The Experts below are selected from a list of 73077 Experts worldwide ranked by ideXlab platform
Lubo Zhang - One of the best experts on this subject based on the ideXlab platform.
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norepinephrine causes epiGenetic Repression of pkce Gene in rodent hearts by activating nox1 dependent reactive oxygen species production
The FASEB Journal, 2012Co-Authors: Fuxia Xiong, Daliao Xiao, Lubo ZhangAbstract:Heart disease is the leading cause of death in the United States. Recent studies demonstrate that fetal programming of PKCe Gene Repression results in ischemia-sensitive phenotype in the heart. The present study tests the hypothesis that increased norepinephrine causes epiGenetic Repression of PKCe Gene in the heart via Nox1-dependent reactive oxygen species (ROS) production. Prolonged norepinephrine treatment increased ROS production in fetal rat hearts and embryonic ventricular myocyte H9c2 cells via a selective increase in Nox1 expression. Norepinephrine-induced ROS resulted in an increase in PKCe promoter methylation at Egr-1 and Sp-1 binding sites, leading to PKCe Gene Repression. N-acetylcysteine, diphenyleneiodonium, and apocynin blocked norepinephrine-induced ROS production and the promoter methylation, and also restored PKCe mRNA and protein to control levels in vivo in fetal hearts and in vitro in embryonic myocyte cells. Accordingly, norepinephrine-induced ROS production, promoter methylation, and PKCe Gene Repression were completely abrogated by knockdown of Nox1 in cardiomyocytes. These findings provide evidence of a novel interaction between elevated norepinephrine and epiGenetic Repression of PKCe Gene in the heart mediated by Nox1-dependent oxidative stress and suggest new insights of molecular mechanisms linking the heightened sympathetic activity to aberrant cardioprotection and increased ischemic vulnerability in the heart.—Xiong, F., Xiao, D., Zhang, L. Norepinephrine causes epiGenetic Repression of PKCe Gene in rodent hearts by activating Nox1-dependent reactive oxygen species production.
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norepinephrine causes epiGenetic Repression of pkce Gene in rodent hearts by activating nox1 dependent reactive oxygen species production
The FASEB Journal, 2012Co-Authors: Fuxia Xiong, Daliao Xiao, Lubo ZhangAbstract:Heart disease is the leading cause of death in the United States. Recent studies demonstrate that fetal programming of PKCe Gene Repression results in ischemia-sensitive phenotype in the heart. The present study tests the hypothesis that increased norepinephrine causes epiGenetic Repression of PKCe Gene in the heart via Nox1-dependent reactive oxygen species (ROS) production. Prolonged norepinephrine treatment increased ROS production in fetal rat hearts and embryonic ventricular myocyte H9c2 cells via a selective increase in Nox1 expression. Norepinephrine-induced ROS resulted in an increase in PKCe promoter methylation at Egr-1 and Sp-1 binding sites, leading to PKCe Gene Repression. N-acetylcysteine, diphenyleneiodonium, and apocynin blocked norepinephrine-induced ROS production and the promoter methylation, and also restored PKCe mRNA and protein to control levels in vivo in fetal hearts and in vitro in embryonic myocyte cells. Accordingly, norepinephrine-induced ROS production, promoter methylation, and PKCe Gene Repression were completely abrogated by knockdown of Nox1 in cardiomyocytes. These findings provide evidence of a novel interaction between elevated norepinephrine and epiGenetic Repression of PKCe Gene in the heart mediated by Nox1-dependent oxidative stress and suggest new insights of molecular mechanisms linking the heightened sympathetic activity to aberrant cardioprotection and increased ischemic vulnerability in the heart.
