Epi-Decoder: Decoding the Local Proteome of a Genomic Locus by Massive Parallel Chromatin Immunoprecipitation Combined with DNA-Barcode Sequencing

Methods Mol Biol. 2022:2458:123-150. doi: 10.1007/978-1-0716-2140-0_8.

Abstract

The genome in a eukaryotic cell is packaged into chromatin and regulated by chromatin-binding and chromatin-modifying factors. Many of these factors and their complexes have been identified before, but how each genomic locus interacts with its surrounding proteins in the nucleus over time and in changing conditions remains poorly described. Measuring protein-DNA interactions at a specific locus in the genome is challenging and current techniques such as capture of a locus followed by mass spectrometry require high levels of enrichment. Epi-Decoder, a method developed in budding yeast, enables systematic decoding of the proteome of a single genomic locus of interest without the need for locus enrichment. Instead, Epi-Decoder uses massive parallel chromatin immunoprecipitation of tagged proteins combined with barcoding a genomic locus and counting of coimmunoprecipitated barcodes by DNA sequencing (TAG-ChIP-Barcode-Seq). In this scenario, DNA barcode counts serve as a quantitative readout for protein binding of each tagged protein to the barcoded locus. Epi-Decoder can be applied to determine the protein-DNA interactions at a wide range of genomic loci, such as coding genes, noncoding genes, and intergenic regions. Furthermore, Epi-Decoder provides the option to study protein-DNA interactions upon changing cellular and/or genetic conditions. In this protocol, we describe in detail how to construct Epi-Decoder libraries and how to perform an Epi-Decoder analysis.

Keywords: ChIP; Chromatin; DNA barcodes; Epi-Decoder; Proteome.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatin Immunoprecipitation / methods
  • Chromatin* / genetics
  • DNA / chemistry
  • Genomics / methods
  • Proteome* / metabolism
  • Sequence Analysis, DNA

Substances

  • Chromatin
  • Proteome
  • DNA