymerase Chain Reaction, RNA Polymerase II, RNA, Small Interfering genetics, Recombinant Fusion Proteins metabolism, Recombinant Proteins metabolism, Trans-Activators genetics, Transcription Factors genetics, Transcription, Genetic, Transcriptional Activation genetics, Transfection, Trans-Activators metabolism, Transcription Factors metabolism
- Abstract
The human activator-recruited cofactor (ARC), a family of large transcriptional coactivator complexes related to the yeast Mediator, was recently identified based on functional association with the activation domains of multiple cellular and viral transcriptional activators, including the herpes simplex viral activator VP16, sterol regulatory element binding protein, and NF-kappaB. Here we describe the biochemical purification and cloning of the 92-kDa ARC/Mediator subunit, ARC92, that is specifically targeted by the activation domain of the VP16 transactivator. Affinity chromatography using the VP16 activation domain followed by peptide microsequencing led to the identification of ARC92 as a specific cellular interaction partner of the VP16 activation domain. ARC92 associates with the VP16 activation domain in vitro and in vivo, and the VP16 binding domain of ARC92 is a strong competitive inhibitor of Gal4-VP16 in vivo. Moreover, small interfering RNA-mediated knockdown of ARC92 in human cells results in selective inhibition of Gal4-VP16 gene activation. Taken together, our results suggest that ARC92 is a direct and specific target of the VP16 transactivator that serves in the context of the ARC/Mediator coactivator as an important transducer of transcription activating signals from the VP16 activation domain to the RNA polymerase II transcriptional machinery.
- Published
- 2004
- Full Text
- View/download PDF
46. A component of the ARC/Mediator complex required for TGF beta/Nodal signalling.
- Author
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Kato Y, Habas R, Katsuyama Y, Näär AM, and He X
- Subjects
- Activins metabolism, Animals, Bone Morphogenetic Proteins pharmacology, Cell Differentiation drug effects, Cell Line, DNA-Binding Proteins metabolism, Gene Expression Regulation, Developmental, Humans, Macromolecular Substances, Mesoderm cytology, Mesoderm drug effects, Mesoderm metabolism, Nerve Growth Factors, Nodal Protein, Promoter Regions, Genetic genetics, Protein Binding, RNA, Messenger genetics, RNA, Messenger metabolism, Smad Proteins, Smad2 Protein, Smad3 Protein, Smad4 Protein, Trans-Activators metabolism, Xenopus laevis genetics, Signal Transduction drug effects, Transforming Growth Factor beta metabolism, Transforming Growth Factor beta pharmacology, Xenopus Proteins chemistry, Xenopus Proteins metabolism, Xenopus laevis embryology, Xenopus laevis metabolism
- Abstract
The transforming growth factor beta (TGF beta) family of cytokines, including Nodal, Activin and bone morphogenetic protein (BMP), have essential roles in development and tumorigenesis. TGF beta molecules activate the Smad family of signal transducers, which form complexes with specific DNA-binding proteins to regulate gene expression. Two discrete Smad-dependent signalling pathways have been identified: TGF beta, Activin and Nodal signal via the Smad2 (or Smad3)-Smad4 complex, whereas BMP signals via the Smad1-Smad4 complex. How distinct Smad complexes regulate specific gene expression is not fully understood. Here we show that ARC105, a component of the activator-recruited co-factor (ARC) complex or the metazoan Mediator complex, is essential for TGF beta/Activin/Nodal/Smad2/3 signal transduction. Expression of ARC105 stimulates Activin/Nodal/Smad2 signalling in Xenopus laevis embryos, inducing axis duplication and mesendoderm differentiation, and enhances TGF beta response in human cells. Depletion of ARC105 inhibits TGF beta/Activin/Nodal/Smad2/3 signalling and Xenopus axis formation, but not BMP/Smad1 signalling. ARC105 protein binds to Smad2/3-Smad4 in response to TGF beta and is recruited to Activin/Nodal-responsive promoters in chromatin in a Smad2-dependent fashion. Thus ARC105 is a specific and key ARC/Mediator component linking TGF beta/Activin/Nodal/Smad2/3 signalling to transcriptional activation.
