Human Rad17 C-terminal tail is phosphorylated by concerted action of CK1δ/ε and CK2 to promote interaction with the 9-1-1 complex

Biochem Biophys Res Commun. 2019 Sep 17;517(2):310-316. doi: 10.1016/j.bbrc.2019.07.076. Epub 2019 Jul 25.

Abstract

The ATR-dependent DNA damage checkpoint is one of the major checkpoint pathways. The interaction between the Rad17-RFC2-5 and 9-1-1 complexes is central to the ATR-Chk1 pathway. However, little is known about the regulation of the interaction. We recently showed that vertebrate Rad17 proteins share a conserved C-terminal tail and that the C-terminal tails have a conserved amino acid motif named iVERGE that must be intact for the interaction between Rad17 and the 9‒1‒1 complex. In human Rad17, the Y665 and S667 residues are conserved in iVERGE. The Rad17-S667 residue is phosphorylated by CK2, and the phosphorylation is important for the interaction with the 9‒1‒1 complex. Here, we show that a C-terminal threonine residue of Rad17, T670 in human Rad17, is constitutively phosphorylated in vivo. The T670 phosphorylation is important for the S667 phosphorylation, and vice versa. Phosphomimetic mutations in the T670 residue promote the interaction with the 9-1-1 complex. The T670 and Y665 residues show functional redundancy, and their roles are dependent on the S667 phosphorylation. Rad17-T670 is phosphorylated by casein kinase 1δ/ε. Our data suggest that iVERGE integrates multiple signaling pathways to regulate the ATR-Chk1 pathway.

Keywords: 9–1–1 complex; ATR; CK1; Cell cycle checkpoint; DNA damage response; Rad17.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Ataxia Telangiectasia Mutated Proteins / metabolism
  • COS Cells
  • Casein Kinase 1 epsilon / metabolism*
  • Casein Kinase II / metabolism*
  • Casein Kinase Idelta / metabolism*
  • Cell Cycle Proteins / chemistry
  • Cell Cycle Proteins / metabolism*
  • Chlorocebus aethiops
  • DNA Damage
  • Humans
  • Phosphorylation
  • Protein Interaction Maps*
  • Signal Transduction

Substances

  • Cell Cycle Proteins
  • HUS1 protein, human
  • Rad17 protein, human
  • ATR protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • Casein Kinase 1 epsilon
  • Casein Kinase II
  • Casein Kinase Idelta