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Dictyostelium as a Model to Assess Site-Specific ADP-Ribosylation Events

  • Anna-Lena Kolb
  • Duen-Wei Hsu
  • Ana B. A. Wallis
  • Seiji Ura
  • Alina Rakhimova
  • Catherine J. Pears
  • Nicholas D. Lakin
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1813)

Abstract

The amoeba Dictyostelium discoideum is a single-cell organism that can undergo a simple developmental program, making it an excellent model to study the molecular mechanisms of cell motility, signal transduction, and cell-type differentiation. A variety of human genes that are absent or show limited conservation in other invertebrate models have been identified in this organism. This includes ADP-ribosyltransferases, also known as poly-ADP-ribose polymerases (PARPs), a family of proteins that catalyze the addition of single or poly-ADP-ribose moieties onto target proteins. The genetic tractability of Dictyostelium and its relatively simple genome structure makes it possible to disrupt PARP gene combinations, in addition to specific ADP-ribosylation sites at endogenous loci. Together, this makes Dictyostelium an attractive model to assess how ADP-ribosylation regulates a variety of cellular processes including DNA repair, transcription, and cell-type specification. Here we describe a range of techniques to study ADP-ribosylation in Dictyostelium, including analysis of ADP-ribosylation events in vitro and in vivo, in addition to approaches to assess the functional roles of this modification in vivo.

Key words

ADP-ribosylation PARP ADP-ribosyltransferase Dictyostelium DNA repair 

Notes

Acknowledgments

We thank members of the Lakin and Pears laboratories for constructive comments during the preparation of this manuscript. N.L.’s laboratory is supported by Cancer Research UK (www.cancerresearch.org.uk; grant C1521/A12353), Medical Research Council (www.mrc.ac.uk; MR/L000164/1), and NC3Rs (www.nc3rs.org.uk; NC/K00137X/1). AR was supported by a Clarendon Award (University of Oxford). C.P.’s laboratory is supported by NC3Rs (www.nc3rs.org.uk; NC/M000834/1).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Anna-Lena Kolb
    • 1
  • Duen-Wei Hsu
    • 1
  • Ana B. A. Wallis
    • 1
  • Seiji Ura
    • 1
  • Alina Rakhimova
    • 1
  • Catherine J. Pears
    • 1
  • Nicholas D. Lakin
    • 1
  1. 1.Department of BiochemistryUniversity of OxfordOxfordUK

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