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A truncated DNA-damage-signaling response is activated after DSB formation in the G1 phase of Saccharomyces cerevisiae

Academic Article
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Overview

authors

  • Janke, R.
  • Herzberg, K.
  • Rolfsmeier, M.
  • Mar, J.
  • Bashkirov, V. I.
  • Haghnazari, E.
  • Cantin, G.
  • Yates III, John
  • Heyer, W. D.

publication date

  • April 2010

journal

  • Nucleic Acids Research  Journal

abstract

  • In Saccharomyces cerevisiae, the DNA damage response (DDR) is activated by the spatio-temporal colocalization of Mec1-Ddc2 kinase and the 9-1-1 clamp. In the absence of direct means to monitor Mec1 kinase activation in vivo, activation of the checkpoint kinase Rad53 has been taken as a proxy for DDR activation. Here, we identify serine 378 of the Rad55 recombination protein as a direct target site of Mec1. Rad55-S378 phosphorylation leads to an electrophoretic mobility shift of the protein and acts as a sentinel for Mec1 activation in vivo. A single double-stranded break (DSB) in G1-arrested cells causes phosphorylation of Rad55-S378, indicating activation of Mec1 kinase. However, Rad53 kinase is not detectably activated under these conditions. This response required Mec1-Ddc2 and loading of the 9-1-1 clamp by Rad24-RFC, but not Rad9 or Mrc1. In addition to Rad55-S378, two additional direct Mec1 kinase targets are phosphorylated, the middle subunit of the ssDNA-binding protein RPA, RPA2 and histone H2A (H2AX). These data suggest the existence of a truncated signaling pathway in response to a single DSB in G1-arrested cells that activates Mec1 without eliciting a full DDR involving the entire signaling pathway including the effector kinases.

subject areas

  • Adaptor Proteins, Signal Transducing
  • Cell Cycle Proteins
  • Checkpoint Kinase 2
  • DNA Breaks, Double-Stranded
  • DNA Repair
  • DNA-Binding Proteins
  • Electrophoretic Mobility Shift Assay
  • G1 Phase
  • Histones
  • Intracellular Signaling Peptides and Proteins
  • Phosphorylation
  • Protein-Serine-Threonine Kinases
  • Saccharomyces cerevisiae
  • Saccharomyces cerevisiae Proteins
  • Serine
  • Signal Transduction
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Identity

PubMed Central ID

  • PMC2853130

International Standard Serial Number (ISSN)

  • 0305-1048

Digital Object Identifier (DOI)

  • 10.1093/nar/gkp1222

PubMed ID

  • 20061370
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Additional Document Info

start page

  • 2302

end page

  • 2313

volume

  • 38

issue

  • 7

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