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Project description:T cell exhaustion limits antitumor immunity, but the molecular determinants of this process remain poorly understood. Using a chronic stimulation assay, we performed genome-wide CRISPR-Cas9 screens to systematically discover regulators of T cell exhaustion, which identified an enrichment of epigenetic factors. In vivo CRISPR screens in murine and human tumor models demonstrated that perturbation of the INO80 and BAF chromatin remodeling complexes improved T cell persistence in tumors. In vivo Perturb-seq revealed distinct transcriptional roles of each complex and that depletion of canonical BAF complex members, including Arid1a, resulted in the maintenance of an effector program and downregulation of exhaustion-related genes in tumor-infiltrating T cells. Finally, Arid1a depletion limited the acquisition of exhaustion-associated chromatin accessibility and led to improved antitumor immunity. In summary, we provide an atlas of the genetic regulators of T cell exhaustion and demonstrate that modulation of epigenetic state can improve T cell responses in cancer immunotherapy.
| S-EPMC9949532 | biostudies-literature
Project description:T cell exhaustion limits anti-tumor immunity, but the molecular determinants of this process remain poorly understood. Using a chronic antigen stimulation assay, we performed genome-wide CRISPR/Cas9 screens to systematically discover genetic regulators of T cell exhaustion, which identified an enrichment of epigenetic factors. In vivo CRISPR screens in murine and human tumor models demonstrated that perturbation of several epigenetic regulators, including members of the INO80 and BAF chromatin remodeling complexes, improved T cell persistence in tumors. In vivo paired CRISPR perturbation and single-cell RNA sequencing revealed distinct transcriptional roles of each complex and that depletion of canonical BAF complex members, including Arid1a, resulted in the maintenance of an effector program and downregulation of terminal exhaustion-related genes in tumor-infiltrating T cells. Finally, Arid1a-depletion limited the global acquisition of chromatin accessibility associated with T cell exhaustion and led to improved anti-tumor immunity after adoptive cell therapy. In summary, we provide a comprehensive atlas of the genetic regulators of T cell exhaustion and demonstrate that modulation of the epigenetic state of T cell exhaustion can improve T cell responses in cancer immunotherapy.
2022-05-26 | GSE203592 | GEO
Project description:T cell exhaustion limits anti-tumor immunity, but the molecular determinants of this process remain poorly understood. Using a chronic antigen stimulation assay, we performed genome-wide CRISPR/Cas9 screens to systematically discover genetic regulators of T cell exhaustion, which identified an enrichment of epigenetic factors. In vivo CRISPR screens in murine and human tumor models demonstrated that perturbation of several epigenetic regulators, including members of the INO80 and BAF chromatin remodeling complexes, improved T cell persistence in tumors. In vivo paired CRISPR perturbation and single-cell RNA sequencing revealed distinct transcriptional roles of each complex and that depletion of canonical BAF complex members, including Arid1a, resulted in the maintenance of an effector program and downregulation of terminal exhaustion-related genes in tumor-infiltrating T cells. Finally, Arid1a-depletion limited the global acquisition of chromatin accessibility associated with T cell exhaustion and led to improved anti-tumor immunity after adoptive cell therapy. In summary, we provide a comprehensive atlas of the genetic regulators of T cell exhaustion and demonstrate that modulation of the epigenetic state of T cell exhaustion can improve T cell responses in cancer immunotherapy.
2022-05-26 | GSE203591 | GEO
Project description:Genome-wide CRISPR screens of T cell exhaustion identify chromatin remodeling factors that limit T cell persistence
| PRJNA841506 | ENA
Project description:Genome-wide CRISPR screens of T cell exhaustion identify chromatin remodeling factors that limit T cell persistence [Perturb-seq]
| PRJNA841559 | ENA
Project description:Genome-wide CRISPR screens of T cell exhaustion identify chromatin remodeling factors that limit T cell persistence [ATAC-seq]
| PRJNA841558 | ENA
Project description:Most genetic alterations that drive melanoma development and resistance to targeted therapy have been uncovered. In contrast, and despite their increasingly recognized contribution, little is known about the non-genetic mechanisms that drive these processes. Here, we performed in vivo gain-of-function CRISPR screens and identified SMAD3, BIRC3 and SLC9A5 as key actors of BRAFi-resistance. We show that their expression levels increase during acquisition of BRAFi-resistance, and remain high in persister cells and during relapse. The upregulation of the SMAD3 transcriptional activity (SMAD3-signature) promotes a mesenchymal-like phenotype and BRAFi-resistance by acting as an upstream transcriptional regulator of potent BRAFi-resistance genes such as EGFR and AXL. This SMAD3-signature predicts resistance to both current melanoma therapies in different cohorts. Critically, chemical inhibition of SMAD3 may constitute amenable target for melanoma since it efficiently abrogates persister cells survival. Interestingly, decrease of SMAD3 activity can also be reached by inhibiting the Aryl hydrocarbon Receptor (AhR), another druggable transcription factor governing SMAD3 expression level. Our work highlights novel drug vulnerabilities that can be exploited to develop long-lasting antimelanoma therapies.
