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A Chromatin Immunoprecipitation Assay to Identify Novel NFAT2 Target Genes in Chronic Lymphocytic Leukemia
Published on: December 4, 2018
Chromatin complex dependencies reveal targeting opportunities in leukemia
Fadi J Najm1, Peter DeWeirdt2, Molly M Moore1
1Gene Regulation Observatory, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
Abstract:
Chromatin regulators are frequently mutated in human cancer and are attractive drug targets. They include diverse proteins that share functional domains and assemble into related multi-subunit complexes. To investigate functional relationships among these regulators, here we apply combinatorial CRISPR knockouts (KOs) to test over 35,000 gene-gene pairings in leukemia cells, using a library of over 300,000 constructs. Top pairs that demonstrate either compensatory non-lethal interactions or synergistic lethality enrich for paralogs and targets that occupy the same protein complex. The screen highlights protein complex dependencies not apparent in single KO screens, for example MCM histone exchange, the nucleosome remodeling and deacetylase (NuRD) complex, and HBO1 (KAT7) complex. We explore two approaches to NuRD complex inactivation. Paralog and non-paralog combinations of the KAT7 complex emerge as synergistic lethal and specifically nominate the ING5 PHD domain as a potential therapeutic target when paired with other KAT7 complex member losses. These findings highlight the power of combinatorial screening to provide mechanistic insight and identify therapeutic targets within redundant networks.
Insights
Combinatorial CRISPR screening reveals gene interactions in leukemia, identifying new therapeutic targets within chromatin regulator complexes like NuRD and KAT7. This approach uncovers dependencies missed by single gene studies.
Area of Science:
- Oncology
- Genetics
- Molecular Biology
Background:
- Chromatin regulators are frequently altered in human cancers, presenting potential therapeutic targets.
- These regulators function within multi-subunit complexes, suggesting complex-level dependencies in cancer.
Purpose of the Study:
- To investigate functional relationships among chromatin regulators using combinatorial CRISPR screening.
- To identify novel gene-gene interactions and potential therapeutic targets within chromatin regulator complexes.
Main Methods:
- Utilized a large-scale combinatorial CRISPR knockout screen testing over 35,000 gene-pair interactions in leukemia cells.
- Employed a library of over 300,000 constructs for comprehensive genetic perturbation.
Main Results:
- Identified gene pairs with compensatory non-lethal or synergistic lethal interactions, enriching for paralogs and members of the same protein complex.
- Highlighted dependencies within complexes such as MCM histone exchange, NuRD, and HBO1 (KAT7) complex, which were not evident from single knockout screens.
- Discovered synergistic lethality in KAT7 complex member combinations, specifically implicating the ING5 PHD domain as a potential therapeutic target.
Conclusions:
- Combinatorial screening is a powerful method for uncovering mechanistic insights into complex biological networks.
- This approach effectively identifies novel therapeutic targets within the redundant networks of chromatin regulators in cancer.
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