Skip to main content Accessibility help

Effect of inhibitors of DNA replication on early zebrafish embryos: evidence for coordinate activation of multiple intrinsic cell-cycle checkpoints at the mid-blastula transition

  • Richard Ikegami (a1), Alma K. Rivera-Bennetts (a1), Deborah L. Brooker (a1) and Thomas D. Yager (a1)


We address the developmental activation, in the zebrafish embryo, of intrinsic cell-cycle checkpoints which monitor the DNA replication process and progression through the cell cycle. Eukaryotic DNA replication is probably carried out by a multiprotein complex containing numerous enzymes and accessory factors that act in concert to effect processive DNA synthesis (Applegren, N. et al. (1995) J. Cell. Biochem. 59, 91–107). We have exposed early zebrafish embryos to three chemical agents which are predicted to specifically inhibit the DNA polymerase α, topoisomerase I and topoisomerase II components of the DNA replication complex. We present four findings: (1) Before mid-blastula transition (MBT) an inhibition of DNA synthesis does not block cells from attempting to proceed through mitosis, implying the lack of functional checkpoints. (2) After MBT, the embryo displays two distinct modes of intrinsic checkpoint operation. One mode is a rapid and complete stop of cell division, and the other is an ‘adaptive’ response in which the cell cycle continues to operate, perhaps in a ‘repair’ mode, to generate daughter nuclei with few visible defects. (3) The embryo does not display a maximal capability for the ‘adaptive’ response until several hours after MBT, which is consistent with a slow rranscriptional control mechanism for checkpoint activation. (4) The slow activation of checkpoints at MBT provides a window of time during which inhibitors of DNA synthesis will induce cytogenetic lesions without killing the embryo. This could be useful in the design of a deletion-mutagenesis strategy.


Corresponding author

T.D. Yager, Division of Developmental Biology and Research Institute, Hospital for Sick Children, Toronto, Ontario, Canada M59 1X8.


