Degradation of ATM-independent decatenation checkpoint function in human cells is secondary to inactivation of p53 and correlated with chromosomal destabilization.
DNA topoisomerase II is required in the cell cycle to decatenate intertwined daughter chromatids prior to mitosis. To study the mechanisms that cells use to accomplish timely chromatid decatenation, the activity of a catenation-responsive checkpoint was monitored in human skin fibroblasts with inherited or acquired defects in the DNA damage G2 checkpoint. G2 delay was quantified shortly after a brief incubation with ICRF-193, which blocks the ability of topoisomerase II to decatenate chromatids, or treatment with ionizing radiation (IR), which damages DNA. Both treatments induced G2 delay in normal human fibroblasts. Ataxia telangiectasia fibroblasts with defective G2 checkpoint response to IR displayed normal G2 delay after treatment with ICRF-193, demonstrating that ATM kinase was not required for signaling when chromatid decatenation was blocked. The G2 delay induced by ICRF-193 was reversed by caffeine, indicating that active checkpoint signaling was involved. ICRF-193-induced G2 delay also was independent of p53 function, being evident in cells expressing HPV16E6 to inactivate p53. However, as fibroblasts expressing HPV16E6 aged in culture, they lost the ability to delay entry to mitosis, both after DNA damage and when decatenation was blocked. This age-related loss of G2 delay in response to ICRF-193 and IR in E6-expressing cells was blocked by induction of telomerase. Expression of telomerase also prevented chromosomal destabilization in aging E6-expressing cells. These observations lead to a new model of genetic instability, in which attenuation of G2 decatenatory checkpoint function permits cells to enter mitosis with insufficiently decatenated chromatids, leading to aneuploidy and polyploidy.
Kaufmann
William K
WK
CB7295, Rm 31-325, Lineberger Comprehensive Cancer Center, UNC-CH, Chapel Hill, North Carolina 27599-7295, USA. wkarlk@med.unc.edu
Campbell
Christine B
CB
Simpson
Dennis A
DA
Deming
Paula B
PB
Filatov
Leonid
L
Galloway
Denise A
DA
Zhao
Xiu J
XJ
Creighton
Andrew M
AM
Downes
C Stephen
CS
D064007:Ataxia Telangiectasia Mutated Proteins; D002110:Caffeine / Q000494:pharmacology; D002453:Cell Cycle / Q000502:physiology*; D018797:Cell Cycle Proteins; D002460:Cell Line; D002875:Chromosomes; D004249:DNA Damage; D004268:DNA-Binding Proteins; D054659:Diketopiperazines; D004305:Dose-Response Relationship, Drug; D004307:Dose-Response Relationship, Radiation; D005347:Fibroblasts / Q000378:metabolism; D016195:G2 Phase; D006801:Humans; D008938:Mitosis; D008954:Models, Biological; D010879:Piperazines / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D015398:Signal Transduction; D019098:Telomerase / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism* / Q000502:physiology; D025521:Tumor Suppressor Proteins
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D054659:Diketopiperazines; D010879:Piperazines; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; C070899:4,4'-(1,2-dimethyl-1,2-ethanediyl)bis-2,6-piperazinedione; D002110:Caffeine; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D019098:Telomerase