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chronic prenatal hypoxia induces epiGenetic programming of pkce Gene Repression in rat hearts
Circulation Research, 2010Co-Authors: Andrew J Patterson, Daliao Xiao, Man Chen, Qin Xue, Lubo ZhangAbstract:Rationale: Epidemiological studies demonstrate a clear association of adverse intrauterine environment with an increased risk of ischemic heart disease in adulthood. Hypoxia is a common stress to the fetus and results in decreased protein kinase C epsilon (PKCe) expression in the heart and increased cardiac vulnerability to ischemia and reperfusion injury in adult offspring in rats. Objectives: The present study tested the hypothesis that fetal hypoxia-induced methylation of cytosine-phosphate-guanine dinucleotides at the PKCe promoter is repressive and contributes to PKCe Gene Repression in the heart of adult offspring. Methods and Results: Hypoxic treatment of pregnant rats from days 15 to 21 of gestation resulted in significant decreases in PKCe protein and mRNA in fetal hearts. Similar results were obtained in ex vivo hypoxic treatment of isolated fetal hearts and rat embryonic ventricular myocyte cell line H9c2. Increased methylation of PKCe promoter at SP1 binding sites, −346 and −268, were demonstrated in both fetal hearts of maternal hypoxia and H9c2 cells treated with 1% O 2 for 24 hours. Whereas hypoxia had no significant effect on the binding affinity of SP1 to the unmethylated sites in H9c2 cells, hearts of fetuses and adult offspring, methylation of both SP1 sites reduced SP1 binding. The addition of 5-aza-2′-deoxycytidine blocked the hypoxia-induced increase in methylation of both SP1 binding sites and restored PKCe mRNA and protein to the control levels. In hearts of both fetuses and adult offspring, hypoxia-induced methylation of SP1 sites was significantly greater in males than in females, and decreased PKCe mRNA was seen only in males. In fetal hearts, there was significantly higher abundance of estrogen receptor α and β isoforms in females than in males. Both estrogen receptor α and β interacted with the SP1 binding sites in the fetal heart, which may explain the sex differences in SP1 methylation in the fetal heart. Additionally, selective activation of PKCe restored the hypoxia-induced cardiac vulnerability to ischemic injury in offspring. Conclusions: The findings demonstrate a direct effect of hypoxia on epiGenetic modification of DNA methylation and programming of cardiac PKCe Gene Repression in a sex-dependent manner, linking fetal hypoxia and pathophysiological consequences in the hearts of adult offspring.
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direct effect of cocaine on epiGenetic regulation of pkcɛ Gene Repression in the fetal rat heart
Journal of Molecular and Cellular Cardiology, 2009Co-Authors: K Meyer, Haitao Zhang, Lubo ZhangAbstract:Abstract Maternal cocaine administration during gestation caused a down-regulation of PKCɛ expression in the heart of adult offspring resulting in an increased sensitivity to ischemia and reperfusion injury. The present study investigated the direct effect of cocaine in epiGenetic modification of PKCɛ Gene Repression in the fetal heart. Hearts were isolated from gestational day 17 fetal rats and treated with cocaine in an ex vivo organ culture system. Cocaine treatment for 48 h resulted in significant decreases in PKCɛ protein and mRNA abundance and increases in CpG methylation at two SP1 binding sites in the PKCɛ promoter region (− 346 and − 268). Electrophoretic mobility shift assays demonstrated that CpG methylation of both SP1 sites inhibited SP1 binding. Consistently, chromatin immunoprecipitation assays showed that cocaine treatment significantly decreased binding of SP1 to the SP1 sites in the intact fetal heart. Reporter Gene assays revealed that site-directed mutations of CpG methylation at both SP1 sites significantly reduced the PKCɛ promoter activity while methylation of a single site at either − 346 or − 268 did not have a significant effect. The causal effect of increased methylation in the cocaine-induced down-regulation of PKCɛ was demonstrated with the use of DNA methylation inhibitors. The presence of either 5-aza-2′-deoxycytodine or procainamide blocked the cocaine-induced increase in SP1 sites methylation and decrease in PKCɛ mRNA. The results demonstrate a direct effect of cocaine in epiGenetic modification of DNA methylation and programming of cardiac PKCɛ Gene Repression linking prenatal cocaine exposure and pathophysiological consequences in the heart of adult offspring.
Robert J Klose - One of the best experts on this subject based on the ideXlab platform.
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prc1 drives polycomb mediated Gene Repression by controlling transcription initiation and burst frequency
Nature Structural & Molecular Biology, 2021Co-Authors: Paula Dobrinic, Aleksander T Szczurek, Robert J KloseAbstract:The Polycomb repressive system plays a fundamental role in controlling Gene expression during mammalian development. To achieve this, Polycomb repressive complexes 1 and 2 (PRC1 and PRC2) bind target Genes and use histone modification-dependent feedback mechanisms to form Polycomb chromatin domains and repress transcription. The inter-relatedness of PRC1 and PRC2 activity at these sites has made it difficult to discover the specific components of Polycomb chromatin domains that drive Gene Repression and to understand mechanistically how this is achieved. Here, by exploiting rapid degron-based approaches and time-resolved genomics, we kinetically dissect Polycomb-mediated Repression and discover that PRC1 functions independently of PRC2 to counteract RNA polymerase II binding and transcription initiation. Using single-cell Gene expression analysis, we reveal that PRC1 acts uniformly within the cell population and that Repression is achieved by controlling transcriptional burst frequency. These important new discoveries provide a mechanistic and conceptual framework for Polycomb-dependent transcriptional control. Here the authors find that Polycomb repressive complex PRC1 functions independently of PRC2 to counteract Pol II binding, regulating transcription initiation and burst frequency.