- Published
- 2002
- Full Text
- View/download PDF
47. Human CRSP interacts with RNA polymerase II CTD and adopts a specific CTD-bound conformation.
- Author
-
Näär AM, Taatjes DJ, Zhai W, Nogales E, and Tjian R
- Subjects
- Binding Sites, Chromatin metabolism, Electrophoresis, Polyacrylamide Gel, Glutathione Transferase metabolism, HeLa Cells, Humans, Immunoblotting, Ligands, Mediator Complex, Microscopy, Electron, Models, Molecular, Protein Binding, Protein Conformation, Protein Structure, Tertiary, Recombinant Fusion Proteins metabolism, RNA Polymerase II chemistry, Trans-Activators chemistry, Trans-Activators metabolism, Transcription, Genetic
- Abstract
Activation of gene transcription in mammalian cells requires several classes of coactivators that participate in different steps of the activation cascade. Using conventional and affinity chromatography, we have isolated a human coactivator complex that interacts directly with the C-terminal domain (CTD) of RNA polymerase II (Pol II). The CTD-binding complex is structurally and functionally indistinguishable from our previously isolated CRSP coactivator complex. The closely related, but transcriptionally inactive, ARC-L complex failed to interact with the CTD, indicating a significant biochemical difference between CRSP and ARC-L that may, in part, explain their functional divergence. Electron microscopy and three-dimensional single-particle reconstruction reveals a conformation for CTD-CRSP that is structurally distinct from unliganded CRSP or CRSP bound to SREBP-1a, but highly similar to CRSP bound to the VP16 activator. Together, our findings suggest that the human CRSP coactivator functions, at least in part, by mediating activator-dependent recruitment of RNA Pol II via the CTD.
- Published
- 2002
- Full Text
- View/download PDF
48. Structure, function, and activator-induced conformations of the CRSP coactivator.
- Author
-
Taatjes DJ, Näär AM, Andel F 3rd, Nogales E, and Tjian R
- Subjects
- CCAAT-Enhancer-Binding Proteins chemistry, CCAAT-Enhancer-Binding Proteins metabolism, Chromatin metabolism, DNA-Binding Proteins chemistry, DNA-Binding Proteins metabolism, HeLa Cells, Herpes Simplex Virus Protein Vmw65 metabolism, Humans, Image Processing, Computer-Assisted, Imaging, Three-Dimensional, Macromolecular Substances, Microscopy, Electron, Models, Genetic, Precipitin Tests, Protein Binding, Protein Structure, Quaternary, Protein Structure, Tertiary, Protein Subunits, Recombinant Fusion Proteins chemistry, Recombinant Fusion Proteins metabolism, Recombinant Proteins metabolism, Sterol Regulatory Element Binding Protein 1, Trans-Activators isolation & purification, Transcription Factors metabolism, Transcriptional Activation, Trans-Activators chemistry, Trans-Activators metabolism, Transcription, Genetic
- Abstract
The human cofactor complexes ARC (activator-recruited cofactor) and CRSP (cofactor required for Sp1 activation) mediate activator-dependent transcription in vitro. Although these complexes share several common subunits, their structural and functional relationships remain unknown. Here, we report that affinity-purified ARC consists of two distinct multisubunit complexes: a larger complex, denoted ARC-L, and a smaller coactivator, CRSP. Reconstituted in vitro transcription with biochemically separated ARC-L and CRSP reveals differential cofactor functions. The ARC-L complex is transcriptionally inactive, whereas the CRSP complex is highly active. Structural determination by electron microscopy (EM) and three-dimensional reconstruction indicate substantial differences in size and shape between ARC-L and CRSP. Moreover, EM analysis of independently derived CRSP complexes reveals distinct conformations induced by different activators. These results suggest that CRSP may potentiate transcription via specific activator-induced conformational changes.
