Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Identifying gene combinations for targeting innate immune cells to enhance T cells activation [CITE-seq]

ABSTRACT: Genetic engineering of hematopoietic cells ex-vivo opens new opportunities for programming immune cells and improving their functionality in response to illness and infections. Molecular methods for genetic manipulation are constantly improving, but it is challenging to reveal which genes are the most beneficial to target and whether multiple genes tackling can further improve the desired outcome. Current screening methods are limited by scale, the number of perturbations, and low content readouts. We developed a hybrid platform called “high multiplicity of perturbations and cellular indexing of transcriptomes and epitopes by sequencing (HMPCITE-seq)” that allows for finding combinations of genes to target and improves desired cellular functions. We used antigen-presenting cells and performed two genome-wide CRISPR screens to enhance the activation and co-stimulation programs and restrict the suppressive programs. Five genes were found in both screens including Cebpb gene. We showed that targeting Cebpb in antigen-presenting cells enhances the expression of costimulatory molecules, prevents attenuate cell migration and phagocytosis, improves T cell proliferation, and restricts tumor growth. To find the genetic interactions between validated targeted genes, and identify gene combinations for targeting, we performed high-order perturbations of validated genes from the screens with single-cell RNA-seq readouts. We found that targeting both Cebpb and Med12 genes shows a better phenotype compared to the single KOs. Thus, the HMPCITE-seq platform, starting from a genome-wide screen, allows the finding of gene combinations to target as demonstrated by functional experiments in-vitro and in-vivo.

ORGANISM(S): Mus musculus

PROVIDER: GSE243244 | GEO | 2023/09/17

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Similar Datasets

Identifying gene combinations for targeting innate immune cells to enhance T cells activation [Cebpb-KO CRISPR screen]

Project description:Genetic engineering of hematopoietic cells ex-vivo opens new opportunities for programming immune cells and improving their functionality in response to illness and infections. Molecular methods for genetic manipulation are constantly improving, but it is challenging to reveal which genes are the most beneficial to target and whether multiple genes tackling can further improve the desired outcome. Current screening methods are limited by scale, the number of perturbations, and low content readouts. We developed a hybrid platform called “high multiplicity of perturbations and cellular indexing of transcriptomes and epitopes by sequencing (HMPCITE-seq)” that allows for finding combinations of genes to target and improves desired cellular functions. We used antigen-presenting cells and performed two genome-wide CRISPR screens to enhance the activation and co-stimulation programs and restrict the suppressive programs. Five genes were found in both screens including Cebpb gene. We showed that targeting Cebpb in antigen-presenting cells enhances the expression of costimulatory molecules, does not attenuate cell migration and phagocytosis, improves T cell proliferation, and restricts tumor growth. To find the genetic interactions between validated targeted genes, and identify gene combinations for targeting, we performed high-order perturbations of validated genes from the screens with single-cell RNA-seq readouts. We found that targeting both Cebpb and Med12 genes shows a better phenotype compared to the single KOs. Thus, the HMPCITE-seq platform, starting from a genome-wide screen, allows the finding of gene combinations to target as demonstrated by functional experiments in-vitro and in-vivo.

2023-09-13 | GSE232543 | GEO

Identifying gene combinations for targeting innate immune cells to enhance T cells activation [Med12 and Cebpb KO]

Project description:Genetic engineering of hematopoietic cells ex-vivo opens new opportunities for programming immune cells and improving their functionality in response to illness and infections. Molecular methods for genetic manipulation are constantly improving, but it is challenging to reveal which genes are the most beneficial to target and whether multiple genes tackling can further improve the desired outcome. Current screening methods are limited by scale, the number of perturbations, and low content readouts. We developed a hybrid platform called “high multiplicity of perturbations and cellular indexing of transcriptomes and epitopes by sequencing (HMPCITE-seq)” that allows for finding combinations of genes to target and improves desired cellular functions. We used antigen-presenting cells and performed two genome-wide CRISPR screens to enhance the activation and co-stimulation programs and restrict the suppressive programs. Five genes were found in both screens including Cebpb gene. We showed that targeting Cebpb in antigen-presenting cells enhances the expression of costimulatory molecules, prevents attenuate cell migration and phagocytosis, improves T cell proliferation, and restricts tumor growth. To find the genetic interactions between validated targeted genes, and identify gene combinations for targeting, we performed high-order perturbations of validated genes from the screens with single-cell RNA-seq readouts. We found that targeting both Cebpb and Med12 genes shows a better phenotype compared to the single KOs. Thus, the HMPCITE-seq platform, starting from a genome-wide screen, allows the finding of gene combinations to target as demonstrated by functional experiments in-vitro and in-vivo.

