Tumor specific modulation of KU70/80 DNA binding activity in breast and bladder human tumor biopsies

Abstract

The Ku70/80 heterodimer is the regulatory subunit of the DNA-dependent protein kinase (DNA-PK) and its DNAbinding activity mediates DNA double-strand breaks repair. Although Ku80 was recently proposed as a caretaker gene involved in the control of genome integrity, no data are available on Ku70/80 DNAbinding activity in human tumors. Heterodimer DNAbinding activity and protein expression were assayed by electrophoretic-mobility-shift-assay (EMSA) and Western blot analysis, in nuclear and cytoplasmic extracts from eight breast, seven bladder primary tumors and three metastatic nodes from breast cancers. Corresponding normal tissues of the same patients were used as controls. Ten out of 15 tumors showed nuclear Kubinding activity 3 ± 10 times higher than in the normal tissues, irrespective of bladder or breast origin. Conversely, in 5/15 primary tumors and in all the metastatic nodes analysed, nuclear Ku-activity was 1.5 ± 4.5-fold lower than in the corresponding normal tissues. Cytoplasmic heterodimer activity signi®cantly diered between tumor and normal tissues, displaying a 2 ± 10-fold increase in neoplastic tissues. Three dierent patterns combining both Ku expression and activity with tumor characteristics were identi®ed. In low aggressive breast tumors p70/p80 proteins were expressed in tumor but not in normal tissues. The heterodimer bindingactivity matched the protein levels. In non-invasive bladder carcinomas no signi®cant dierences in protein expression between tumor and the corresponding normal tissues were found, however heterodimer binding-activity was increased in tumor samples. In breast and bladder tumors, at the advanced stage and in node metastases, the binding activity was strongly reduced in tumor biopsies, however no dierences were demonstrated between normal and tumor protein levels. Our results suggest a dierent modulation of Ku70/80 DNA-binding activity in human neoplastic tissues, possibly related to tumor progression. Findings provide further data on tissue-speci®c protein expression and post-translational regulation of heterodimer activity. Oncogene (2001) 20, 739 ± 747.

The DNA repair complex Ku70/86 modulates Apaf1 expression upon DNA damage

“…37 In fact, as the impairment of DNA repair process produces genetic instability and, in turn, cancer development, it has been suggested that Ku's loss-of-function induces tumorigenesis. [38][39][40] Conversely, its gain-of-function has been associated with hyperproliferation and evading apoptosis of different tumour histotypes. 41,42 On the basis of the results obtained in this work, it could be speculated that the above-mentioned resistance phenomena are related to a Ku-dependent dysregulation of Apaf1 expression.…”

Are aneuploidy and chromosome breakage caused by a CINgle mechanism?

“…In contrast to a proposed repair defect, many CIN tumors show upregulation of DSB repair pathways such as non-homologous end joining. 31,32 A related theory proposed the existence of a number of fragile sites on the chromosome. 30 Due either to the lack of repair or to an opening of the chromatin during active transcription, the chromosome would be more prone to rupture at specific sites.…”

“…A continuous de novo generation of breaks, increasing in number with the complexity of translocations, explains why DSB repair is activated in many carcinomas. 31,32 Also telomerase appears to be involved in a general repair pathway that "heals" chromosomes after induction of DSB. 52 In accordance with the previous observation, telomerase reactivation in a hereditary breakage syndrome takes place only after cells have gone through crisis.…”

Centromere-localized breaks indicate the generation of DNA damage by the mitotic spindle

“…But even though mutation of DSB repair genes results in DNA damage after irradiation, little effect was found in untreated cells (5,6). In addition, sporadic carcinomas bear few if any mutations in DSB repair genes, and many tumor samples show augmented instead of reduced repair activity (7,8). The role of DSB repair in de novo generation of structural defects thus remains uncertain.…”