Centrosomal Localization of DNA Damage Checkpoint Proteins

J Cell Biochem. 2007 May 15;101(2):451-65. doi: 10.1002/jcb.21195.

Abstract

During mitosis, the phosphatidylinositol-3 (PI-3) family-related DNA damage checkpoint kinases ATM and ATR were found on the centrosomes of human cells. ATRIP, an interaction partner of ATR, as well as Chk1 and Chk2, the downstream targets of ATR or ATM, were also localized to the centrosomes. Surprisingly, the DNA-PK inhibitor vanillin enhanced the level of ATM on centrosomes. Accordingly, DNA-PKcs, the catalytic subunit of DNA-PK, was also found on the centrosomes. Vanillin altered the phosphorylation of Chk2 in the centrosomes and in whole cell extracts. Nucleoplasmic ATM co-immunoprecipitated with Ku70/86, the DNA binding subunits of DNA-PK, while vanillin diminished this association. Vanillin did not affect microtubule polymerization at the centrosomes but, surprisingly, caused a transient enhancement of alpha-tubulin foci in the nucleus. Interestingly, gamma-tubulin was also present in the nucleus and co-immunoprecipitated with ATR or BRCA1. DNA damage led to a reduction of the mentioned checkpoint proteins on the centrosomes but increased the level of gamma-tubulin at this organelle. Taken together, these results indicate that DNA damage checkpoint proteins may control the formation of gamma-tubulin and/or the kinetics of microtubule formation at the centrosomes, and thereby couple them to the DNA damage response.

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Animals
  • Ataxia Telangiectasia Mutated Proteins
  • BRCA1 Protein / metabolism
  • Benzaldehydes / metabolism
  • Cell Cycle Proteins / metabolism*
  • Cell Line
  • Centrosome / metabolism*
  • Checkpoint Kinase 1
  • Checkpoint Kinase 2
  • DNA Damage
  • DNA-Activated Protein Kinase / antagonists & inhibitors
  • DNA-Activated Protein Kinase / metabolism
  • DNA-Binding Proteins / metabolism*
  • Enzyme Inhibitors / metabolism
  • Exodeoxyribonucleases / metabolism*
  • Humans
  • Microtubules / metabolism
  • Phosphoproteins / metabolism*
  • Protein Kinases / metabolism
  • Protein Subunits / metabolism
  • Protein-Serine-Threonine Kinases / metabolism*
  • Tubulin / metabolism
  • Tumor Suppressor Proteins / metabolism*

Substances

  • ATRIP protein, human
  • Adaptor Proteins, Signal Transducing
  • BRCA1 Protein
  • BRCA1 protein, human
  • Benzaldehydes
  • Cell Cycle Proteins
  • DNA-Binding Proteins
  • Enzyme Inhibitors
  • Phosphoproteins
  • Protein Subunits
  • Tubulin
  • Tumor Suppressor Proteins
  • vanillin
  • Protein Kinases
  • Checkpoint Kinase 2
  • ATM protein, human
  • ATR protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • CHEK1 protein, human
  • CHEK2 protein, human
  • Checkpoint Kinase 1
  • DNA-Activated Protein Kinase
  • Protein-Serine-Threonine Kinases
  • Exodeoxyribonucleases
  • three prime repair exonuclease 1