2023-09-13 | GSE233796 | GEO

Identifying gene combinations for targeting innate immune cells to enhance T cells activation [RNA-Seq]

Project description:Genetic engineering of hematopoietic cells ex-vivo opens new opportunities for programming immune cells and improving their functionality in response to illness and infections. Molecular methods for genetic manipulation are constantly improving, but it is challenging to reveal which genes are the most beneficial to target and whether multiple genes tackling can further improve the desired outcome. Current screening methods are limited by scale, the number of perturbations, and low content readouts. We developed a hybrid platform called “high multiplicity of perturbations and cellular indexing of transcriptomes and epitopes by sequencing (HMPCITE-seq)” that allows for finding combinations of genes to target and improves desired cellular functions. We used antigen-presenting cells and performed two genome-wide CRISPR screens to enhance the activation and co-stimulation programs and restrict the suppressive programs. Five genes were found in both screens including Cebpb gene. We showed that targeting Cebpb in antigen-presenting cells enhances the expression of costimulatory molecules, prevents attenuate cell migration and phagocytosis, improves T cell proliferation, and restricts tumor growth. To find the genetic interactions between validated targeted genes, and identify gene combinations for targeting, we performed high-order perturbations of validated genes from the screens with single-cell RNA-seq readouts. We found that targeting both Cebpb and Med12 genes shows a better phenotype compared to the single KOs. Thus, the HMPCITE-seq platform, starting from a genome-wide screen, allows the finding of gene combinations to target as demonstrated by functional experiments in-vitro and in-vivo.

2023-09-13 | GSE211213 | GEO

Identifying gene combinations for targeting innate immune cells to enhance T cells activation [DNA]

2023-09-13 | GSE206832 | GEO

Compressed phenotypic screening empowers scalable biological discovery (PDAC screens)

Project description:High-throughput phenotypic screens leveraging biochemical perturbations and high-content readouts are poised to advance therapeutic discovery, yet they remain constrained by limitations of scale. To address this, we establish a method of pooling exogenous perturbations followed by computational deconvolution to compress a screen’s required sample, labor, and financial input. We benchmark the approach with a bioactive small molecule library and a high-content imaging readout, demonstrating the feasibility and increased efficiency of compressed experimental designs compared to conventional approaches. To prove generalizability, we apply compressed screening in two different biological discovery campaigns. In the first, we use early-passage pancreatic cancer organoids to map transcriptional responses to alibrary oftumor-microenvironmentrecombinant protein ligands. We uncover reproducible phenotypic shifts induced by specific ligands that are distinct from canonical reference signatures and uniquely correlate with clinical outcome.In the second, we examine the modulatory effects of a known mechanism of action chemical compound library on primary human peripheral blood mononuclear cell immune responses. Through contrastive analyses, we identify molecules that potentiate and/or inhibit cell-type specific transcriptional features, uncover pleiotropic effects for individual compounds across diverse cell types, and realize a systems-level view of drug responses. In sum, our approach empowers phenotypic screens with information-rich readouts to advance drug discovery efforts as well as basic biological inquiry.

2024-05-13 | GSE267243 | GEO

Efficient combinatorial targeting of RNA transcripts in single cells with Cas13 RNA Perturb-seq

Project description:Pooled CRISPR screens coupled with single-cell RNA-sequencing have enabled systematic interrogation of gene function and regulatory networks. Here, we introduce Cas13 RNA Perturb-seq (CaRPool-seq) which leverages the RNA-targeting CRISPR/Cas13d system and enables efficient combinatorial perturbations alongside multimodal single-cell profiling. CaRPool-seq encodes multiple perturbations on a cleavable array which is associated with a detectable barcode sequence, allowing for the simultaneous targeting of multiple genes. We compared CaRPool-seq to existing Cas9-based methods, highlighting its unique strength to efficiently profile combinatorially perturbed cells. Finally, we apply CaRPool-seq to perform multiplexed combinatorial perturbations of myeloid differentiation regulators in an acute myeloid leukemia (AML) model system and identify extensive interactions between different chromatin regulators that can enhance or suppress AML differentiation phenotypes.

2022-09-29 | GSE213957 | GEO

Engineered CRISPR-Cas12a for higher-order combinatorial chromatin perturbations (amplicon)

Project description:Multiplexed genetic perturbations are critical for testing functional interactions among coding or non-coding genetic elements. Compared to double-stranded DNA cutting, repressive chromatin formation using CRISPR interference (CRISPRi) avoids genotoxicity and is more effective for perturbing non-coding regulatory elements in pooled assays. However, current CRISPRi pooled screening approaches are limited to targeting 1-3 genomic sites per cell. We engineer an Acidaminococcus Cas12a (AsCas12a) variant, multiplexed transcriptional interference AsCas12a (multiAsCas12a), that incorporates R1226A, a mutation that stabilizes the ribonucleoprotein:DNA complex via DNA nicking. The multiAsCas12a-KRAB fusion improves CRISPRi activity over DNase-dead AsCas12a-KRAB fusions, often rescuing the activities of lentivirally delivered CRISPR RNAs (crRNA) that are inactive when used with the latter. multiAsCas12a-KRAB supports CRISPRi using 6-plex crRNA arrays in high-throughput pooled screens. Using multiAsCas12a-KRAB, we discover enhancer elements and dissect the combinatorial function of cis-regulatory elements in human cells. These results instantiate a group testing framework for efficiently surveying numerous combinations of chromatin perturbations for biological discovery and engineering.

2024-03-05 | GSE260830 | GEO

Engineered CRISPR-Cas12a for higher-order combinatorial chromatin perturbations (CRISPR screen)

2024-03-06 | GSE260829 | GEO

Engineered CRISPR-Cas12a for higher-order combinatorial chromatin perturbations (RNA-Seq)

2024-03-06 | GSE260828 | GEO

Engineered CRISPR-Cas12a for higher-order combinatorial chromatin perturbations (Nanopore)

2024-03-06 | GSE260827 | GEO