A mitotic topoisomerase II checkpoint in budding yeast is required for genome stability but acts independently of Pds1/securin.

Topoisomerase II (Topo II) performs topological modifications on double-stranded DNA molecules that are essential for chromosome condensation, resolution, and segregation. In mammals, G2 and metaphase cell cycle delays induced by Topo II poisons have been proposed to be the result of checkpoint activation in response to the catenation state of DNA. However, the apparent lack of such controls in model organisms has excluded genetic proof that Topo II checkpoints exist and are separable from the conventional DNA damage checkpoint controls. But here, we define a Topo II-dependent G2/M checkpoint in a genetically amenable eukaryote, budding yeast, and demonstrate that this checkpoint enhances cell survival. Conversely, a lack of the checkpoint results in aneuploidy. Neither DNA damage-responsive pathways nor Pds1/securin are needed for this checkpoint. Unusually, spindle assembly checkpoint components are required for the Topo II checkpoint, but checkpoint activation is not the result of failed chromosome biorientation or a lack of spindle tension. Thus, compromised Topo II function activates a yeast checkpoint system that operates by a novel mechanism.

[1]  R. Lock,et al.  Inhibition of p 34 cdc 2 Kinase Activity by Etoposide or Irradiation as a Mechanism of G 2 Arrest in Chinese Hamster Ovary Cells 1 , 2006 .

[2]  R. Müller,et al.  The Mitotic Spindle Checkpoint Is a Critical Determinant for Topoisomerase-based Chemotherapy* , 2005, Journal of Biological Chemistry.

[3]  U. Surana,et al.  DNA replication checkpoint prevents precocious chromosome segregation by regulating spindle behavior. , 2004, Molecular cell.

[4]  R. Margolis,et al.  Inhibition of DNA decatenation, but not DNA damage, arrests cells at metaphase. , 2004, Molecular cell.

[5]  A. Mikhailov,et al.  Topoisomerase II and histone deacetylase inhibitors delay the G2/M transition by triggering the p38 MAPK checkpoint pathway , 2004, The Journal of cell biology.

[6]  K. Yanagisawa,et al.  Identification of Decatenation G2 Checkpoint Impairment Independently of DNA Damage G2 Checkpoint in Human Lung Cancer Cell Lines , 2004, Cancer Research.

[7]  H. Koyama,et al.  Genetic evidence for involvement of two distinct nonhomologous end-joining pathways in repair of topoisomerase II-mediated DNA damage. , 2004, Biochemical and biophysical research communications.

[8]  J. R. Daum,et al.  Differential Kinetochore Requirements for Establishment and Maintenance of the Spindle Checkpoint Are Dependent on the Mechanism of Checkpoint Activation in Saccharomyces cerevisiae , 2004, Cell cycle.

[9]  W. Earnshaw,et al.  Analysis of Scc1‐deficient cells defines a key metaphase role of vertebrate cohesin in linking sister kinetochores , 2004, EMBO reports.

[10]  K. Yanagisawa,et al.  Identification of Decatenation G 2 Checkpoint Impairment Independently of DNA Damage G 2 Checkpoint in Human Lung Cancer Cell Lines , 2004 .

[11]  D. J. Clarke,et al.  S-phase checkpoint controls mitosis via an APC-independent Cdc20p function , 2003, Nature Cell Biology.

[12]  S. Mateos,et al.  DNA strand breaks induced by the anti-topoisomerase II bis-dioxopiperazine ICRF-193. , 2003, Mutation research.

[13]  R. Rothstein,et al.  Cell Cycle-Regulated Centers of DNA Double-Strand Break Repair , 2003, Cell cycle.

[14]  D. J. Clarke,et al.  Replication-Coupled Topoisomerase II Templates the Mitotic Chromosome Scaffold? , 2003, Cell cycle.

[15]  A. Mikhailov,et al.  DNA Damage during Mitosis in Human Cells Delays the Metaphase/Anaphase Transition via the Spindle-Assembly Checkpoint , 2002, Current Biology.