Hide All
Adachi, Y. & Laemmli, U.K. (1992). Identification of nuclear pre-replication centers poised for DNA synthesis in Xenopus egg extracts: immunolocalization study of replication protein. A. J. Cell Biol. 119, 115.
Alexander, S.P. & Rieder, C.L. (1991). Chromosome motion during attachment to the vertebrate spindle: initial salta-tory-like behavior of chromosomes and quantitative analysis of force production by nascent kinetochore fibers. J. Cell Biol. 113, 805–15.
Applegren, N., Hickey, R.J., Kleinschmidt, A.M., Zhou, Q., Coll, J., Wills, P., Swaby, R., Wei, Y., Quan, J.Y. & Lee, M.Y. (1995). Further characterization of the human cell multi-protein DNA replication complex. J. Cell Biochem. 59, 91107.
Araki, H., Leem, S.-H., Phongdara, A. & Sugino, A. (1995). Dpb11, which interacts with DNA polymerase II (e) in Saccharomyces cerevisiae, has a dual role in S-phase progression and at a cell cycle checkpoint. Proc. Natl. Acad. Sci. USA 92, 11791–5.
Blaustein, A.R., Hoffman, P.D., Hokit, D.G., Kiesecher, J.M., Walls, S.C. & Hays, J.B. (1994). UV repair and resistance to solar UV-B in amphibian eggs: a link to population declines? Proc. Natl. Acad. Sci. USA 91, 1791–5.
Bryant, P.J., Watson, K.J., Justice, R.W. & Woods, D.F. (1993). Tumor suppressor genes encoding proteins required for cell interactions and signal transduction in Drosophila. Development (Suppl)., 239–49.
Buschman, E., Lepage, P. & Gros, P. (1994). P-glycoprotein homologues. Cancer Treat. Res. 73, 1739.
Chaudhary, N. & Courvalin, J.C. (1993). Stepwise reassembly of the nuclear envelope at the end of mitosis. J. Cell Biol. 122, 295306.
Chong, J.P.J., Thommes, P. & Blow, J.J. (1996). The role of MCM/P1 proteins in the licensing of DNA replication. Trends Biochem. Sci. 21, 102–6.
Cochet-Meilhac, M. & Chambon, P. (1974). Animal DNA-dependent RNA polymerases. II. Mechanism of the inhibition of RNA polymerases B by amatoxins. Biochim. Biophys. Acta 353, 160–84.
Collodi, P., Miranda, C, Zhao, X., Buhler, D.R. & Barnes, D.W. (1994). Induction of zebrafish hepatic cytochrome P450 in vivo and in cell culture. Xenobiotica 24, 487–93.
Daga, R.R., Thode, G. & Amores, A. (1996). Chromosome complement, C-banding, Ag-NOR and replication banding in the zebrafish Danio rerio. Chromosome Res. 4, 2932.
Darzynkiewicz, Z. (1995). Apoptosis in anti-tumor strategies: modulation of cell cycle or differentiation. J. Cell. Biochem. 58, 151–9.
Dasso, M. & Newport, J.W. (1990). Completion of DNA replication is monitored by a feedback system that controls the initiation of mitosis in vitro: studies in Xenopus. Cell 61, 811–23.
Del Bino, G., Skierski, J.S. & Darzynkiewicz, Z. (1991). The concentration-dependent diversity of effects of DNA topoisomerase I and II inhibitors on the cell cycle of HL-60 cells. Exp. Cell Res. 195, 485–91.
Devlin, R.H., McNeil, B.K., Solar, I.I. & Donaldson, E.M. (1994). A rapid PCR-based test for Y-chromosomal DNA allows simple production of all-female strains of chinook salmon. Aquaculture 128, 211–20.
DiNardo, S., Voelkel, K. & Sternglanz, R. (1984). DNA topoisomerase II mutant of Saccharomyces cerevisiae: topoisomerase II is required for segregation of daughter molecules at the termination of DNA replication. Proc. Natl. Acad. Sci. USA 81, 2616–20.
Downes, C.S., Clarke, D.J., Mullinger, A.M., Gimenez-Abian, J.F., Creighton, A.M. & Johnson, R.T. (1994). A topoisomerase II dependent G2 cycle checkpoint in mammalian cells. Nature 372, 467–70.
Driever, W., Solnica-Krezel, L., Schier, A.F., Neuhauss, S.C.F., Malicki, J., Stemple, D.L., Stanier, D.Y.R., Zwart-kruis, F., Abdelilah, S., Rangini, Z., Belak, J. & Boggs, C. (1996). A genetic screen for mutations affecting embryo-genesis in zebrafish. Development 123, 3746.