| S-SCDT-EMM-2020-13466 | biostudies-other
Project description:Comprehensive identification of factors that can specify neuronal fate could provide valuable insights into lineage specification and reprogramming, but systematic interrogation of transcription factors, and their interactions with each other, has proven technically challenging. We developed a CRISPR activation (CRISPRa) approach to systematically identify regulators of neuronal-fate specification. We activated expression of all endogenous transcription factors and other regulators via a pooled CRISPRa screen in embryonic stem cells, revealing genes including epigenetic regulators such as Ezh2 that can induce neuronal fate. Systematic CRISPR-based activation of factor pairs allowed us to generate a genetic interaction map for neuronal differentiation, with confirmation of top individual and combinatorial hits as bona fide inducers of neuronal fate. Several factor pairs could directly reprogram fibroblasts into neurons, which shared similar transcriptional programs with endogenous neurons. This study provides an unbiased discovery approach for systematic identification of genes that drive cell-fate acquisition.
| S-EPMC6214761 | biostudies-literature
Project description:BackgroundThe COVID-19 pandemic has resulted in 275 million infections and 5.4 million deaths as of December 2021. While effective vaccines are being administered globally, there is still a great need for antiviral therapies as antigenically novel SARS-CoV-2 variants continue to emerge across the globe. Viruses require host factors at every step in their life cycle, representing a rich pool of candidate targets for antiviral drug design.MethodsTo identify host factors that promote SARS-CoV-2 infection with potential for broad-spectrum activity across the coronavirus family, we performed genome-scale CRISPR knockout screens in two cell lines (Vero E6 and HEK293T ectopically expressing ACE2) with SARS-CoV-2 and the common cold-causing human coronavirus OC43. Gene knockdown, CRISPR knockout, and small molecule testing in Vero, HEK293, and human small airway epithelial cells were used to verify our findings.ResultsWhile we identified multiple genes and functional pathways that have been previously reported to promote human coronavirus replication, we also identified a substantial number of novel genes and pathways. The website https://sarscrisprscreens.epi.ufl.edu/ was created to allow visualization and comparison of SARS-CoV2 CRISPR screens in a uniformly analyzed way. Of note, host factors involved in cell cycle regulation were enriched in our screens as were several key components of the programmed mRNA decay pathway. The role of EDC4 and XRN1 in coronavirus replication in human small airway epithelial cells was verified. Finally, we identified novel candidate antiviral compounds targeting a number of factors revealed by our screens.ConclusionsOverall, our studies substantiate and expand the growing body of literature focused on understanding key human coronavirus-host cell interactions and exploit that knowledge for rational antiviral drug development.
| S-EPMC8792531 | biostudies-literature
Project description:Most genetic alterations that drive melanoma development and resistance to targeted therapy have been uncovered. In contrast, and despite their increasingly recognized contribution, little is known about the non-genetic mechanisms that drive these processes. Here, we performed in vivo gain-of-function CRISPR screens and identified SMAD3, BIRC3, and SLC9A5 as key actors of BRAFi resistance. We show that their expression levels increase during acquisition of BRAFi resistance and remain high in persister cells and during relapse. The upregulation of the SMAD3 transcriptional activity (SMAD3-signature) promotes a mesenchymal-like phenotype and BRAFi resistance by acting as an upstream transcriptional regulator of potent BRAFi-resistance genes such as EGFR and AXL. This SMAD3-signature predicts resistance to both current melanoma therapies in different cohorts. Critically, chemical inhibition of SMAD3 may constitute amenable target for melanoma since it efficiently abrogates persister cells survival. Interestingly, decrease of SMAD3 activity can also be reached by inhibiting the Aryl hydrocarbon Receptor (AhR), another druggable transcription factor governing SMAD3 expression level. Our work highlights novel drug vulnerabilities that can be exploited to develop long-lasting antimelanoma therapies.
| S-EPMC8103100 | biostudies-literature