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Detecting and Quantifying pADPr In Vivo

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Poly(ADP-Ribose) Polymerase

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1608))

Abstract

Poly(ADP-ribose) polymerases (PARP) participate in diverse biological processes contributing to cellular homeostasis or exacerbating injury. PARP catalyzes the addition of ADP-ribose molecules (pADPr) to the target proteins, a process termed poly-ADP-ribosylation. Overactivation of PARP, as reflected by increased poly-ADP-ribosylation, accumulation of pADPr-modified proteins or free pADPr, contributes to depletion of NAD+ and mitochondrial dysfunction, potentially leading to cell death. Since PARP overactivation and increases in free pADPr have been identified as key contributors to the pathobiology of many diseases, monitoring PARP-1 activation by detecting and quantifying pADPr may provide valuable mechanistic insights as well as facilitating therapeutic drug monitoring for PARP inhibitors.

Several non-isotopic immunodetection methods for quantifying pADPr are discussed: western blotting of poly-ADP-ribosylated proteins, cellular localization of pADPr by immunohistochemistry, quantification of pADPr by enzyme-linked immunoassay and small scale two-dimensional gel electrophoresis.

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Acknowledgements

This work was supported by P50 NS30318 (RSC) and T32 HD40686 (YL, MAS).

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Authors and Affiliations

  1. Department of Pediatrics, Baylor College of Medicine, Houston, TX, USA

    Yi-Chen Lai M.D.

  2. Section of Pediatric Critical Care Medicine, Texas Children’s Hospital, 6621 Fannin Street, WT6-006, Houston, TX, 77030, USA

    Yi-Chen Lai M.D.

  3. Department of Critical Care Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

    Rajesh K. Aneja M.D. & Robert S. B. Clark M.D.

  4. Children’s Hospital of Pittsburgh of UPMC, Pittsburgh, PA, USA

    Rajesh K. Aneja M.D., Margaret A. Satchell M.D. & Robert S. B. Clark M.D.

  5. Safar Center for Resuscitation Research, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

    Robert S. B. Clark M.D.

Authors
  1. Yi-Chen Lai M.D.
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  2. Rajesh K. Aneja M.D.
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  3. Margaret A. Satchell M.D.
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  4. Robert S. B. Clark M.D.
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Corresponding author

Correspondence to Yi-Chen Lai M.D. .

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Editors and Affiliations

  1. Cancer Biology Program, Fox Chase Cancer Center, Philadelphia, Pennsylvania, USA

    Alexei V. Tulin

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Lai, YC., Aneja, R.K., Satchell, M.A., Clark, R.S.B. (2017). Detecting and Quantifying pADPr In Vivo. In: Tulin, A. (eds) Poly(ADP-Ribose) Polymerase. Methods in Molecular Biology, vol 1608. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6993-7_3

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  • DOI: https://doi.org/10.1007/978-1-4939-6993-7_3

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6992-0

  • Online ISBN: 978-1-4939-6993-7

  • eBook Packages: Springer Protocols

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