Du, S.J., Devlin, R.H. & Hew, C.L. (1993). Genomic structure of growth hormone genes in chinook salmon (Onco-rhynchus tshawytscha): presence of two functional genes, GH-I and GH-II, and a male-specific pseudogene GH-psi. DNA Cell Biol. 12, 739–51.
Edgar, B.A. & Datar, S.A. (1996). Zygotic degradation of two maternal cdc25 mRNAs terminates Drosophila's early cell cycle program. Genes Dev. 10, 1966–77.
Felsenfeld, A.L. (1996). Defining the boundaries of zebrafish developmental genetics. Nature Genet. 14, 258–63.
Forbes, S.H., Knudsen, K.L., North, T.W. & Allendorf, F.W. (1994). One of two growth hormone genes in coho salmon is sex-linked. Proc. Natl. Acad. Sci. USA 91, 1628–31.
Francesconi, S., De Recondo, A.M. & Baldacci, G. (1995). DNA polymerase delta is required for the replication feedback control of cell-cycle progression in Schizo-saccharomyces pombe. Mol. Gen. Genet. 246, 561–9.
Friedberg, E.C., Walker, G.C. & Siede, W. (eds.) (1995). DNA Repair and Mutagenesis. Washington, DC: ASM Press.
Fritz, A., Rozowski, M., Walker, C. & Westerfield, M. (1996). Identification of selected gamma-ray induced deficiencies in zebrafish using multiplex polymerase chain reaction. Genetics 144, 1735–45.
Gaiano, N., Amsterdam, A., Kawakami, K., Allende, M., Becker, T. & Hopkins, N. (1996). Insertional mutagenesis and rapid cloning of essential genes in zebrafish. Nature 383, 829–32.
Gasser, S.M., Laroche, T., Falquet, J., Boy de la Tour, E. & Laemmli, U.K. (1986). Metaphase chromosome structure involvement of topoisomerase II. J. Mol. Biol. 188, 613–29.
Glover, T.W. & Stein, C.K. (1988). Chromosome breakage and recombination at fragile sites. Am. J. Hum. Genet. 43, 265–73.
Gromova, I.I., Thomsen, B. & Razin, S.V. (1995). Different topoisomerase II antitumor drugs direct similar specific long-range fragmentation of an amplified c-MYC gene locus in living cells and in high-salt-extracted nuclei. Proc. Natl. Acad. Sci. USA 92, 102–6.
Haaf, T. & Schmid, M. (1991). Chromosome topology in mammalian interphase nuclei. Exp. Cell Res. 192, 325–32.
Haffter, P., Granato, M., Brand, M., Mullins, M.C., Ham-merschmidt, M., Kane, D.A., Odenthal, J., van Eeden, F.J.M., Jiang, Y.-J., Heisenberg, C.-P, Kelsh, R.N., Furu-tani-Seiki, M., Vogelsang, E., Beuchle, D., Schach, U., Fabian, C. & Nusslein-Volhard, C. (1996). The identification of genes with unique and essential functions in the development of the zebrafish, Danio rerio. Development 123, 136.
Hally, M.K., Rasch, E.M., Mainwaring, H.R. & Bruce, R.C. (1986). Cytophotometric evidence of variation in genome size of desmognathine salamanders. Histochemistry 85, 185–92.
Hartwell, L.H. & Kastan, M.B. (1994). Cell-cycle control and cancer. Science 266, 1821–8.
Heichman, K. A. & Roberts, J.M. (1994). Rules to replicate by. Cell 79, 557–62.
Hinegardner, R. & Rosen, D.E. (1972). Cellular DNA content and the evolution of teleostean fishes. Am. Nat. 166, 621–44.
Hofferer, L., Winterhalter, K.H. & Althaus, F.R. (1995). Xenopus eggs lysates repair heat-generated DNA nicks with an average patch size of 36 nucleotides. Nucleic Acids Res. 23, 1396–7.
Horiguchi, T. & Tanida, S. (1995). Rescue of Schizosaccharo-myces pombe from camptothecin-mediated death by a DNA topoisomerase I inhibitor, TAN-1518 A. Biochem. Pharmacol. 49, 1395–401.
Howe, J.A. & Newport, J.W. (1996). A developmental timer regulates degradation of cyclin E1 at the midblastula transition during Xenopus embryogenesis. Proc. Natl. Acad. Sci. USA 93, 2060–4.
Iarovaia, O., Hancock, R., Lagarkova, M., Miassod, R. & Razin, S.V. (1996). Mapping of genomic DNA loop organization in a 500-kilobase region of the Drosophila X chromosome by the topoisomerase II-mediated DNA loop excision protocol. Mol. Cell. Biol. 16, 302–8.
Johnson, S.L., Gates, M.A., Johnson, M., Talbot, W.S., Home, S., Baik, K., Rude, S., Wong, J.R. & Postlethwait, J.H. (1996). Centromere-linkage analysis and consolidation of the zebrafish genetic map. Genetics 142, 1277–88.
