Cysteine Rich Protein 2 is a copper-responsive regulator of skeletal muscle differentiation. [CUT&Run]
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE252156 | GEO | 2024/05/30
REPOSITORIES: GEO
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Project description:Copper (Cu) is an essential trace element for diverse biological reactions such as respiration, neurotransmitter synthesis, oxidative stress and transcriptional regulation. If Cu homeostasis is disrupted, several pathological conditions can develop, leading to alterations of neuronal, cognitive, and muscular development. Known Cu+-binding transcription factors in mammalian cells are Atox1, Mtf1 and Sp1. We identified Crip2 as a novel Cu+-responsive transcriptional regulator that is required for the differentiation of primary myoblasts derived from mouse satellite cells. Functional characterization of CRISPR/Cas9-mediated deletion of Crip2 showed that myoblasts fail to differentiate and manifest a decrease in expression of the differentiation markers Myogenin and Myosin heavy chain. RNA-seq and CUT&RUN analysis were performed to understand the effect on Crip2 in the regulation of gene expression in proliferating and differentiating primary myoblasts stimulated with Cu.
2024-05-30 | GSE252158 | GEO
Project description:MTF1 is a highly conserved metal-binding transcription factor in eukaryotes. MTF1 binds to DNA sequence motifs, termed metal response elements (MREs) to induce the expression of genes involved in metal and oxidative stress homeostasis. MTF1 is responsive to both metal excess and deprivation, and can also protect cells from oxidative and hypoxic stresses. Disruption of metal homeostasis leads to the development of several pathological states. Despite its roles in these processes , MTF1 has been shown to be required for developmental processes such as embryonic liver formation. In this study, we used multiple strategies to understand the mechanism by which MTF1 functions in skeletal muscle differentiation and to determine the role cellular copper (Cu) status plays in this process. We provide the functional relationships between MTF1, Cu, myogenic gene promoters, and MyoD, an specific component of the myogenic transcriptional machinery in differentiating primary myoblasts derived from mouse satellite cells. We found that MTF1 is induced and translocated to the nucleus upon initiation of myogenesis. Consistent with previous studies from our laboratory , addition of non-toxic concentrations of Cu promote myogenesis and enhanced MTF1 expression. CRISPR/Cas9-mediated depletion of MTF1 demonstrated that MTF1 is essential for proper development of skeletal muscle, as partial Mtf1 knockdown leads to apoptosis to differentiating myoblasts. MTF1 was also found to bind Cu at a carboxy-terminal tetra-cysteine cluster, which may contribute to the mobilization of Cu to the nucleus during myogenesis. ChIP-seq and ChIP-qPCR analyses showed that MTF1 binds at the promoter regions of myogenic genes as part of a complex with MyoD, the master transcriptional regulator of the myogenic lineage. These results have the potential to initiate a new area of research in MTF1 function and in the regulation of myogenesis. Furthermore, these studies set the basis to understand the role of Cu at the transcriptional level, affects growth and development and will contribute to the largely unexplored are of muscular phenotypes observed in human pathologies associated to Cu misbalance, such as Menkes’ and Wilson’s diseases.