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live cell single particle tracking of prc1 reveals a highly dynamic system with low target site occupancy
Nature Communications, 2021Co-Authors: Miles K Huseyin, Robert J KloseAbstract:Polycomb repressive complex 1 (PRC1) is an essential chromatin-based repressor of Gene transcription. How PRC1 engages with chromatin to identify its target Genes and achieve Gene Repression remains poorly defined, representing a major hurdle to our understanding of Polycomb system function. Here, we use genome engineering and single particle tracking to dissect how PRC1 binds to chromatin in live mouse embryonic stem cells. We observe that PRC1 is highly dynamic, with only a small fraction stably interacting with chromatin. By integrating subunit-specific dynamics, chromatin binding, and abundance measurements, we discover that PRC1 exhibits low occupancy at target sites. Furthermore, we employ perturbation approaches to uncover how specific components of PRC1 define its kinetics and chromatin binding. Together, these discoveries provide a quantitative understanding of chromatin binding by PRC1 in live cells, suggesting that chromatin modification, as opposed to PRC1 complex occupancy, is central to Gene Repression. How PRC1 recognises and interacts with its target Genes remains poorly understood. Here, the authors use genome engineering and single particle tracking to dissect how PRC1 binds to chromatin in live mouse embryonic stem cells, revealing that this repressor is highly dynamic, with only a small fraction stably interacting with chromatin.
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prc1 drives polycomb mediated Gene Repression by controlling transcription initiation and burst frequency
bioRxiv, 2020Co-Authors: Paula Dobrinic, Aleksander T Szczurek, Robert J KloseAbstract:The Polycomb repressive system plays a fundamental role in controlling Gene expression during mammalian development. To achieve this, Polycomb repressive complexes 1 and 2 (PRC1 and PRC2) bind target Genes and use histone modification-dependent feedback mechanisms to form Polycomb chromatin domains and repress transcription. The interrelatedness of PRC1 and PRC2 activity at these sites has made it difficult to discover the specific components of Polycomb chromatin domains that drive Gene Repression and to understand mechanistically how this is achieved. Here, by exploiting rapid degron-based approaches and time-resolved genomics we kinetically dissect Polycomb-mediated Repression and discover that PRC1 functions independently of PRC2 to counteract RNA polymerase II binding and transcription initiation. Using single-cell Gene expression analysis, we reveal that PRC1 acts uniformly within the cell population, and that Repression is achieved by controlling transcriptional burst frequency. These important new discoveries provide a mechanistic and conceptual framework for Polycomb-dependent transcriptional control.
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live cell single particle tracking of prc1 reveals a highly dynamic system with low target site occupancy
bioRxiv, 2020Co-Authors: Miles K Huseyin, Robert J KloseAbstract:Polycomb repressive complex 1 (PRC1) is an essential chromatin-based repressor of Gene transcription. However, how PRC1 engages with chromatin to identify its target Genes and achieve Gene Repression remains poorly defined, representing a major hurdle to our understanding of Polycomb system function. Here we use genome engineering and single particle tracking to dissect how PRC1 binds to chromatin in live mouse embryonic stem cells. We reveal that PRC1 is highly dynamic, with only a small fraction stably interacting with chromatin. By integrating subunit-specific dynamics, chromatin binding, and abundance measurements, we discover that PRC1 exhibits surprisingly low occupancy at target sites. Furthermore, we employ perturbation approaches to uncover how specific components of PRC1 define its kinetics and chromatin binding. Together, these discoveries provide a quantitative understanding of chromatin binding by PRC1 in live cells, and suggests that chromatin modification, as opposed to PRC1 complex occupancy, is central to Gene Repression.
John Svaren - One of the best experts on this subject based on the ideXlab platform.
