Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1115932 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR29155700.fastq.gz | Fastqsanger.gz | |||
| SRR29155701.fastq.gz | Fastqsanger.gz | |||
| SRR29155702.fastq.gz | Fastqsanger.gz | |||
| SRR29155703_1.fastq.gz | Fastqsanger.gz | |||
| SRR29155703_2.fastq.gz | Fastqsanger.gz |
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Project description:Large-scale chromosomal aberrations are prevalent in human cancer but their functional effects remain poorly understood. We describe the establishment of chromosome-engineered hepatocellular carcinoma cell lines using a dual guided CRISPR-Cas9 genome editing strategy. A mega base-sized region on chromosome 8p (chr8p) was heterozygously deleted mimicking a frequently observed copy number alteration (CNA) of solid tumors. Using this unique model system, we delineated the functional consequences of chr8p loss and its impact on metastatic behavior and patient survival. We found that metastasis associated genes on chr8p act in concert to induce an aggressive and invasive phenotype characteristic for chr8p deleted tumors. The conjunction of engineered chromosome-scale CNAs and genome-wide CRISPR-Cas9 viability screenings served as a powerful tool for the discovery of novel synthetic lethal targets and vulnerabilities accompanying patient-specific copy number changes. Using this target identification strategy, we showed that chr8p deletion sensitizes tumor cells to targeting of the reactive oxygen sanitizing enzyme Nudix hydrolase 17 (NUDT17).
2023-12-27 | GSE220319 | GEO
Project description:Large-scale chromosomal aberrations are prevalent in human cancer but their functional effects remain poorly understood. We describe the establishment of chromosome-engineered hepatocellular carcinoma cell lines using a dual guided CRISPR-Cas9 genome editing strategy. A mega base-sized region on chromosome 8p (chr8p) was heterozygously deleted mimicking a frequently observed copy number alteration (CNA) of solid tumors. Using this unique model system, we delineated the functional consequences of chr8p loss and its impact on metastatic behavior and patient survival. We found that metastasis associated genes on chr8p act in concert to induce an aggressive and invasive phenotype characteristic for chr8p deleted tumors. The conjunction of engineered chromosome-scale CNAs and genome-wide CRISPR-Cas9 viability screenings served as a powerful tool for the discovery of novel synthetic lethal targets and vulnerabilities accompanying patient-specific copy number changes. Using this target identification strategy, we showed that chr8p deletion sensitizes tumor cells to targeting of the reactive oxygen sanitizing enzyme Nudix hydrolase 17 (NUDT17).
2023-12-27 | GSE220312 | GEO
Project description:Genome-scale pooled CRISPR screens are powerful tools for identifying genetic dependencies across varied cellular processes. The vast majority of CRISPR screens reported to date have focused exclusively on the perturbation of protein-coding gene function. However, protein-coding genes comprise <2% of the sequence space in the human genome leaving a substantial portion of the genome uninterrogated. Noncoding regions of the genome harbor important regulatory elements (e.g. promoters, enhancers, suppressors) that influence cellular processes but high-throughput methods for evaluating their essentiality have yet to be established. Here, we describe a CRISPR-based screening approach that facilitates the functional profiling of thousands of noncoding regulatory elements in parallel. We selected the tumor suppressor p53 as a model system and designed a pooled CRISPR library targeting thousands of p53 binding sites throughout the genome. Following transduction into dCas9-KRAB-expressing cells we identified several regulatory elements that influence cell proliferation. Moreover, we uncovered multiple elements that are required for the p53-mediated DNA damage response. Surprisingly, many of these elements are located deep within intergenic regions of the genome that have no prior functional annotations. This work diversifies the applications for pooled CRISPR screens and provides a framework for future functional studies focused on noncoding regulatory elements.
