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High affinity chromodomains engineered for improved detection of histone modifications and enhanced CRISPR-based gene repression

ABSTRACT: Histone methylation is an important post-translational modification that plays a crucial role in regulating cellular functions, and its dysregulation is implicated in cancer and developmental defects. Therefore, systematic characterization of histone methylation is necessary to elucidate complex biological processes, identify biomarkers, and ultimately, enable drug discovery. Studying histone methylation relies on the use of antibodies, but these suffer from batch-to-batch variation, are costly, and cannot be used in live cells. Chromatin-modification reader domains are potential affinity reagents for methylated histones, but their application is limited by their modest affinities. We used phage display to identify key residues that greatly enhance the affinities of Cbx chromodomains for methylated histone marks and developed a general strategy for enhancing the affinity of most chromodomains of the human Cbx protein family. Our strategy allowed us to develop powerful probes for genome-wide binding analysis and live cell imaging. Furthermore, we used optimized chromodomains to develop extremely potent CRISPR-based repressors for tailored gene silencing. Our results highlight the power of engineered chromodomains for analyzing protein interaction networks involving chromatin and represent a modular platform for efficient gene silencing.

ORGANISM(S): Mus musculus

PROVIDER: GSE188439 | GEO | 2022/09/08

REPOSITORIES: GEO

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