To map the genomic interaction sites of chromatin proteins, two related methods were developed and experimentally explored in Saccharomyces cerevisiae. The ChIC method (chromatin immunocleavage) consists of tethering a fusion protein (pA-MN) consisting of micrococcal nuclease (MN) and staphylococcal protein A to specifically bound antibodies. The nuclease is kept inactive during the tethering process (no Ca2+). The ChEC method (chromatin endogenous cleavage) consists of expressing fusion proteins in vivo, where MN is C-terminally fused to the proteins of interest. The specifically tethered nucleases are activated with Ca2+ ions to locally introduce double-stranded DNA breaks. We demonstrate that ChIC and ChEC map proteins with a 100-200 bp resolution and excellent specificity. One version of the method is applicable to formaldehyde-fixed nuclei, another to native cells with comparable results. Among various model experiments, these methods were used to address the conformation of yeast telomeres.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Annotation Extension||Reference|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Assay||Construct||Conditions||Strain Background||Reference|