2019-10-04 | GSE116331 | GEO
Project description:Copper (Cu)-binding transcription factors are emerging as important regulators of gene expression during skeletal muscle differentiation. Our group has shown that the classic Metal Transcription Factor 1, MTF1, works cooperatively with the myogenic factor MyoD to promote skeletal muscle differentiation and ensure viability of murine primary myoblasts. Preliminary evidence showed that MTF1 interacts with members of the mammalian SWItch/Sucrose Non-Fermentable (mSWI/SNF) family of ATP-dependent chromatin remodeling enzymes to promote differential gene expression in proliferating and differentiating myoblasts. mSWI/SNF complexes are assembled into three major subfamilies that are distinguished by the presence of mutually exclusive subunits: cBAF (canonical BRG1 or BRM-Associated Factor), PBAF (Polybromo containing BAF), or ncBAF (non-canonical BAF). The mechanisms by which each subfamily contributes to the establishment or function of specific cell lineages are poorly understood. We determined the contributions of the BAF, ncBAF, and PBAF complexes to myoblast proliferation, differentiation, and metal homeostasis via knock-down (KD) of a distinguishing subunit of each complex (Baf250A, Brd9, and Baf180, respectively). Biochemical, molecular and RNA-seq analyses showed that Baf250A KD reduced myoblast proliferation rate and differentiation. Brd9 KD resulted in a minor proliferation defect and a delay in myogenesis. KD of Baf180 and other PBAF-specific subunits did not impact proliferation or differentiation but affected the maintenance of metal homeostasis in myoblasts. The data support a model for differential roles and interactions between MTF1 and a novel category of Cu-binding transcription factors and the SWI/SNF enzymes in the development and homeostasis of the skeletal muscle lineage.
2024-10-26 | GSE253376 | GEO
Project description:Copper (Cu)-binding transcription factors are emerging as important regulators of gene expression during skeletal muscle differentiation. Our group has shown that the classic Metal Transcription Factor 1, MTF1, works cooperatively with the myogenic factor MyoD to promote skeletal muscle differentiation and ensure viability of murine primary myoblasts. Preliminary evidence showed that MTF1 interacts with members of the mammalian SWItch/Sucrose Non-Fermentable (mSWI/SNF) family of ATP-dependent chromatin remodeling enzymes to promote differential gene expression in proliferating and differentiating myoblasts. mSWI/SNF complexes are assembled into three major subfamilies that are distinguished by the presence of mutually exclusive subunits: cBAF (canonical BRG1 or BRM-Associated Factor), PBAF (Polybromo containing BAF), or ncBAF (non-canonical BAF). The mechanisms by which each subfamily contributes to the establishment or function of specific cell lineages are poorly understood. We determined the contributions of the BAF, ncBAF, and PBAF complexes to myoblast proliferation, differentiation, and metal homeostasis via knock-down (KD) of a distinguishing subunit of each complex (Baf250A, Brd9, and Baf180, respectively). Biochemical, molecular and RNA-seq analyses showed that Baf250A KD reduced myoblast proliferation rate and differentiation. Brd9 KD resulted in a minor proliferation defect and a delay in myogenesis. KD of Baf180 and other PBAF-specific subunits did not impact proliferation or differentiation but affected the maintenance of metal homeostasis in myoblasts. The data support a model for differential roles and interactions between MTF1 and a novel category of Cu-binding transcription factors and the SWI/SNF enzymes in the development and homeostasis of the skeletal muscle lineage.
2024-10-26 | GSE253377 | GEO
Project description:The transition metal copper (Cu) is an essential element for all living organisms. In plants, Cu plays key roles in electron transport chains of chloroplasts and mitochondria, as well as in cell wall metabolism, ethylene perception, molybdenum cofactor synthesis and oxidative stress protection. Because of its physiological importance, suboptimal concentrations of Cu trigger a re-organization of metabolism to economize on Cu, and pronounced Cu deficiency causes severe growth reduction and defects in male fertility. However, when present in excess, Cu can be highly toxic. Therefore, Cu uptake, utilization and cellular concentrations are strictly regulated. A number of components of the Cu-homeostatic network of Arabidopsis have already been identified. However, the mechanisms that control responses of plant gene expression to Cu-deficiency are only partly understood. In Chlamydomonas reinhardtii, the transcription factor Crr1 is required for activating and/or repressing the expression of a number of genes in response to Cu deficiency. This protein contains a plant-specific DNA-binding domain (SBP domain), ankyrin repeats and a C-terminal cysteine-rich region with similarity to a Drosophila metallothionein (MT). In Arabidopsis, there is a family of 16 proteins with SBP domains named SPL family (SBP-like) of which a subset of proteins have been implicated in flowering time control and floral development. Among all of them, AtSPL7 is the most similar to Crr1 (27 % identity), and AtSPL1 (25 % identity), AtSPL12 (24 % identity) and AtSPL14 (23 % identity) share a common protein architecture. The goal of this work was to obtain a complete account of the response of the Arabidopsis thaliana transcriptome to Cu deficiency, as well as to determine the role of AtSPL7 in the transcriptional response to Cu deficiency. For this purpose, three independent experiments were carried out including Col-0 wild-type and spl7-2 mutant plants grown in Cu-sufficient and Cu-deficient hydroponic media, respectively. Root and shoot transcriptomes were established by RNA-Seq for quantitative comparisons.