Kane, D.A. & Kimmel, C.B. (1993). The zebrafish midblastula transition. Development 119, 447–56.
Kane, D.A., Maischein, H.M., Brand, M., van Eeden, F.J., Furutani-Seiki, M., Granato, M., Haffter, P., Hammer-schmidt, M., Heisenberg, C.P, Jiang, Y.T., Kelsh, R.N., Mullins, M.C., Odenthal, J., Warga, R.M. & Nusslein-Volhard, C. (1996). The zebrafish early arrest mutants. Development 123, 5766.
Kane, D.A., Warga, R.M. & Kimmel, C.B. (1992). Mitotic domains in the early embryo of the zebrafish. Nature 360, 735–7.
Kim, R.A. & Wang, J.C. (1989). Function of DNA topoiso-merases as replication swivels in Saccharomyces cerevisiae. J. Mol. Biol. 208, 257–67.
Kimmel, C.B., Ballard, W.W., Kimmel, S.R., Ullmann, B. & Schilling, T.F. (1995). Stages of embryonic development of the zebrafish. Dev. Dynam. 203, 253310.
King, R.W., Peters, J.M., Tugendreich, A., Rolfe, M., Hieter, P. & Kirschner, M.W. (1995). A 20S complex containing CDC27 and CDC16 catalyzes the mitosis-specific conjugation of ubiquitin to cyclin B. Cell 81, 279–88.
Knapik, E.W., Goodman, A., Atkinson, S.O., Roberts, C.T., Shiozawa, M., Sim, C.U., Weksler-Zangen, S., Trolliet, M.R., Futrell, C, Innes, B.A., Koike, G., McLaughlin, M.G., Pierre, L, Simon, J.S., Vilallonga, E., Roy, M., Chiang, P.-W., Fishman, M.C., Driever, W. & Jacob, H.J. (1996). A reference cross DNA panel for zebrafish (Danio rerio) anchored with simple sequence length polymorphisms. Development 123, 451–60.
Kumagai, A. & Dunphy, W.G. (1995). Control of the Cdc2/ cyclin B complex in Xenopus egg extracts arrested at a G2/ M checkpoint with DNA synthesis inhibitors. Mol. Biol. Cell 6, 199213.
Lam, W.L., Lee, T.-S. & Gilbert, W. (1996). Active transposition in zebrafish. Proc. Natl. Acad. Sci. USA 93, 10870–5.
Lee, S.-H., Kim, D.-K. & Drissi, R. (1995). Human xeroderma pigmentosum Group A protein interacts with human replication protein A and inhibits DNA replication. J. Biol. Chem. 270, 21800–5.
Lehman, C.W., Jeong-Yu, S., Trautman, J.K. & Carroll, D. (1994). Repair of heteroduplex DNA in Xenopus laevis oocytes. Genetics 138, 459–70.
Li, J.J. & Deshaies, R.J. (1993). Exercising self-restraint: discouraging illicit acts of S and M in eukaryotes. Cell 74, 223–6.
Li, C.J., Averboukh, L. & Pardee, A.B. (1993 a). β-Lapachone, a novel DNA topoisomerase I inhibitor with a mode of action different from camptothecin. J. Biol. Chem. 268, 22463–8.
Li, C, Cao, L.-G., Wand, Y.-L. & Baril, E.F. (1993 b). Further purification and characterization of a multienzyme complex for DNA synthesis in human cells. J. Cell. Biochem. 53, 405–19.
Lincke, C.R., Broeks, A., The, I., Plasterk, R.H.A. & Borst, P. (1993). The expression of two P-glycoprotein (pgp) genes in transgenic Caenorhabditis elegans is confined to intestinal cells. EMBO J. 12, 1615–20.
Malkas, L.H., Hickey, R.J., Li, C, Pedersen, N. & Baril, E.F. (1990). A 21S enzyme complex from HeLa cells that functions in Simian Virus 40 DNA replication in vitro. Biochemistry 29, 6362–74.
Margolis, R.L. & Andreassen, P.R. (1993). The telophase disc: its possible role in mammalian cell cleavage. Bio-Essays 15, 201–7.
Matsumoto, Y, Kim, K. & Bogenhagen, D.F. (1994). Proliferating cell nuclear antigen-dependent abasic site repair in Xenopus laevis oocytes: an alternative pathway of base excision DNA repair. Mol. Cell. Biol. 14, 6187–97.
Mullins, M.C., Hammerschmidt, M, Haffter, P. & Nusselin-Volhard, C. (1994). Large-scale mutagenesis in the zebra-fish: in search of genes controlling development in a vertebrate. Curr. Biol. 4, 189202.
Murakami, H. & Okayama, H. (1995). A kinase from fission yeast responsible for blocking mitosis in S phase. Nature 374, 817–19.
Nagano, H., Hirai, S., Okano, K. & Ikegami, S. (1981). Achromosomal cleavage of fertilization starfish eggs in the presence of aphidicolin. Dev. Biol. 85, 409–15.
