Mutational analysis of the poly(ADP-ribosyl)ation sites of the transcription factor CTCF provides an insight into the mechanism of its regulation by poly(ADP-ribosyl)ation

Dawn Farrar, Sushma Rai, Igor Chernukhin, Maja Jagodic, Yoko Ito, Samer Yammine, Rolf Ohlsson, Adele Murrell, Elena Klenova

Research output: Contribution to journalArticle

53 Citations (Scopus)

Abstract

Poly(ADP-ribosyl)ation of the conserved multifunctional transcription factor CTCF was previously identified as important to maintain CTCF insulator and chromatin barrier functions. However, the molecular mechanism of this regulation and also the necessity of this modification for other CTCF functions remain unknown. In this study, we identified potential sites of poly(ADP-ribosyl)ation within the N-terminal domain of CTCF and generated a mutant deficient in poly(ADP-ribosyl)ation. Using this CTCF mutant, we demonstrated the requirement of poly(ADP-ribosyl)ation for optimal CTCF function in transcriptional activation of the p19ARF promoter and inhibition of cell proliferation. By using a newly generated isogenic insulator reporter cell line, the CTCF insulator function at the mouse Igf2-H19 imprinting control region (ICR) was found to be compromised by the CTCF mutation. The association and simultaneous presence of PARP-1 and CTCF at the ICR, confirmed by single and serial chromatin immunoprecipitation assays, were found to be independent of CTCF poly(ADP-ribosyl)ation. These results suggest a model of CTCF regulation by poly(ADP-ribosyl)ation whereby CTCF and PARP-1 form functional complexes at sites along the DNA, producing a dynamic reversible modification of CTCF. By using bioinformatics tools, numerous sites of CTCF and PARP-1 colocalization were demonstrated, suggesting that such regulation of CTCF may take place at the genome level.
Original languageEnglish
Pages (from-to)1199-1216
Number of pages18
JournalMolecular and Cellular Biology
Volume30
Issue number5
Early online date28 Dec 2009
DOIs
Publication statusPublished - Mar 2010

Keywords

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • Base Sequence
  • Binding Sites
  • Cell Line
  • Cell Proliferation
  • DNA Primers
  • Genomic Imprinting
  • HeLa Cells
  • Humans
  • Hybrid Cells
  • Insulin-Like Growth Factor II
  • Mice
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Mutation
  • Poly Adenosine Diphosphate Ribose
  • Poly(ADP-ribose) Polymerases
  • RNA, Long Noncoding
  • RNA, Untranslated
  • Recombinant Proteins
  • Repressor Proteins
  • Transfection

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