2013-10-14 | GSE24696 | GEO
Project description:The transition metal copper (Cu) is an essential element for all living organisms. In plants, Cu plays key roles in electron transport chains of chloroplasts and mitochondria, as well as in cell wall metabolism, ethylene perception, molybdenum cofactor synthesis and oxidative stress protection. Because of its physiological importance, suboptimal concentrations of Cu trigger a re-organization of metabolism to economize on Cu, and pronounced Cu deficiency causes severe growth reduction and defects in male fertility. However, when present in excess, Cu can be highly toxic. Therefore, Cu uptake, utilization and cellular concentrations are strictly regulated. A number of components of the Cu-homeostatic network of Arabidopsis have already been identified. However, the mechanisms that control responses of plant gene expression to Cu-deficiency are only partly understood. In Chlamydomonas reinhardtii, the transcription factor Crr1 is required for activating and/or repressing the expression of a number of genes in response to Cu deficiency. This protein contains a plant-specific DNA-binding domain (SBP domain), ankyrin repeats and a C-terminal cysteine-rich region with similarity to a Drosophila metallothionein (MT). In Arabidopsis, there is a family of 16 proteins with SBP domains named SPL family (SBP-like) of which a subset of proteins have been implicated in flowering time control and floral development. Among all of them, AtSPL7 is the most similar to Crr1 (27 % identity), and AtSPL1 (25 % identity), AtSPL12 (24 % identity) and AtSPL14 (23 % identity) share a common protein architecture. The goal of this work was to obtain a complete account of the response of the Arabidopsis thaliana transcriptome to Cu deficiency, as well as to determine the role of AtSPL7 in the transcriptional response to Cu deficiency. For this purpose, three independent experiments were carried out including Col-0 wild-type and spl7-2 mutant plants grown in Cu-sufficient and Cu-deficient hydroponic media, respectively. Root and shoot transcriptomes were established by RNA-Seq for quantitative comparisons. Sampling of root and shoot tissues of wild-type (WT, Col-0) and spl7-2 mutant plants (the mutant is knock-down for the transcription factor SPL7, which plays a key role in the transcriptional regulation of the copper deficiency responses) cultivated hydroponically in a modified Hoagland's solution under Cu-sufficient (control) and Cu-deficient conditions.
2013-10-14 | E-GEOD-24696 | biostudies-arrayexpress
Project description:Anthropogenic pollution has increased the levels of heavy metals in the environment. Bacterial populations continue to thrive in highly polluted environments and bacteria must have mechanisms to counter heavy metal stress. We chose to examine the response of the environmentally-relevant organism Pseudomonas aeruginosa to two different copper treatments. A short, 45 min exposure to copper was done in the Cu shock treatment to examine the immediate transcriptional profile to Cu stress. The Cu adapted treatment was designed to view the transcriptional profile of cells that were actively growing in the presence of Cu. Experiment Overall Design: We analyzed 2 biological replicates of Pseudomonas aeruginosa exposed to a 45 min Cu shock as compared to a control that was exposed to HCl to bring the pH to similar levels. We analyzed 2 biological replicates of Pseudomonas aeruginosa that were grown in the presence of Cu for approx. 6h (Cu adapted) as compared to an untreated control.
