Identification of Proteins Associated with an IFNγ-Responsive Promoter by a Retroviral Expression System for enChIP Using CRISPR

摘要: Isolation of specific genomic regions retaining molecular interactions is essential for comprehensive identification molecules associated with the regions. Recently, we developed engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP) technology purification Here, a retroviral expression system enChIP using CRISPR. We showed that target locus can be purified high efficiency by this system. also contamination potential off-target sites negligible if guide RNA (gRNA) site has sufficiently long unique sequence in its seed sequence. combined stable isotope labeling amino acids cell culture (SILAC) analysis identified proteins whose association interferon (IFN) regulatory factor-1 (IRF-1) promoter region increases response to IFNγ stimulation. The list contained many novel context IFNγ-induced gene as well related histone deacetylase complexes involvement been suggested IFNγ-mediated expression. Finally, confirmed increased IRF-1 ChIP. Thus, our results CRISPR would useful biochemical genome functions including transcription and epigenetic regulation.