Analysis of Chromatin ADP-Ribosylation at the Genome-wide Level and at Specific Loci by ADPr-ChAP

Giody Bartolomei, Mario Leutert, Massimiliano Manzo, Tuncay Baubec, Michael O Hottiger

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Chromatin ADP-ribosylation regulates important cellular processes. However, the exact location and magnitude of chromatin ADP-ribosylation are largely unknown. A robust and versatile method for assessing chromatin ADP-ribosylation is therefore crucial for further understanding its function. Here, we present a chromatin affinity precipitation method based on the high specificity and avidity of two well-characterized ADP-ribose binding domains to map chromatin ADP-ribosylation at the genome-wide scale and at specific loci. Our ADPr-ChAP method revealed that in cells exposed to oxidative stress, ADP-ribosylation of chromatin scales with histone density, with highest levels at heterochromatic sites and depletion at active promoters. Furthermore, in growth factor-induced adipocyte differentiation, increased chromatin ADP-ribosylation was observed at PPARγ target genes, whose expression is ADP-ribosylation dependent. In combination with deep-sequencing and conventional chromatin immunoprecipitation, the established ADPr-ChAP provides a valuable resource for the bioinformatic comparison of ADP-ribosylation with other chromatin modifications and for addressing its role in other biologically important processes.

Original languageEnglish
Pages (from-to)474-485
Number of pages12
JournalMolecular Cell
Volume61
Issue number3
DOIs
Publication statusPublished - 4 Feb 2016
Externally publishedYes

Bibliographical note

Copyright © 2016 Elsevier Inc. All rights reserved.

Keywords

  • 3T3-L1 Cells
  • Adenosine Diphosphate Ribose/metabolism
  • Adipocytes/drug effects
  • Adipogenesis
  • Animals
  • Binding Sites
  • Cell Line, Tumor
  • Cell Nucleus/drug effects
  • Chromatin/genetics
  • Chromatin Immunoprecipitation/methods
  • Computational Biology
  • Gene Expression Regulation
  • Growth Hormone/pharmacology
  • High-Throughput Nucleotide Sequencing
  • Humans
  • Hydrogen Peroxide/pharmacology
  • Mice
  • Oxidative Stress
  • PPAR gamma/genetics
  • Poly (ADP-Ribose) Polymerase-1
  • Poly(ADP-ribose) Polymerases/genetics
  • Promoter Regions, Genetic
  • Protein Binding
  • Protein Structure, Tertiary
  • Transfection

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