2019-08-21 | GSE126320 | GEO
Project description:CHO cells are major hosts for the industrial production of therapeutic proteins and their production stability is of considerable economic significance. It is widely known that CHO cells can rapidly acquire genetic alterations, which affects their genetic homogeneity over time. However, the role of non-genetic mechanisms, including epigenetic mechanisms such as DNA methylation, remains poorly understood. We have now used whole-genome bisulfite sequencing to establish single-base methylation maps of eight independent CHO cell lines. Our results identify CpG islands and low-methylated regions as conserved elements with dynamic DNA methylation. Interestingly, methylation patterns were found to cluster clearly along the three main branches of CHO evolution, with no directional changes over short culture periods. Furthermore, multi-ome single-cell sequencing of 9,833 nuclei from three independent cultures revealed dynamic subpopulation structures characterized by robust expression differences in pathways related to protein production. Our findings thus provide novel insights into the epigenetic heterogeneity of CHO cells and support the development of epigenetic biomarkers that trace the emergence of subpopulations in CHO cultures.
2024-11-04 | GSE273684 | GEO
Project description:CHO cells are major hosts for the industrial production of therapeutic proteins and their production stability is of considerable economic significance. It is widely known that CHO cells can rapidly acquire genetic alterations, which affects their genetic homogeneity over time. However, the role of non-genetic mechanisms, including epigenetic mechanisms such as DNA methylation, remains poorly understood. We have now used whole-genome bisulfite sequencing to establish single-base methylation maps of eight independent CHO cell lines. Our results identify CpG islands and low-methylated regions as conserved elements with dynamic DNA methylation. Interestingly, methylation patterns were found to cluster clearly along the three main branches of CHO evolution, with no directional changes over short culture periods. Furthermore, multi-ome single-cell sequencing of 9,833 nuclei from three independent cultures revealed dynamic subpopulation structures characterized by robust expression differences in pathways related to protein production. Our findings thus provide novel insights into the epigenetic heterogeneity of CHO cells and support the development of epigenetic biomarkers that trace the emergence of subpopulations in CHO cultures.
2024-11-04 | GSE273851 | GEO
Project description:CHO cells are major hosts for the industrial production of therapeutic proteins and their production stability is of considerable economic significance. It is widely known that CHO cells can rapidly acquire genetic alterations, which affects their genetic homogeneity over time. However, the role of non-genetic mechanisms, including epigenetic mechanisms such as DNA methylation, remains poorly understood. We have now used whole-genome bisulfite sequencing to establish single-base methylation maps of eight independent CHO cell lines. Our results identify CpG islands and low-methylated regions as conserved elements with dynamic DNA methylation. Interestingly, methylation patterns were found to cluster clearly along the three main branches of CHO evolution, with no directional changes over short culture periods. Furthermore, multi-ome single-cell sequencing of 9,833 nuclei from three independent cultures revealed dynamic subpopulation structures characterized by robust expression differences in pathways related to protein production. Our findings thus provide novel insights into the epigenetic heterogeneity of CHO cells and support the development of epigenetic biomarkers that trace the emergence of subpopulations in CHO cultures.
2024-11-04 | GSE272869 | GEO
Project description:Essentiality assays are commonly practiced as important tools for the discovery of gene functions. Growth/no growth screens of single gene knockout strain collections are often utilized to test the predictive power of genome-scale models. False positive predictions occur when computational analysis predicts a gene to be non-essential, however experimental screens deem the gene to be essential. One explanation for this inconsistency is that the model contains the wrong information, possibly an incorrectly annotated alternative pathway or isozyme reaction. Inconsistencies could also be attributed to experimental limitations, such as growth tests with arbitrary time cut-offs. The focus of this study was to resolve such inconsistencies to better understand isozyme activities and gene essentiality. Gene-deletion strains associated with false positive predictions of gene essentiality on defined minimal medium for Escherichia coli were targeted for extended growth tests followed by population sequencing.