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regulation of peripheral nerve myelin maintenance by Gene Repression through polycomb repressive complex 2
The Journal of Neuroscience, 2015Co-Authors: Ki H, Rajini Srinivasan, Holly A Hung, Huafeng Xie, Stuart H Orkin, John SvarenAbstract:Myelination of peripheral nerves by Schwann cells requires coordinate regulation of Gene Repression as well as Gene activation. Several chromatin remodeling pathways critical for peripheral nerve myelination have been identified, but the functions of histone methylation in the peripheral nerve have not been elucidated. To determine the role of histone H3 Lys27 methylation, we have Generated mice with a Schwann cell-specific knock-out of Eed, which is an essential subunit of the polycomb repressive complex 2 (PRC2) that catalyzes methylation of histone H3 Lys27. Analysis of this mutant revealed no significant effects on early postnatal development of myelin. However, its loss eventually causes progressive hypermyelination of small-diameter axons and apparent fragmentation of Remak bundles. These data identify the PRC2 complex as an epigenomic modulator of mature myelin thickness, which is associated with changes in Akt phosphorylation. Interestingly, we found that Eed inactivation causes deRepression of several Genes, e.g., Sonic hedgehog (Shh) and Insulin-like growth factor-binding protein 2 (Igfbp2), that become activated after nerve injury, but without activation of a primary regulator of the injury program, c-Jun. Analysis of the activated Genes in cultured Schwann cells showed that Igfbp2 regulates Akt activation. Our results identify an epigenomic pathway required for establishing thickness of mature myelin and repressing Genes that respond to nerve injury.
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active Gene Repression by the egr2 nab complex during peripheral nerve myelination
Journal of Biological Chemistry, 2008Co-Authors: Gennifer M. Mager, Sungwook Jang, Lawrence Wrabetz, Rajini Srinivasan, Rebecca Ward, John SvarenAbstract:Abstract The Egr2/Krox20 transactivator is required for activation of many myelin-associated Genes during peripheral nerve myelination by Schwann cells. However, recent work has indicated that Egr2 not only activates Genes required for peripheral nerve myelination but may also be involved in Gene Repression. The NAB (NGFI-A/Egr-binding) corepressors interact with Egr2 and are required for proper coordination of myelin formation. Therefore, NAB proteins could mediate Repression of some Egr2 target Genes, although direct Repression by Egr2 or NAB proteins during myelination has not been demonstrated. To define the physiological role of NAB coRepression in Gene Repression by Egr2, we tested whether the Egr2·NAB complex directly repressed specific target Genes. A screen for NAB-regulated Genes identified several (including Id2, Id4, and Rad) that declined during the course of peripheral nerve myelination. In vivo chromatin immunoprecipitation analysis of the myelinating sciatic nerve was used to show developmental association of both Egr2 and NAB2 on the Id2, Id4, and Rad promoters as they were repressed during the myelination process. In addition, NAB2 represses transcription by interaction with the chromodomain helicase DNA-binding protein 4 (CHD4) subunit of the nucleosome remodeling and deacetylase chromatin remodeling complex, and we demonstrate that CHD4 occupies NAB-repressed promoters in a developmentally regulated manner in vivo. These results illustrate a novel aspect of Genetic regulation of peripheral nerve myelination by showing that Egr2 directly represses Genes during myelination in conjunction with NAB corepressors. Furthermore, Repression of Id2 was found to augment activation of Mpz (myelin protein zero) expression.
Chase L Beisel - One of the best experts on this subject based on the ideXlab platform.
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rapid and scalable characterization of crispr technologies using an e coli cell free transcription translation system
bioRxiv, 2017Co-Authors: Ryan Marshall, Colin S Maxwell, Scott P Collins, Michelle L Luo, Thomas Jacobsen, Chase L Beisel, Vincent NoireauxAbstract:CRISPR-Cas systems have offered versatile technologies for genome engineering, yet their implementation has been outpaced by the ongoing discovery of new Cas nucleases and anti-CRISPR proteins. Here, we present the use of E. coli cell-free transcription-translation systems (TXTL) to vastly improve the speed and scalability of CRISPR characterization and validation. Unlike prior approaches that require protein purification or live cells, TXTL can express active CRISPR machinery from added plasmids and linear DNA, and TXTL can output quantitative dynamics of DNA cleavage and Gene Repression. To demonstrate the applicability of TXTL, we rapidly measure guide RNA-dependent DNA cleavage and Gene Repression for single- and multi-effector CRISPR-Cas systems, accurately predict the strength of Gene Repression in E. coli, quantify the inhibitory activity of anti-CRISPR proteins, and develop a fast and scalable high-throughput screen for protospacer-adjacent motifs. These examples underscore the potential of TXTL to facilitate the characterization and application of CRISPR technologies across their many uses.
