Systematic characterization of BAF mutations provides insights into intracomplex synthetic lethalities in human cancers

Schick, Sandra; Rendeiro, André F.; Runggatscher, Kathrin; Ringler, Anna; Boidol, Bernd; Hinkel, Melanie; Marynen, Peter; Vulliard, Loan; Penz, Thomas; Parapatics, Katja; Schmidl, Christian; Menche, Jörg; Boehmelt, Guido; Petronczki, Mark; Müller, André C.; Bock, Christoph; Kubicek, Stefan

doi:10.1038/s41588-019-0477-9

Cited by 105 publications

(137 citation statements)

References 59 publications

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“…Compared with genes for which intron targeting sgRNAs did not result in isolation of GFP-positive cells, successfully targeted genes have higher average expression in HAP1 cells ( Fig. 1E; Schick et al 2019). By single-cell dilution, we then isolated 335 clonal cell lines for which a massively parallel multiplex sgRNA amplicon sequencing strategy unambiguously identified the integrated sgRNAs indicating a single tagged protein (Supplemental Note S1; Supplemental Table S3).…”

Section: Resultsmentioning

confidence: 99%

Pooled protein tagging, cellular imaging, and in situ sequencing for monitoring drug action in real time

2020

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The levels and subcellular localizations of proteins regulate critical aspects of many cellular processes and can become targets of therapeutic intervention. However, high-throughput methods for the discovery of proteins that change localization either by shuttling between compartments, by binding larger complexes, or by localizing to distinct membraneless organelles are not available. Here we describe a scalable strategy to characterize effects on protein localizations and levels in response to different perturbations. We use CRISPR-Cas9-based intron tagging to generate cell pools expressing hundreds of GFP-fusion proteins from their endogenous promoters and monitor localization changes by time-lapse microscopy followed by clone identification using in situ sequencing. We show that this strategy can characterize cellular responses to drug treatment and thus identify nonclassical effects such as modulation of protein–protein interactions, condensate formation, and chemical degradation.

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Section: Resultsmentioning

confidence: 99%

Pooled protein tagging, cellular imaging, and in situ sequencing for monitoring drug action in real time

2020

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“…Although we could show that knockout efficiency is only slightly decreased in diploid compared to haploid HAP1 cells, the effect is considered to be stronger in polyploid cancer cell lines where a full knockout requires out-of-frame editing of all alleles of a gene (Van Campenhout et al 2019) . HAP1 cells with specific gene knockouts have therefore been applied in many studies addressing various research questions (Davis et al 2015;Schick et al 2019;Lenk et al 2019;Smits et al 2019;Gerhards et al 2018;Baggen et al 2019) . The exact cellular context of this cell line is, however, not fully understood, especially since HAP1 cells do not share major characteristics with their parental chronic myeloid leukemia (CML) cell line KBM7.…”

Section: Discussionmentioning

confidence: 99%

Pooled CRISPR screening at high sensitivity with an empirically designed sgRNA library

Henkel

Rauscher

Schmitt

et al. 2020

Preprint

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Given their broad utility in functionally annotating genomes, the experimental design of genome-scale CRISPR screens can vary greatly and criteria for optimal experimental implementation and library composition are still emerging. In this study, we report advantages of conducting viability screens in selected Cas9 single cell clones in contrast to Cas9 bulk populations. We further systematically analyzed published CRISPR screens in human cells to identify single-guide (sg)RNAs with consistent high on-target and low off-target activity. Selected guides were collected in a new genome-scale sgRNA library, which efficiently identifies core and context-dependent essential genes. In summary, we show how empirically designed libraries in combination with an optimised experimental design increase the dynamic range in gene essentiality screens at reduced library coverage.

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“…Comprehensive analysis of synthetic lethality between 2 factors in all subunits of the SWI/SNF chromatin remodeling complex showed that the SMARCA4‐ARID2, SMARCA4‐ACTB, and SMARCC1‐SMARCC2 pairs are significant for synthetic lethality . Considering synthetic lethality among these pairs, the SMARCA4 and ARID2 relationship would be a potential target for synthetic lethal therapy because SMARCA4 and ARID2 are frequently mutated in cancer.…”

Section: Synthetic Lethal Targets Based On Targeting the Interaction mentioning

confidence: 99%

Synthetic lethal therapy based on targeting the vulnerability of SWI/SNF chromatin remodeling complex‐deficient cancers

Sasaki

Ogiwara

2020

Cancer Science

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The SWI/SNF chromatin remodeling complex is composed of approximately 15 subunits, and approximately 20% of all cancers carry mutations in the genes encoding these subunits. Most of the genetic alterations in these genes are loss‐of‐function mutations. The identification of vulnerability based on synthetic lethality in cancers with SWI/SNF chromatin remodeling complex deficiency contributes to precision medicine. The SWI/SNF chromatin remodeling complex is involved in transcription, DNA repair, DNA replication, and chromosomal segregation. Cancers with deficiency in the SWI/SNF chromatin remodeling complex show increased vulnerability derived from the loss of these functions. Synthetic lethal targets have been identified based on vulnerabilities in the functions of the SWI/SNF chromatin remodeling complex. In this review article, we propose a precision medicine strategy using chemotherapeutic methods, such as molecular targeted therapy and immunotherapy, based on harnessing synthetic lethality in cancers with deficiency in the SWI/SNF chromatin remodeling complex.

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Systematic characterization of BAF mutations provides insights into intracomplex synthetic lethalities in human cancers

Cited by 105 publications

References 59 publications

Pooled protein tagging, cellular imaging, and in situ sequencing for monitoring drug action in real time

Pooled protein tagging, cellular imaging, and in situ sequencing for monitoring drug action in real time

Pooled CRISPR screening at high sensitivity with an empirically designed sgRNA library

Synthetic lethal therapy based on targeting the vulnerability of SWI/SNF chromatin remodeling complex‐deficient cancers

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