[16]  Kim Nasmyth,et al.  Segregating Sister Genomes: The Molecular Biology of Chromosome Separation , 2002, Science.

[17]  S. Elledge,et al.  The SUMO-1 isopeptidase Smt4 is linked to centromeric cohesion through SUMO-1 modification of DNA topoisomerase II. , 2002, Molecular cell.

[18]  D. J. Clarke,et al.  A Topoisomerase II-Dependent Checkpoint in G2-Phase Plant Cells Can Be Bypassed by Ectopic Expression of Mitotic Cyclin B2 , 2002, Cell cycle.

[19]  Steven B Haase,et al.  Improved Flow Cytometric Analysis of the Budding Yeast Cell Cycle , 2002, Cell cycle.

[20]  S. Mateos,et al.  Testing the SCE mechanism with non-poisoning topoisomerase II inhibitors. , 2001, Mutation research.

[21]  R. Paules,et al.  The human decatenation checkpoint , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[22]  D. J. Clarke,et al.  Mec1p regulates Pds1p levels in S phase: complex coordination of DNA replication and mitosis , 2001, Nature Cell Biology.

[23]  J. Piette,et al.  Ku antigen is required to relieve G2 arrest caused by inhibition of DNA topoisomerase II activity by the bisdioxopiperazine ICRF-193 , 2001, Oncogene.

[24]  J. Roca,et al.  Circular Minichromosomes Become Highly Recombinogenic in Topoisomerase-deficient Yeast Cells* , 2001, The Journal of Biological Chemistry.

[25]  D. J. Clarke,et al.  A Novel Role of the Budding Yeast Separin Esp1 in Anaphase Spindle Elongation , 2001, The Journal of cell biology.

[26]  D. Burke,et al.  The spindle checkpoint: two transitions, two pathways. , 2000, Trends in cell biology.

[27]  G. Goshima,et al.  Establishing Biorientation Occurs with Precocious Separation of the Sister Kinetochores, but Not the Arms, in the Early Spindle of Budding Yeast , 2000, Cell.

[28]  D. J. Clarke,et al.  Checkpoints controlling mitosis , 2000, BioEssays : news and reviews in molecular, cellular and developmental biology.

[29]  D. J. Clarke,et al.  Premitotic chromosome individualization in mammalian cells depends on topoisomerase II activity , 2000, Chromosoma.

[30]  D. J. Clarke,et al.  The Pds1 anaphase inhibitor and Mec1 kinase define distinct checkpoints coupling S phase with mitosis in budding yeast , 1999, Current Biology.

[31]  M. Hoyt,et al.  Dominant alleles of Saccharomyces cerevisiae CDC20 reveal its role in promoting anaphase. , 1998, Genetics.

[32]  Anthony L. Johnson,et al.  Budding yeast RSI1/APC2, a novel gene necessary for initiation of anaphase, encodes an APC subunit , 1998, The EMBO journal.

[33]  J W Sedat,et al.  Mitosis in living budding yeast: anaphase A but no metaphase plate. , 1997, Science.

[34]  Andrew W. Murray,et al.  GFP tagging of budding yeast chromosomes reveals that protein–protein interactions can mediate sister chromatid cohesion , 1996, Current Biology.

[35]  D. J. Clarke,et al.  A postprophase topoisomerase II-dependent chromatid core separation step in the formation of metaphase chromosomes , 1995, The Journal of cell biology.

[36]  D N Mastronarde,et al.  Three-dimensional ultrastructural analysis of the Saccharomyces cerevisiae mitotic spindle , 1995, The Journal of cell biology.

[37]  D. J. Clarke,et al.  A topoisomerase II-dependent G2 cycle checkpoint in mammalian cells , 1994, Nature.

[38]  P. Philippsen,et al.  New heterologous modules for classical or PCR‐based gene disruptions in Saccharomyces cerevisiae , 1994, Yeast.

[39]  D. J. Clarke,et al.  A topoisomerase II-dependent G2 cycle checkpoint in mammalian cells , 1994, Nature.