Navas, T.A., Zhou, Z. & Elledge, S.J. (1995). DNA poly-merase e links the DNA replication machinery to the S phase checkpoint. Cell 80, 2939.
Nelson, D.R., Kamataki, T., Waxman, D.J., Gugngerich, F.P., Estabrook, R.W., Feyereisen, R., Gonzalez, F.J., Coon, M.J., Gunsalus, I.C., Gotoh, O., et al. (1993). The P450 super-family: update on new sequences, gene mapping, accession numbers, early trivial names of enzymes, and nomenclature. DNA Cell Biol. 12, 151.
Neter, J., Wasserman, W. & Whitmore, G.A. (1978). Applied Statistics. Boston: Allyn and Bacon.
Newport, J. & Dasso, M. (1989). On the coupling between DNA replication and mitosis. J. Cell Sci. (Suppl.) 12, 149–60.
Nicklas, R.B. (1988). The forces that move chromosomes in mitosis. Annu. Rev. Biophys. Biophys. Chem. 17, 431–49.
Nurse, P. (1994). Ordering S phase and M phase in the cell cycle. Cell 79, 547–50.
Patel, N.H., Martin-Bianco, E., Coleman, K.G., Poole, S.J., Ellis, M.C., Kornberg, T.B. & Goodman, C.S. (1989). Expression of engrailed proteins in arthropods, annelids and chordates. Cell 58, 955–68.
Paulovich, A.G. & Hartwell, L.H. (1995). A checkpoint regulates the rate of progression through S phase in S. cerevisiae in response to DNA damage. Cell 82, 841–7.
Pijnacker, L.P. & Ferwerda, M.A. (1995). Zebrafish chromosome banding. Genome 38, 1052–5.
Pommier, Y. (1996). Eukaryotic DNA topoisomerase I: genome gatekeeper and its intruders, camptothecins. Semin. Oncol. 23, 310.
Raff, J.W. & Glover, D.M. (1988). Nuclear and cytoplasmic mitotic cycles continue in Drosophila embryos in which DNA synthesis is inhibited with aphidicolin. J. Cell Biol. 107, 2009–19.
Raff, M.C. (1992). Social controls on cell survival and cell death. Nature 356, 397400.
Raff, M.C. (1996). Size control: the regulation of cell numbers in animal development. Cell 86, 173–5.
Razin, S.V., Petrov, P. & Hancock, R. (1991). Precise localization of the alpha-globin gene cluster within one of the 20-to 300-kilobase DNA fragments released by cleavage of chicken chromosomal DNA at topoisomerase II sites in vivo: evidence that the fragments are DNA loops or domains. Proc. Natl. Acad. Sci. USA 88, 8515–19.
Richardson, H.E., O'Keefe, L.V., Reed, S.I. & Saint, R. (1993). A Drosophila G1-specific cyclin E homolog exhibits different modes of expression during embryogenesis. Development 119, 673–90.
Rieder, C.L., Cole, R.W., Khodjakov, A. & Sluder, G. (1995). The checkpoint delaying anaphase in response to chromosome monoorientation is mediated by inhibitory signal produced by unattached kinetochores. J. Cell Biol. 130, 941–8.
Romanowski, P. & Madine, M.A. (1996). Mechanisms restricting DNA replication to one per cell cycle: MCMs, pre-replicative complexes and kinases. Trends Cell Biol. 6, 184–8.
Saiki, T, Kyozuka, K., Osanai, K. & Hamaguchi, Y (1991). Chromosomal behavior in starfish (Asterina pectinifera) zygotes under the effect of aphidicolin, an inhibitor of DNA polymerase. Exp. Cell Res. 192, 380–8.
Sanchez, Y. & Elledge, S.J. (1995). Stopped for repairs. Bioessays 17, 545–8.
Schinkel, A.H., Smit, J.J., van Tellingen, O., Beijnen, J.H., Wagenaar, E., van Deemter, L., Mol, C.A., van der Valk, M.A., Robanus-Maandag, E.C., te Riele, H.P., et al. (1994). Disruption of the mouse mdrla P-glycoprotein gene leads to a deficiency in the blood-brain barrier and to increased sensitivity to drugs. Cell 77, 491502.
Shamu, C.E. & Murray, A.W. (1992). Sister chromatid separation in frog egg extracts requires DNA topoisomerase II activity during anaphase. J. Cell Biol. 117, 921–34.
Sheaff, R. & Kuchta, R. (1991). Mechanism of DNA polymerase alpha inhibition by aphidicolin. Biochemistry 30, 8590–7.
Sheldrick, K.S. & Carr, A.M. (1993). Feedback controls and G2 checkpoints: fission yeast as a model system. Bioessays 15, 775–81.
