CRISPR Activation/Interference Screen to Identify Genetic Networks in HDAC-Inhibitor-Resistant Cells.
Methods Mol Biol
; 2589: 429-454, 2023.
Article
en En
| MEDLINE
| ID: mdl-36255641
ABSTRACT
Epigenetic alterations have been identified in various tumor types. In part, these alterations are mediated via increased histone deacetylase activity. Although preclinical results of monotherapies with histone deacetylase inhibitors (HDACi) are promising, success in clinical trials is limited. Reasons for these limitations may be de novo or acquired resistance to HDAC inhibitors that could be overcome with rational combination therapies. This requires knowledge of resistance mechanism along with the involved genetic networks. One way to identify such genetic networks is the implementation of a CRISPR-based technology allowing transcriptional repression (CRISPRi) and activation (CRISPRa) at a genome-wide scale. We describe a simple approach to amplify and validate sgRNA libraries, generate a myeloid progenitor cell line expressing catalytically dead Cas9 (dCas9) fusion proteins with transcriptional effectors to repress or activate genetic regions of interest and demonstrate a complementary genome-wide HDACi resistance screening approach. Furthermore, we present bioinformatics tools for quality control and analysis of the sequencing data.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Redes Reguladoras de Genes
/
Inhibidores de Histona Desacetilasas
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2023
Tipo del documento:
Article
País de afiliación:
Alemania