Human cells contain a DNA-activated protein kinase that phosphorylates simian virus 40 T antigen, mouse p53, and the human Ku autoantigen.

Abstract

HeLa cells contain a serine/threonine protein kinase (DNA-PK) that is strongly activated in vitro by low concentrations of double-stranded DNA (dsDNA). Activation was specific for dsDNA; both natural DNAs and synthetic oligonucleotides functioned as kinase activators. The fact that DNA-PK activity was rapidly inhibited by incubation with dsDNA and ATP suggests that DNA-PK activity also may be regulated by autophosphorylation. During gel filtration, DNA-PK activity behaved as a 350-kDa protein, and highly purified DNA-PK contained a dsDNA-binding, 350-kDa polypeptide that was phosphorylated in a dsDNA-dependent manner. We conclude that this 350-kDa polypeptide is likely to be DNA-PK. Previously we showed that the dsDNA-activated kinase phosphorylates two threonines at the N terminus of hsp90 alpha (S. P. Lees-Miller and C. W. Anderson, J. Biol. Chem. 264:17275-17280, 1989). Here we show that DNA-PK also phosphorylates the simian virus 40 large tumor antigen, the mouse tumor-suppressor protein p53, the human Ku autoantigen, and two unidentified HeLa DNA-associated polypeptides of 52 and 110 kDa. Identification of these and other newly identified DNA-binding substrates suggest that the dsDNA-activated kinase may regulate transcription, DNA replication, or cell growth.

• Authors: S P Lees-Miller, Y R Chen, C W Anderson
• Journal: Molecular and cellular biology (Mol Cell Biol) Vol. 10 Issue 12 Pg. 6472-81 (Dec 1990) ISSN: 0270-7306 [Print] United States
• PMID: 2247067 (Publication Type: Journal Article, Research Support, U.S. Gov't, Non-P.H.S., Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Antigens, Nuclear
• Antigens, Polyomavirus Transforming
• Antigens, Surface
• DNA-Binding Proteins
• Tumor Suppressor Protein p53
• Protein Kinases
• DNA Helicases
• XRCC5 protein, human
• Xrcc6 protein, human
• Xrcc6 protein, mouse
• Ku Autoantigen

Topics

• Amino Acid Sequence
• Animals
• Antigens, Nuclear
• Antigens, Polyomavirus Transforming
• Antigens, Surface
• DNA Helicases
• DNA-Binding Proteins (metabolism)
• HeLa Cells (enzymology)
• Humans
• Kinetics
• Ku Autoantigen
• Mice
• Molecular Sequence Data
• Phosphorylation
• Protein Kinases (isolation & purification, metabolism)
• Substrate Specificity
• Tumor Suppressor Protein p53 (metabolism)