- Published
- 2002
- Full Text
- View/download PDF
49. Transcriptional coactivator complexes.
- Author
-
Näär AM, Lemon BD, and Tjian R
- Subjects
- Acetyltransferases metabolism, Adenosine Triphosphate metabolism, Chromatin genetics, DNA-Binding Proteins chemistry, Histone Acetyltransferases, Nuclear Proteins chemistry, Trans-Activators chemistry, Trans-Activators physiology, Transcription Factor TFIID, Transcription Factors, TFII chemistry, Chromatin metabolism, DNA-Binding Proteins physiology, Nuclear Proteins physiology, Saccharomyces cerevisiae Proteins, TATA-Binding Protein Associated Factors, Transcription Factors, TFII physiology, Transcriptional Activation
- Abstract
The last two decades have witnessed a tremendous expansion in our knowledge of the mechanisms employed by eukaryotic cells to control gene activity. A critical insight to transcriptional control mechanisms was provided by the discovery of coactivators, a diverse array of cellular factors that connect sequence-specific DNA binding activators to the general transcriptional machinery, or that help activators and the transcriptional apparatus to navigate through the constraints of chromatin. A number of coactivators have been isolated as large multifunctional complexes, and biochemical, genetic, molecular, and cellular strategies have all contributed to uncovering many of their components, activities, and modes of action. Coactivator functions can be broadly divide into two classes: (a) adaptors that direct activator recruitment of the transcriptional apparatus, (b) chromatin-remodeling or -modifying enzymes. Strikingly, several distinct coactivator complexes nonetheless share many subunits and appear to be assembled in a modular fashion. Such structural and functional modularity could provide the cell with building blocks from which to construct a versatile array of coactivator complexes according to its needs. The extent of functional interplay between these different activities in gene-specific transcriptional regulation is only now becoming apparent, and will remain an active area of research for years to come.
- Published
- 2001
- Full Text
- View/download PDF
50. Ligand-dependent transcription activation by nuclear receptors requires the DRIP complex.
- Author
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Rachez C, Lemon BD, Suldan Z, Bromleigh V, Gamble M, Näär AM, Erdjument-Bromage H, Tempst P, and Freedman LP
- Subjects
- Amino Acid Sequence, Animals, Carrier Proteins physiology, Chromatin physiology, Cloning, Molecular, Drosophila, HeLa Cells, Humans, Ligands, Macromolecular Substances, Mediator Complex, Mediator Complex Subunit 1, Mice, Molecular Sequence Data, Nuclear Proteins chemistry, Sequence Homology, Amino Acid, Nuclear Proteins physiology, Receptors, Calcitriol physiology, Trans-Activators, Transcription Factors, Transcriptional Activation
- Abstract
Nuclear receptors modulate the transcription of genes in direct response to small lipophilic ligands. Binding to ligands induces conformational changes in the nuclear receptors that enable the receptors to interact with several types of cofactor that are critical for transcription activation (transactivation). We previously described a distinct set of ligand-dependent proteins called DRIPs, which interact with the vitamin D receptor (VDR); together, these proteins constitute a new cofactor complex. DRIPs bind to several nuclear receptors and mediate ligand-dependent enhancement of transcription by VDR and the thyroid-hormone receptor in cell-free transcription assays. Here we report the identities of thirteen DRIPs that constitute this complex, and show that the complex has a central function in hormone-dependent transactivation by VDR on chromatin templates. The DRIPs are almost indistinguishable from components of another new cofactor complex called ARC, which is recruited by other types of transcription activators to mediate transactivation on chromatin-assembled templates. Several DRIP/ARC subunits are also components of other potentially related cofactors, such as CRSP, NAT, SMCC and the mouse Mediator, indicating that unique classes of activators may share common sets or subsets of cofactors. The role of nuclear-receptor ligands may, in part, be to recruit such a cofactor complex to the receptor and, in doing so, to enhance transcription of target genes.
- Published
- 1999
- Full Text
- View/download PDF
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