2018-07-19 | GSE117303 | GEO
Project description:Small-scale bioreactors used in Chinese hamster ovary (CHO) cell line development allow transcriptomic studies on multiple cell lines. Here we define the CHO cell long non-coding RNA (lncRNA) transcriptome from cells grown in controlled miniature bioreactors under fed-batch conditions using RNA-Seq to identify lncRNAs and how the expression of these changes throughout growth and between IgG producers. We identify lncRNAs associated with productivity and growth characteristics, finding that Adapt15, linked to ER stress, GAS5, linked to mTOR signalling/growth arrest, and PVT1, linked to Myc expression, are differentially regulated during fed-batch culture and whose expression relates to productivity and growth. Changes in (non)-coding RNA expression between the seed train and the equivalent day of fed-batch culture are also reported, showing large differences in gene expression between these, and compared with existing datasets. Collectively, we present a comprehensive lncRNA CHO cell profiling and identify targets for engineering growth and productivity characteristics of CHO cells.
2019-11-20 | GSE140669 | GEO
Project description:Small-scale bioreactors used in Chinese hamster ovary (CHO) cell line development allow transcriptomic studies on multiple cell lines. Here we define the CHO cell long non-coding RNA (lncRNA) transcriptome from cells grown in controlled miniature bioreactors under fed-batch conditions using RNA-Seq to identify lncRNAs and how the expression of these changes throughout growth and between IgG producers. We identify lncRNAs associated with productivity and growth characteristics, finding that Adapt15, linked to ER stress, GAS5, linked to mTOR signalling/growth arrest, and PVT1, linked to Myc expression, are differentially regulated during fed-batch culture and whose expression relates to productivity and growth. Changes in (non)-coding RNA expression between the seed train and the equivalent day of fed-batch culture are also reported, showing large differences in gene expression between these, and compared with existing datasets. Collectively, we present a comprehensive lncRNA CHO cell profiling and identify targets for engineering growth and productivity characteristics of CHO cells.
2019-11-20 | GSE140668 | GEO
Project description:<p><strong>Background:</strong> Cell line specific, genome-scale metabolic models enable rigorous and systematic in silico investigation of cellular metabolism. Such models have recently become available for Chinese hamster ovary (CHO) too. However, a key ingredient, namely an experimentally validated biomass function that summarizes the cellular composition, was so far missing in all genome-scale metabolic models of CHO cell lines. Here, we close this gap by providing extensive experimental data on the biomass composition of 13 parental and producer CHO cell lines under various conditions.</p><p><strong>Results:</strong> We report protein, lipid, DNA, RNA and carbohydrate content, cell dry mass, and protein and lipid composition. Furthermore, we present meticulous data on exchange rates between cells and environment and provide detailed experimental protocols on how to determine all of the above. The biomass composition is converted into cell line and condition specific biomass functions for usage in cell line specific, genome-scale metabolic models of CHO. Finally, flux-balance analysis (FBA) is used to demonstrate consistency between in silico predictions and experimental analysis.</p><p><strong>Conclusions:</strong> Our study reveals a strong variability of the total protein content and cell dry mass across cell lines. Yet, the relative amino acid composition is independent of the cell lines and conditions and thus needs not be explicitly measured for each new cell line. In contrast, the lipid composition is strongly influenced by the growth media and needs to be determined for each case. These cell line specific variations in the biomass composition have a small impact on growth rate predictions with FBA, as inaccuracies in the predictions are typically dominated by inaccuracies in the exchange rate spectra. Cell specific biomass variations become only important if the experimental errors in the exchange rate spectra drop below twenty percent.</p><p><br></p><p><strong>Links:</strong></p><p>All data and code for data processing are available at <a href='http://dx.doi.org/10.17632/g3wz7spyc9.1' rel='noopener noreferrer' target='_blank'>Data Mendeley</a>.</p><p>Cell line specific metabolic models were deposited in <a href='https://www.ebi.ac.uk/biomodels/(Chelliah et al, 2014)' rel='noopener noreferrer' target='_blank'>BioModels</a>, identifiers are listed in the study sample table.</p>
2020-07-13 | MTBLS1210 | MetaboLights