DNA end binding activity and Ku70/80 heterodimer expression in human colorectal tumor.
OBJECTIVE
To determine the DNA binding activity and protein levels of the Ku70/80 heterodimer, the functional mediator of the NHEJ activity, in human colorectal carcinogenesis.


METHODS
The Ku70/80 DNA-binding activity was determined by electrophoretic mobility shift assays in 20 colon adenoma and 15 colorectal cancer samples as well as matched normal colonic tissues. Nuclear and cytoplasmic protein expression was determined by immunohistochemistry and Western blot analysis.


RESULTS
A statistically significant difference was found in both adenomas and carcinomas as compared to matched normal colonic mucosa (P<0.00). However, changes in binding activity were not homogenous with approximately 50% of the tumors showing a clear increase in the binding activity, 30% displaying a modest increase and 15% showing a decrease of the activity. Tumors, with increased DNA-binding activity, also showed a statistically significant increase in Ku70 and Ku86 nuclear expression, as determined by Western blot and immunohistochemical analyses (P<0.001). Cytoplasmic protein expression was found in pathological samples, but not in normal tissues either from tumor patients or from healthy subjects.


CONCLUSIONS
Overall, our DNA-binding activity and protein level are consistent with a substantial activation of the NHEJ pathway in colorectal tumors. Since the NHEJ is an error prone mechanism, its abnormal activation can result in chromosomal instability and ultimately lead to tumorigenesis.
Mazzarelli
Paola
P
Laboratory of Molecular Medicine and Biotechnology, Università Campus Bio-Medico, Via Longoni, 83, Rome 00155, Italy.
Parrella
Paola
P
Seripa
Davide
D
Signori
Emanuela
E
Perrone
Giuseppe
G
Rabitti
Carla
C
Borzomati
Domenico
D
Gabbrielli
Armando
A
Matera
Maria Giovanna
MG
Gravina
Carolina
C
Caricato
Marco
M
Poeta
Maria Luana
ML
Rinaldi
Monica
M
Valeri
Sergio
S
Coppola
Roberto
R
Fazio
Vito Michele
VM
D000236:Adenoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D034961:Antigens, Nuclear / Q000737:chemistry / Q000378:metabolism*; D002277:Carcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D015179:Colorectal Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D004247:DNA / Q000378:metabolism*; D051747:DNA-Activated Protein Kinase / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D019281:Dimerization; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D000072200:Ku Autoantigen; D008297:Male; D008875:Middle Aged; D009687:Nuclear Proteins / Q000378:metabolism
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004247:DNA; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen