Phosphoproteomics reveals distinct modes of Mec1/ATR signaling during DNA replication

Mol Cell. 2015 Mar 19;57(6):1124-1132. doi: 10.1016/j.molcel.2015.01.043. Epub 2015 Mar 5.

Abstract

The Mec1/Tel1 kinases (human ATR/ATM) play numerous roles in the DNA replication stress response. Despite the multi-functionality of these kinases, studies of their in vivo action have mostly relied on a few well-established substrates. Here we employed a combined genetic-phosphoproteomic approach to monitor Mec1/Tel1 signaling in a systematic, unbiased, and quantitative manner. Unexpectedly, we find that Mec1 is highly active during normal DNA replication, at levels comparable or higher than Mec1's activation state induced by replication stress. This "replication-correlated" mode of Mec1 action requires the 9-1-1 clamp and the Dna2 lagging-strand factor and is distinguishable from Mec1's action in activating the downstream kinase Rad53. We propose that Mec1/ATR performs key functions during ongoing DNA synthesis that are distinct from their canonical checkpoint role during replication stress.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Ataxia Telangiectasia Mutated Proteins / genetics
  • Ataxia Telangiectasia Mutated Proteins / metabolism
  • Cell Cycle Proteins / genetics
  • Cell Cycle Proteins / metabolism
  • Checkpoint Kinase 2 / genetics
  • Checkpoint Kinase 2 / metabolism
  • DNA Replication*
  • Humans
  • Intracellular Signaling Peptides and Proteins / genetics
  • Intracellular Signaling Peptides and Proteins / metabolism*
  • Phosphoproteins / analysis
  • Phosphoproteins / genetics
  • Phosphoproteins / metabolism
  • Phosphorylation
  • Protein-Serine-Threonine Kinases / genetics
  • Protein-Serine-Threonine Kinases / metabolism*
  • Proteomics / methods*
  • Proto-Oncogene Proteins c-ets / genetics
  • Proto-Oncogene Proteins c-ets / metabolism*
  • Repressor Proteins / genetics
  • Repressor Proteins / metabolism*
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Signal Transduction

Substances

  • Cell Cycle Proteins
  • Ddc1 protein, S cerevisiae
  • ETS translocation variant 6 protein
  • Intracellular Signaling Peptides and Proteins
  • Phosphoproteins
  • Proto-Oncogene Proteins c-ets
  • Repressor Proteins
  • Saccharomyces cerevisiae Proteins
  • Checkpoint Kinase 2
  • ATR protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • MEC1 protein, S cerevisiae
  • Protein-Serine-Threonine Kinases
  • TEL1 protein, S cerevisiae
  • RAD53 protein, S cerevisiae