Similar Datasets
Project description:We previously performed a tiling CRISPR activation (CRISPRa) screen in Jurkat T cells to discover enhancers controlling IL2RA expression. This screen identified six regions where recruitment of dCas9-VP64 was sufficient to drive increased expression of IL2RA. We named these putative enhancers CRISPRa Responsive Elements (CaREs). To examine transcriptome-wide consequences of dCas9-VP64 recruitment to IL2RA CaREs, we performed RNA-Seq on HuT78 cells stably expressing dCas9-VP64 and gRNAs targeting the IL2RA TSS, CaRE3, or CaRE4, or stimulated with anti-CD3/CD28 antibodies.
2017-04-26 | GSE98177 | GEO
Project description:Background: Dystrophic epidermolysis bullosa (DEB) is a skin blistering disease caused by mutations in COL7A1, which encodes type VII collagen (C7). There is no cure for DEB, but previous work has shown potential therapeutic benefit in increasing production of even partially functional C7. Genome-wide screens using CRISPR-Cas9 have enabled the identification of genes involved in cancer development, drug resistance, and other genetic diseases, suggesting that they could be used to identify novel drivers of C7 production. Methods: A keratinocyte C7 reporter cell line was created by integrating a tdTomato fluorescent marker into the last exon of the endogenous COL7A1 gene. A genome-wide CRISPR activation (CRISPRa) screen was performed with the C7_tdTomato reporter to identify genes and pathways that increase C7 expression. High tdTomato-expressing cells were sorted and sequenced to identify the single guide RNAs (sgRNAs) that became enriched relative to the starting material. Pathway analysis was performed on the corresponding genes to identify regulators and pathways that influence C7 expression. A targeted drug screen was performed in three different keratinocyte cell lines based on the CRISPRa screen results, and C7 upregulation was evaluated. Results: The C7_tdTomato cell line was validated as an effective reporter cell line for detection of C7 upregulation. The CRISPRa screen identified two genes, DENND4B and TYROBP as top hits based on enrichment in the high fluorescence population and representation with multiple sgRNAs. Pathway analysis of the CRISPRa screen showed enrichment of toll-like receptor and interferon-related upstream regulators and functions related to calcium uptake and immune signaling. Several compounds in the targeted drug screen increased C7 expression in at least one keratinocyte line, but only kaempferol, a plant flavonoid, significantly increased C7 mRNA and protein in a DEB patient line. Conclusions: The novel C7_tdTomato reporter cell line can be used to screen compounds for increased C7 expression. The CRISPRa screen combined with a fluorescent reporter cell line can be used to reveal mechanistic regulators of gene expression and therapeutic targets for rare genetic diseases. Kaempferol has shown promising results in increasing C7 production and should be further evaluated as a potential therapeutic for DEB patients.
2022-01-31 | GSE163024 | GEO
Project description:CRISPRa screen for drivers of COL7A1 expression in keratinocytes
| PRJNA684343 | ENA
Project description:Functional validation of the genome-scale CRISPR-activation screen by transcriptomic profiling of MGT#4-reporter expressing IDH-wt human glioma initiating cells (hGICs) upon CRISPRa-mediated overexpression of the predicted pro-mesenchymal drivers.
2023-11-27 | GSE236153 | GEO
Project description:Sodium-lithium countertransport activity (SLC) is an intermediate phenotype for salt sensitive hypertension; elevated SLC corresponds to increased risk of hypertension. Genetic epidemiology studies indicate that SLC is heritable in humans and nonhuman primates; however, little is known about genetic determinants explaining variation in SLC. In this study we identified dietary salt-responsive gene networks in baboon kidneys and salt-responsive networks that differ between baboons with normal and high blood pressure phenotypes.
2018-12-31 | GSE18484 | GEO
Project description:Regulation of cytokine production in stimulated T cells can be disrupted in autoimmunity, immunodeficiencies, and cancer. Systematic discovery of stimulation-dependent cytokine regulators requires both loss-of-function and gain-of-function studies, which have been challenging in primary human cells. We now report genome-wide CRISPR activation (CRISPRa) and interference (CRISPRi) screens in primary human T cells to identify gene networks controlling interleukin 2 and interferon gamma production. Arrayed CRISPRa confirmed key hits and enabled multiplexed secretome characterization, revealing reshaped cytokine responses. Coupling CRISPRa screening with single-cell RNA-seq enabled deep molecular characterization of screen hits, revealing how perturbations tuned T cell activation and promoted cell states characterized by distinct cytokine expression profiles. Together, these screens reveal genes that reprogram critical immune cell functions, which could inform the design of immunotherapies.
2021-05-13 | GSE174284 | GEO
Project description:Regulation of cytokine production in stimulated T cells can be disrupted in autoimmunity, immunodeficiencies, and cancer. Systematic discovery of stimulation-dependent cytokine regulators requires both loss-of-function and gain-of-function studies, which have been challenging in primary human cells. We now report genome-wide CRISPR activation (CRISPRa) and interference (CRISPRi) screens in primary human T cells to identify gene networks controlling interleukin 2 and interferon gamma production. Arrayed CRISPRa confirmed key hits and enabled multiplexed secretome characterization, revealing reshaped cytokine responses. Coupling CRISPRa screening with single-cell RNA-seq enabled deep molecular characterization of screen hits, revealing how perturbations tuned T cell activation and promoted cell states characterized by distinct cytokine expression profiles. Together, these screens reveal genes that reprogram critical immune cell functions, which could inform the design of immunotherapies.
2021-05-13 | GSE174255 | GEO
Project description:Regulation of cytokine production in stimulated T cells can be disrupted in autoimmunity, immunodeficiencies, and cancer. Systematic discovery of stimulation-dependent cytokine regulators requires both loss-of-function and gain-of-function studies, which have been challenging in primary human cells. We now report genome-wide CRISPR activation (CRISPRa) and interference (CRISPRi) screens in primary human T cells to identify gene networks controlling interleukin 2 and interferon gamma production. Arrayed CRISPRa confirmed key hits and enabled multiplexed secretome characterization, revealing reshaped cytokine responses. Coupling CRISPRa screening with single-cell RNA-seq enabled deep molecular characterization of screen hits, revealing how perturbations tuned T cell activation and promoted cell states characterized by distinct cytokine expression profiles. Together, these screens reveal genes that reprogram critical immune cell functions, which could inform the design of immunotherapies.
2022-02-04 | GSE190846 | GEO
Project description:Regulation of cytokine production in stimulated T cells can be disrupted in autoimmunity, immunodeficiencies, and cancer. Systematic discovery of stimulation-dependent cytokine regulators requires both loss-of-function and gain-of-function studies, which have been challenging in primary human cells. We now report genome-wide CRISPR activation (CRISPRa) and interference (CRISPRi) screens in primary human T cells to identify gene networks controlling interleukin 2 and interferon gamma production. Arrayed CRISPRa confirmed key hits and enabled multiplexed secretome characterization, revealing reshaped cytokine responses. Coupling CRISPRa screening with single-cell RNA-seq enabled deep molecular characterization of screen hits, revealing how perturbations tuned T cell activation and promoted cell states characterized by distinct cytokine expression profiles. Together, these screens reveal genes that reprogram critical immune cell functions, which could inform the design of immunotherapies.
2022-02-04 | GSE190604 | GEO