Expression and activity of DNA-dependent protein kinase in normal human leukocytes.
BACKGROUND
DNA-PK is a serine/threonine protein kinase that has been shown to play an important role in V(D)J recombination, Ig switching and DNA double strand break (DSB) repair. We have investigated DNA-PK in whole cell extracts from B and T lymphocytes, monocytes and granulocytes and the effects of phorbolmyristate acetate (PMA) and Concanavalin A (Con A) on DNA-PK in B and T lymphocytes.


METHODS
Lymphocytes were separated from human peripheral blood using the E-rosette technique and DNA-PK was analysed using Western blot, DNA-PK kinase activity assay and flow cytometry.


RESULTS
The levels of DNA-PK activity and content varied among the leukocytes. PMA and Con A stimulation consistently increased both DNA-PK activity and content in B lymphocytes as cells entered the S phase. Up-regulation could normally not be seen in T lymphocytes, although cells entered the S phase following PMA and Con A stimulation.


CONCLUSIONS
Mitogenic stimulation up-regulates DNA-PK activity and protein expression in B lymphocytes. T lymphocytes show interindividual differences in DNA-PK. Regulation of DNA-PK is not strictly cell cycle dependent for all cell types.
Eriksson
A
A
Department of Oncology and Pathology, Cancer Centre Karolinska, Karolinska Hospital, Stockholm, Sweden.
Lewensohn
R
R
Nilsson
A
A
D000328:Adult; D017209:Apoptosis; D001402:B-Lymphocytes / Q000187:drug effects / Q000201:enzymology; D001665:Binding Sites; D002453:Cell Cycle; D003208:Concanavalin A / Q000494:pharmacology; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins; D006098:Granulocytes / Q000201:enzymology; D006801:Humans; D007962:Leukocytes / Q000187:drug effects / Q000201:enzymology*; D008934:Mitogens / Q000494:pharmacology; D009000:Monocytes / Q000187:drug effects / Q000201:enzymology; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis / Q000378:metabolism*; D013601:T-Lymphocytes / Q000187:drug effects / Q000201:enzymology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D004268:DNA-Binding Proteins; D008934:Mitogens; D009687:Nuclear Proteins; D003208:Concanavalin A; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D013755:Tetradecanoylphorbol Acetate