Unit

UNIT 18.7 Nonisotopic Methods for Determination of Poly(ADP-Ribose) Levels and Detection of Poly(ADP-Ribose) Polymerase

  1. Jean François Haince1,
  2. Guy G. Poirier1,
  3. Jim B. Kirkland2

Published Online: 1 FEB 2004

DOI: 10.1002/0471143030.cb1807s21

Current Protocols in Cell Biology

Current Protocols in Cell Biology

How to Cite

Haince, J. F., Poirier, G. G. and Kirkland, J. B. 2004. Nonisotopic Methods for Determination of Poly(ADP-Ribose) Levels and Detection of Poly(ADP-Ribose) Polymerase. Current Protocols in Cell Biology. 21:18.7:18.7.1–18.7.26.

Author Information

  1. 1

    Laval University Medical Research Center, Ste.-Foy, Quebec, Canada

  2. 2

    University of Guelph, Guelph, Ontario, Canada

Publication History

  1. Published Online: 1 FEB 2004
  2. Published Print: DEC 2003

Abstract

Poly(ADP-ribosyl)ation is a post-translational modification catalyzed mostly by the 116-kDa enzyme poly(ADP-ribose) polymerase-1 (PARP-1), a nuclear enzyme that transfers an ADP-ribose moiety onto a limited number of nuclear proteins, including itself. When cells are exposed to environmental stresses such as alkylating agents or free radicals, there is up to a 500-fold increase in net poly(ADP-ribose) synthesis in response to DNA strand breaks. The enzyme responsible for 80% to 90% of this stimulated poly(ADP-ribose) synthesis is PARP-1, while other PARPs are responsible for the remaining 10% to 20%. The physiological meaning of these phenomena is not clear; however, it can be interpreted as a way of translating an event occurring on DNA to the nucleus by protein modification and finally to the cytoplasm via NAD+ depletion. It has also been proposed that the presence of negatively charged poly(ADP-ribose) at the site of DNA damage may play several roles in regulation of base excision repair, p53 functions, and apoptosis. This unit describes protocols for measuring the levels of poly(ADP-ribose) in cells using nonisotopic reagents and for identifying the poly(ADP-ribose) polymerase enzymes present in cells.

Keywords:

  • poly(ADP-ribose);
  • poly(ADP-ribose)polymerase;
  • apoptosis;
  • DNA damage