Interaction between human MCM7 and Rad17 proteins is required for replication checkpoint signaling

EMBO J. 2004 Nov 24;23(23):4660-9. doi: 10.1038/sj.emboj.7600463. Epub 2004 Nov 11.

Abstract

Human Rad17 (hRad17) is centrally involved in the activation of cell-cycle checkpoints by genotoxic agents or replication stress. Here we identify hMCM7, a core component of the DNA replication apparatus, as a novel hRad17-interacting protein. In HeLa cells, depletion of either hRad17 or hMCM7 with small-interfering RNA suppressed ultraviolet (UV) light- or aphidicolin-induced hChk1 phosphorylation, and abolished UV-induced S-phase checkpoint activation. Similar results were obtained after transfection of these cells with a fusion protein containing the hMCM7-binding region of hRad17. The hMCM7-depleted cells were also defective for the formation of ATR-containing nuclear foci after UV irradiation, suggesting that hMCM7 is required for stable recruitment of ATR to damaged DNA. These results demonstrate that hMCM7 plays a direct role in the transmission of DNA damage signals from active replication forks to the S-phase checkpoint machinery in human cells.

Publication types

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

MeSH terms

  • Aphidicolin / pharmacology
  • Ataxia Telangiectasia Mutated Proteins
  • Cell Cycle Proteins / genetics
  • Cell Cycle Proteins / metabolism
  • Cell Cycle Proteins / physiology*
  • Cell Line, Tumor
  • Checkpoint Kinase 1
  • Checkpoint Kinase 2
  • DNA Damage
  • DNA Replication / physiology*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / physiology*
  • Green Fluorescent Proteins / genetics
  • HeLa Cells
  • Humans
  • Minichromosome Maintenance Complex Component 7
  • Nuclear Proteins / genetics
  • Nuclear Proteins / physiology*
  • Phosphorylation
  • Protein Binding
  • Protein Kinases / metabolism
  • Protein Serine-Threonine Kinases / metabolism
  • Recombinant Fusion Proteins / genetics
  • S Phase / physiology
  • Signal Transduction
  • Transfection
  • Two-Hybrid System Techniques
  • Ultraviolet Rays

Substances

  • Cell Cycle Proteins
  • DNA-Binding Proteins
  • Nuclear Proteins
  • Rad17 protein, human
  • Recombinant Fusion Proteins
  • Green Fluorescent Proteins
  • Aphidicolin
  • Protein Kinases
  • Checkpoint Kinase 2
  • ATR protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • CHEK1 protein, human
  • CHEK2 protein, human
  • Checkpoint Kinase 1
  • Protein Serine-Threonine Kinases
  • MCM7 protein, human
  • Minichromosome Maintenance Complex Component 7