2007-10-24 | E-GEOD-4152 | biostudies-arrayexpress
Project description:Copper (Cu) regulates hypoxia-inducible factor-1 (HIF-1) transcription activity by affecting the selectivity of HIF-1α targeting to the promoters of the affected genes. Here, we made an effort to provide a comprehensive understanding of Cu regulation of the selectivity of HIF-1α targeting across genome. We used tetraethylenepentamine (TEPA), a Cu selective chelator, to reduce Cu content in the cells. In hypoxia, we conducted chromatin immunoprecipitation combined with massively parallel DNA sequencing (ChIP-seq) to globally map the binding sites of HIF-1α, Pol Ⅱ (RNA polymeraseⅡ) and histone H3K27ac. We also performed RNA-sequencing (RNA-seq) in EA.hy926 cells under hypoxia (1% O2) with or without Cu depression to determine the profile of mRNA expression. Our analyses identified 3197 HIF-1α binding sites under hypoxia. Cu depression by TEPA reduced 1820 binding sites from the 3197, but induced additional 274 new binding sites. We analyzed these binding sites in the promoter and putative enhancer regions, coupled with their mRNA expression profiles, and found 281 Cu-dependent and 10 Cu-independent HIF-1α target genes. We found that the core bases “GGAA” and “TTCC” constituted the critical motifs for the binding sites of Cu-dependent genes. This study thus revealed that Cu, by affecting the binding of HIF-1α to the critical motifs in the promoter and putative enhancer regions of HIF-1 regulated genes, leads to the selectivity of HIF-1 regulated expression of Cu-dependent genes.
2019-09-24 | GSE120524 | GEO
Project description:Copper (Cu) regulates hypoxia-inducible factor-1 (HIF-1) transcription activity by affecting the selectivity of HIF-1α targeting to the promoters of the affected genes. Here, we made an effort to provide a comprehensive understanding of Cu regulation of the selectivity of HIF-1α targeting across genome. We used tetraethylenepentamine (TEPA), a Cu selective chelator, to reduce Cu content in the cells. In hypoxia, we conducted chromatin immunoprecipitation combined with massively parallel DNA sequencing (ChIP-seq) to globally map the binding sites of HIF-1α, Pol Ⅱ (RNA polymeraseⅡ) and histone H3K27ac. We also performed RNA-sequencing (RNA-seq) in EA.hy926 cells under hypoxia (1% O2) with or without Cu depression to determine the profile of mRNA expression. Our analyses identified 3197 HIF-1α binding sites under hypoxia. Cu depression by TEPA reduced 1820 binding sites from the 3197, but induced additional 274 new binding sites. We analyzed these binding sites in the promoter and putative enhancer regions, coupled with their mRNA expression profiles, and found 281 Cu-dependent and 10 Cu-independent HIF-1α target genes. We found that the core bases “GGAA” and “TTCC” constituted the critical motifs for the binding sites of Cu-dependent genes. This study thus revealed that Cu, by affecting the binding of HIF-1α to the critical motifs in the promoter and putative enhancer regions of HIF-1 regulated genes, leads to the selectivity of HIF-1 regulated expression of Cu-dependent genes.
2019-09-24 | GSE120523 | GEO
Project description:Anthropogenic pollution has increased the levels of heavy metals in the environment. Bacterial populations continue to thrive in highly polluted environments and bacteria must have mechanisms to counter heavy metal stress. We chose to examine the response of the environmentally-relevant organism Pseudomonas aeruginosa to two different copper treatments. A short, 45 min exposure to copper was done in the Cu shock treatment to examine the immediate transcriptional profile to Cu stress. The Cu adapted treatment was designed to view the transcriptional profile of cells that were actively growing in the presence of Cu. Keywords: stress response
2006-10-04 | GSE4152 | GEO