ADP-ribosylation of ADPR-transferase and topoisomerase I in intact mouse epidermal cells JB6

Biochemistry. 1989 Mar 7;28(5):2034-40. doi: 10.1021/bi00431a011.

Abstract

Poly(ADP-ribosylation) [poly(ADPR)] is a posttranslational modification of chromosomal proteins that affects the structural and functional properties of chromatin. We have studied poly(ADPR) of ADPR-transferase and topoisomerase I in intact mouse epidermal cells JB6 (clone 41) by a combination of affinity chromatography on phenylboronate and immunoblotting with monoclonal antibodies against poly(ADPR) chains and polyclonal antibodies against ADPR-transferase and topoisomerase I, respectively. Constitutive, steady-state poly(ADPR) substitution of ADPR-transferase was estimated at 4% and that of topoisomerase I at 0.1%. Active oxygen produced extracellularly by xanthine-xanthine oxidase and the methylating agent N-methyl-N'-nitro-N-nitrosoguanidine transiently increased the level of poly(ADPR) substitution of these enzymes by a factor of 6-10. While the poly(ADPR) substitution of ADPR-transferase remained elevated after 60 min of incubation, the poly(ADPR) substitution of topoisomerase I had returned to control values within this time. Benzamide (100 microM) partially prevented the stimulation of poly(ADPR) synthesis by these agents. We speculate that self-inactivation of ADPR-transferase by poly(ADPR) represents a feedback mechanism that has the function to avoid excessive poly(ADPR) synthesis and concomitant NAD and ATP depletion. Inactivation of topoisomerase I in the neighborhood of DNA breakage may temporarily shut down DNA replication and allow DNA repair to occur.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal
  • Benzamides
  • Cells, Cultured
  • Chromatography, Affinity
  • DNA Topoisomerases, Type I / metabolism*
  • Immunoblotting
  • Methylnitronitrosoguanidine
  • Nucleoside Diphosphate Sugars / metabolism*
  • Oxygen
  • Poly Adenosine Diphosphate Ribose / metabolism*
  • Poly(ADP-ribose) Polymerases / metabolism*
  • Protein Processing, Post-Translational

Substances

  • Antibodies, Monoclonal
  • Benzamides
  • Nucleoside Diphosphate Sugars
  • Methylnitronitrosoguanidine
  • Poly Adenosine Diphosphate Ribose
  • benzamide
  • Poly(ADP-ribose) Polymerases
  • DNA Topoisomerases, Type I
  • Oxygen