KA1-targeted regulatory domain mutations activate Chk1 in the absence of DNA damage

Sci Rep. 2015 Jun 3;5:10856. doi: 10.1038/srep10856.

Abstract

The Chk1 protein kinase is activated in response to DNA damage through ATR-mediated phosphorylation at multiple serine-glutamine (SQ) residues within the C-terminal regulatory domain, however the molecular mechanism is not understood. Modelling indicates a high probability that this region of Chk1 contains a kinase-associated 1 (KA1) domain, a small, compact protein fold found in multiple protein kinases including SOS2, AMPK and MARK3. We introduced mutations into Chk1 designed to disrupt specific structural elements of the predicted KA1 domain. Remarkably, six of seven Chk1 KA1 mutants exhibit constitutive biological activity (Chk1-CA) in the absence of DNA damage, profoundly arresting cells in G2 phase of the cell cycle. Cell cycle arrest induced by selected Chk1-CA mutants depends on kinase catalytic activity, which is increased several-fold compared to wild-type, however phosphorylation of the key ATR regulatory site serine 345 (S345) is not required. Thus, mutations targeting the putative Chk1 KA1 domain confer constitutive biological activity by circumventing the need for ATR-mediated positive regulatory phosphorylation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Catalysis
  • Cell Proliferation
  • Checkpoint Kinase 1
  • DNA Damage*
  • Enzyme Activation
  • G2 Phase Cell Cycle Checkpoints / genetics
  • Humans
  • Models, Biological
  • Molecular Sequence Data
  • Mutation*
  • Phosphorylation
  • Protein Interaction Domains and Motifs / genetics*
  • Protein Kinases / chemistry
  • Protein Kinases / genetics*
  • Protein Kinases / metabolism*
  • Son of Sevenless Proteins / chemistry
  • Son of Sevenless Proteins / genetics
  • Son of Sevenless Proteins / metabolism

Substances

  • SOS2 protein, human
  • Son of Sevenless Proteins
  • Protein Kinases
  • CHEK1 protein, human
  • Checkpoint Kinase 1