Shima, A. & Shimada, A. (1991). Development of a possible nonmammalian test system for radiation-induced germ-cell mutagenesis using a fish, the Japanese medaka (Oryzias latipes). Proc. Natl. Acad. Sci. USA 88, 2545–9.
Shima, A. & Shimada, A. (1994). The Japanese medaka, Oryzias latipes, as a new model organism for studying environmental germ-cell mutagenesis. Environ. Health Perspect. 102 (Suppl. 12), 33–5.
Shimizu, T. (1994). The prevention of smaller blastomeres of early Tubifex embryos from entering mitosis by unrepli-cated DNA. Dev. Biol. 161, 274–8.
Shivji, M.K., Grey, S.J., Strausfeld, U.P., Wood, R.D. & Blow, J.J. (1994). Cipl inhibits DNA replication but not PCNA-dependent nucleotide excision-repair. Curr. Biol. 4, 1062–8.
Simbulan-Rosenthal, C.M., Rosenthal, D.S., Hilz, H., Hickey, R., Malkas, L., Applegren, N., Wu, Y, Bers, G. & Smulson, M.E. (1996). The expression of poly(ADP-ribose) poly-merase during differentiation-linked DNA replication reveals that it is a component of the multiprotein DNA replication complex. Biochemistry 35, 11622–33.
Sluder, G. & Lewis, K. (1987). Relationship between nuclear DNA synthesis and centrosome reproduction in sea urchin eggs. J. Exp. Zool. 244, 89100.
Smythe, C. & Newport, J.W. (1992). Coupling of mitosis to the completion of S phase in Xenopus occurs via modulation of the tyrosine kinase that phosphorylates p34cdc2. Cell 68, 787–97.
Svejstrup, J.Q., Christiansen, K., Gromova, I.I., et al. (1991). New techniques for uncoupling the cleavage and religation reactions of eukaryotic topoisomerase I. The mode of action of camptothecin at a specific recognition site. J. Mol. Biol. 222, 669–78.
Tsao, Y.P., D'Arpa, P. & Liu, L.F. (1992). The involvement of active DNA synthesis in camptothecin-induced G2 arrest: altered regulation of p34cdc2/cyclin B. Cancer Res. 52, 1823–9.
Uemura, T., Ohkura, H., Adachi, Y, Morino, K., Shiozaki, K. & Yanagida, M. (1987). DNA topoisomerase II is required for condensation and separation of mitotic chromosomes in S. pombe. Cell 50, 917–25.
Walker, C. & Streisinger, G. (1983). Induction of mutagenesis by gamma-rays in pregonial germ cells of zebrafish embryos. Genetics 103, 125–36.
Walker, D.H., DePaoli-Roach, A.A. & Mailer, J.L. (1992). Multiple roles for protein phosphatase 1 in regulating the Xenopus early embryonic cell cycle. Mol. Biol. Cell 3, 687–98.
Wang, Y & Burke, D.J. (1995). Checkpoint genes required to delay cell division in response to nocodazole respond to impaired Kinetochore function in the yeast Saccharomyces cerevisiae. Mol. Cell Biol. 15, 6838–44.
Westerfield, M. (1995). The Zebrafish Book, 3rd edn. Eugene, OR: University of Oregon Press.
Wright, S.J. & Schatten, G. (1990). Teniposide, a topoisomerase II inhibitor, prevents chromosome condensation and separation but not decondensation in fertilized surf clam (Spisula solidissima) oocytes. Dev. Biol. 142, 224–32.
Zachariae, W., Shin, T.H., Galova, M., Obermaier, B. & Nasmyth, K. (1996). Identification of subunits of the anaphase-promoting complex of Saccharomyces cerevisiae. Science 274, 1201–4.
Zhang, Y, Wang, Z. & Ravid, K. (1996). The cell cycle in polyploid megakaryocytes is associated with reduced activity of cyclin Bl-dependent cdc2 kinase. J. Biol. Chem. 217, 4266–72.


Effect of inhibitors of DNA replication on early zebrafish embryos: evidence for coordinate activation of multiple intrinsic cell-cycle checkpoints at the mid-blastula transition

  • Richard Ikegami (a1), Alma K. Rivera-Bennetts (a1), Deborah L. Brooker (a1) and Thomas D. Yager (a1)


Full text views

Total number of HTML views: 0
Total number of PDF views: 0 *
Loading metrics...

Abstract views

Total abstract views: 0 *
Loading metrics...

* Views captured on Cambridge Core between <date>. This data will be updated every 24 hours.

Usage data cannot currently be displayed