[40]  D. J. Clarke,et al.  Topoisomerase II inhibition prevents anaphase chromatid segregation in mammalian cells independently of the generation of DNA strand breaks. , 1993, Journal of cell science.

[41]  A. Murray,et al.  Sister chromatid separation in frog egg extracts requires DNA topoisomerase II activity during anaphase , 1992, The Journal of cell biology.

[42]  T. Mitchison,et al.  Cell cycle control of higher-order chromatin assembly around naked DNA in vitro , 1991, The Journal of cell biology.

[43]  T. Andoh,et al.  Inhibition of topoisomerase II by antitumor agents bis(2,6-dioxopiperazine) derivatives. , 1991, Cancer research.

[44]  P. Cook.  The nucleoskeleton and the topology of replication , 1991, Cell.

[45]  K. Deisseroth,et al.  Cross-resistance of an amsacrine-resistant human leukemia line to topoisomerase II reactive DNA intercalating agents. Evidence for two topoisomerase II directed drug actions. , 1991, Biochemistry.

[46]  U. K. Laemmli,et al.  Chromosome assembly in vitro: Topoisomerase II is required for condensation , 1991, Cell.

[47]  W. Earnshaw,et al.  Mitotic chromatin condensation in vitro using somatic cell extracts and nuclei with variable levels of endogenous topoisomerase II , 1990, The Journal of cell biology.

[48]  R. Hancock,et al.  DNA topoisomerase II is required for formation of mitotic chromosomes in Chinese hamster ovary cells: studies using the inhibitor 4'-demethylepipodophyllotoxin 9-(4,6-O-thenylidene-beta-D-glucopyranoside). , 1990, Biochemistry.

[49]  R. Lock,et al.  Inhibition of p34cdc2 kinase activity by etoposide or irradiation as a mechanism of G2 arrest in Chinese hamster ovary cells. , 1990, Cancer research.

[50]  Curt Wittenberg,et al.  An essential G1 function for cyclin-like proteins in yeast , 1989, Cell.

[51]  R. McCarroll,et al.  Time of replication of yeast centromeres and telomeres , 1988, Cell.

[52]  L. Hartwell,et al.  The RAD9 gene controls the cell cycle response to DNA damage in Saccharomyces cerevisiae. , 1988, Science.

[53]  J. Newport.  Nuclear reconstitution in vitro: Stages of assembly around protein-free DNA , 1987, Cell.

[54]  J. Wang,et al.  Yeast DNA Topoisomerases and Their Structural Genes , 1987 .

[55]  T. Uemura,et al.  The nucleotide sequence of the fission yeast DNA topoisomerase II gene: structural and functional relationships to other DNA topoisomerases. , 1986, The EMBO journal.

[56]  N. Cozzarelli,et al.  Biochemical topology: applications to DNA recombination and replication. , 1986, Science.

[57]  D. Botstein,et al.  DNA topoisomerase II is required at the time of mitosis in yeast , 1985, Cell.

[58]  W. Earnshaw,et al.  Topoisomerase II is a structural component of mitotic chromosome scaffolds , 1985, The Journal of cell biology.

[59]  T. Uemura,et al.  Isolation of type I and II DNA topoisomerase mutants from fission yeast: single and double mutants show different phenotypes in cell growth and chromatin organization. , 1984, The EMBO journal.

[60]  R. Sternglanz,et al.  DNA topoisomerase II mutant of Saccharomyces cerevisiae: topoisomerase II is required for segregation of daughter molecules at the termination of DNA replication. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[61]  J. Ducore,et al.  Effects of the epipodophyllotoxin VP-16-213 on cell cycle traverse, DNA synthesis, and DNA strand size in cultures of human leukemic lymphoblasts. , 1983, Cancer research.

[62]  A. Varshavsky,et al.  Arrest of segregation leads to accumulation of highly intertwined catenated dimers: Dissection of the final stages of SV40 DNA replication , 1981, Cell.

[63]  A. M. Creighton,et al.  The effect of bisdioxopiperazines on the synthesis of deoxyribonucleic acid, ribonucleic acid and protein in growing mouse-embryo fibroblasts. , 1969, The Biochemical journal.