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repurposing endogenous type i crispr cas systems for programmable Gene Repression
Nucleic Acids Research, 2015Co-Authors: Michelle L Luo, Adam S Mullis, Ryan T Leenay, Chase L BeiselAbstract:CRISPR-Cas systems have shown tremendous promise as heterologous tools for genome editing and transcriptional regulation. Because these RNA-directed immune systems are found in most prokaryotes, an opportunity exists to harness the endogenous systems as convenient tools in these organisms. Here, we report that the Type I-E CRISPR-Cas system in Escherichia coli can be co-opted for programmable transcriptional Repression. We found that deletion of the signature cas3 Gene converted this immune system into a programmable Gene regulator capable of reversible Gene silencing of heterologous and endogenous Genes. Targeting promoter regions yielded the strongest Repression, whereas targeting coding regions showed consistent strand bias. Furthermore, multi-targeting CRISPR arrays could Generate complex phenotypes. This strategy offers a simple approach to convert many endogenous Type I systems into transcriptional regulators, thereby expanding the available toolkit for CRISPR-mediated Genetic control while creating new opportunities for genome-wide screens and pathway engineering.
Xiaobing Shi - One of the best experts on this subject based on the ideXlab platform.
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recognition of unmethylated histone h3 lysine 4 links bhc80 to lsd1 mediated Gene Repression
Nature, 2007Co-Authors: Robe E Collins, Xiaobing Shi, Rossella De Cegli, Roma Alpatov, Joh R Horto, O Gozani, Xiaodong Cheng, Yang ShiAbstract:BHC80 is a component of the LSD1 co-repressor complex that demethylates histone H3 at lysine 4. The PHD domain of BHC80 interacts with the histone H3 tail only when lysine 4 is unmethylated, and BHC80 function is coupled to that of LSD1 in Gene Repression. Histone methylation is crucial for regulating chromatin structure, Gene transcription and the epiGenetic state of the cell. LSD1 is a lysine-specific histone demethylase that represses transcription by demethylating histone H3 on lysine 4 (ref. 1). The LSD1 complex contains a number of proteins, all of which have been assigned roles in events upstream of LSD1-mediated demethylation2,3,4 apart from BHC80 (also known as PHF21A), a plant homeodomain (PHD) finger-containing protein. Here we report that, in contrast to the PHD fingers of the bromodomain PHD finger transcription factor (BPTF) and inhibitor of growth family 2 (ING2), which bind methylated H3K4 (H3K4me3)5,6, the PHD finger of BHC80 binds unmethylated H3K4 (H3K4me0), and this interaction is specifically abrogated by methylation of H3K4. The crystal structure of the PHD finger of BHC80 bound to an unmodified H3 peptide has revealed the structural basis of the recognition of H3K4me0. Knockdown of BHC80 by RNA inhibition results in the de-Repression of LSD1 target Genes, and this Repression is restored by the reintroduction of wild-type BHC80 but not by a PHD-finger mutant that cannot bind H3. Chromatin immunoprecipitation showed that BHC80 and LSD1 depend reciprocally on one another to associate with chromatin. These findings couple the function of BHC80 to that of LSD1, and indicate that unmodified H3K4 is part of the ‘histone code’7. They further raise the possibility that the Generation and recognition of the unmodified state on histone tails in General might be just as crucial as post-translational modifications of histone for chromatin and transcriptional regulation.
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ing2 phd domain links histone h3 lysine 4 methylation to active Gene Repression
Nature, 2006Co-Authors: Xiaobing Shi, Tao Hong, Kay L Walte, Mark D Ewal, Eriko Michishita, Tiffany Hung, Dyla Carney, Pedro V Pena, Moha R Kaadige, Nicolas LacosteAbstract:Dynamic regulation of diverse nuclear processes is intimately linked to covalent modifications of chromatin. Much attention has focused on methylation at lysine 4 of histone H3 (H3K4), owing to its association with euchromatic genomic regions. H3K4 can be mono-, di- or tri-methylated. Trimethylated H3K4 (H3K4me3) is preferentially detected at active Genes, and is proposed to promote Gene expression through recognition by transcription-activating effector molecules. Here we identify a novel class of methylated H3K4 effector domains--the PHD domains of the ING (for inhibitor of growth) family of tumour suppressor proteins. The ING PHD domains are specific and highly robust binding modules for H3K4me3 and H3K4me2. ING2, a native subunit of a repressive mSin3a-HDAC1 histone deacetylase complex, binds with high affinity to the trimethylated species. In response to DNA damage, recognition of H3K4me3 by the ING2 PHD domain stabilizes the mSin3a-HDAC1 complex at the promoters of proliferation Genes. This pathway constitutes a new mechanism by which H3K4me3 functions in active Gene Repression. Furthermore, ING2 modulates cellular responses to genotoxic insults, and these functions are critically dependent on ING2 interaction with H3K4me3. Together, our findings establish a pivotal role for trimethylation of H3K4 in Gene Repression and, potentially, tumour suppressor mechanisms.