Expression and DNA binding activity of the Ku heterodimer in bladder carcinoma.
BACKGROUND
The Ku protein is a tightly associated heterodimer, comprised of 70-kilodalton (kD) and 86-kD subunits, that forms the DNA-dependent protein kinase (DNA-PK) complex together with the 470-kD DNA-PKcs catalytic subunit, and is involved mainly in DNA double-strand breaks (DSBs) repair. The objective of the current study was to investigate the expression and DNA-binding activity of the Ku protein in fresh tissues from patients with bladder carcinoma and to compare it with that in nontumor tissues obtained from the same organ. Moreover, the DNA-binding activity of Ku was assessed after exposure of the tumor cells to 1 or 2 grays (Gy) of X-rays. Furthermore, the level of phosphorylated Ku was analyzed in both the nuclear and cytoplasmic compartment of normal tissue after exposure to 2 Gy of X-rays.


METHODS
The expression and DNA-binding activity of Ku protein were assessed in tumor samples from patients who all were diagnosed with transitional cell carcinoma (TCC) of the bladder using Western blot analysis and the electrophoretic mobility shift assay, respectively.


RESULTS
Enhanced Ku activity and expression were found in tumor tissue compared with normal tissue for each patient. Moreover, variations in Ku activity were found in a dose-dependent manner after the tumor cells were exposed to 1 or 2 Gy of X-rays. A decrease in phosphorylated Ku in the cytoplasm and a parallel increase in the nucleus of normal tissue cells were observed after exposure to X-rays.


CONCLUSIONS
The results of the current study suggest a possible role of Ku in regulating the DNA-PK activity of DSBs repair in bladder tumors.
Stronati
L
L
Section of Toxicology and Biomedical Sciences, ENEA CR Casaccia, Via Anguillarese 301, 00060 Rome, Italy.
Gensabella
G
G
Lamberti
C
C
Barattini
P
P
Frasca
D
D
Tanzarella
C
C
Giacobini
S
S
Toscano
M G
MG
Santacroce
C
C
Danesi
D T
DT
D000368:Aged; D000369:Aged, 80 and over; D034961:Antigens, Nuclear; D015153:Blotting, Western; D002295:Carcinoma, Transitional Cell / Q000235:genetics* / Q000473:pathology; D018736:DNA Adducts; D004265:DNA Helicases; D015342:DNA Probes; D004260:DNA Repair; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000096:biosynthesis*; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000528:radiation effects; D006801:Humans; D000072200:Ku Autoantigen; D008297:Male; D008875:Middle Aged; D009687:Nuclear Proteins / Q000096:biosynthesis*; D017346:Protein Serine-Threonine Kinases / Q000096:biosynthesis / Q000378:metabolism*; D011832:Radiation Injuries; D014407:Tumor Cells, Cultured; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000473:pathology
D034961:Antigens, Nuclear; D018736:DNA Adducts; D015342:DNA Probes; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen