Degradation of ATM-independent checkpoint function in human cells is secondary to inactivation of p53 and correlated with chromosome destabilization

WK Kaufmann, CB Campbell, DA Simpson, PB Deming, L Filatov, DA Galloway, XJ Zhao, AM Creighton, Stephen Downes

    Research output: Contribution to journalArticle

    Abstract

    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.
    LanguageEnglish
    Pages210-219
    JournalCell Cycle
    Volume1
    Issue number3
    Publication statusPublished - May 2002

    Fingerprint

    Chromatids
    Chromosomes
    Fibroblasts
    Ionizing Radiation
    Mitosis
    DNA Damage
    Type II DNA Topoisomerase
    Telomerase
    Ataxia Telangiectasia
    Polyploidy
    Genetic Models
    Aneuploidy
    Caffeine
    Cell Cycle
    Phosphotransferases
    4,4'-(1,2-dimethyl-1,2-ethanediyl)bis-2,6-piperazinedione
    Skin

    Cite this

    Kaufmann, WK., Campbell, CB., Simpson, DA., Deming, PB., Filatov, L., Galloway, DA., ... Downes, S. (2002). Degradation of ATM-independent checkpoint function in human cells is secondary to inactivation of p53 and correlated with chromosome destabilization. Cell Cycle, 1(3), 210-219.
    Kaufmann, WK ; Campbell, CB ; Simpson, DA ; Deming, PB ; Filatov, L ; Galloway, DA ; Zhao, XJ ; Creighton, AM ; Downes, Stephen. / Degradation of ATM-independent checkpoint function in human cells is secondary to inactivation of p53 and correlated with chromosome destabilization. In: Cell Cycle. 2002 ; Vol. 1, No. 3. pp. 210-219.
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    abstract = "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.",
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    Kaufmann, WK, Campbell, CB, Simpson, DA, Deming, PB, Filatov, L, Galloway, DA, Zhao, XJ, Creighton, AM & Downes, S 2002, 'Degradation of ATM-independent checkpoint function in human cells is secondary to inactivation of p53 and correlated with chromosome destabilization', Cell Cycle, vol. 1, no. 3, pp. 210-219.

    Degradation of ATM-independent checkpoint function in human cells is secondary to inactivation of p53 and correlated with chromosome destabilization. / Kaufmann, WK; Campbell, CB; Simpson, DA; Deming, PB; Filatov, L; Galloway, DA; Zhao, XJ; Creighton, AM; Downes, Stephen.

    In: Cell Cycle, Vol. 1, No. 3, 05.2002, p. 210-219.

    Research output: Contribution to journalArticle

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    AU - Kaufmann, WK

    AU - Campbell, CB

    AU - Simpson, DA

    AU - Deming, PB

    AU - Filatov, L

    AU - Galloway, DA

    AU - Zhao, XJ

    AU - Creighton, AM

    AU - Downes, Stephen

    PY - 2002/5

    Y1 - 2002/5

    N2 - 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.

    AB - 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.

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