• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of jcellbiolHomeThe Rockefeller University PressEditorsContactInstructions for AuthorsThis issue
J Cell Biol. Jun 2, 1994; 125(6): 1303–1312.
PMCID: PMC2290920

Cyclin-like accumulation and loss of the putative kinetochore motor CENP-E results from coupling continuous synthesis with specific degradation at the end of mitosis

Abstract

CENP-E is a kinesin-like protein that binds to kinetochores through the early stages of mitosis, but after initiation of anaphase, it relocalizes to the overlapping microtubules in the midzone, ultimately concentration in the developing midbody. By immunoblotting of cells separated at various positions in the cell cycle using centrifugal elutriation, we show that CENP-E levels increase progressively across the cycle peaking at approximately 22,000 molecules/cell early in mitosis, followed by an abrupt (> 10 fold) loss at the end of mitosis. Pulse-labeling with [35S]methionine reveals that beyond a twofold increase in synthesis between G1 and G2, interphase accumulation results primarily from stabilization of CENP-E during S and G2. Despite localizing in the midbody during normal cell division, CENP-E loss at the end of mitosis is independent of cytokinesis, since complete blockage of division with cytochalasin has no affect on CENP-E loss at the M/G1 transition. Thus, like mitotic cyclins, CENP-E accumulation peaks before cell division, and it is specifically degraded at the end of mitosis. However, CENP-E degradation kinetically follows proteolysis of cyclin B in anaphase. Combined with cyclin A destruction before the end of metaphase, degradation of as yet unidentified components at the metaphase/anaphase transition, and cyclin B degradation at or after the anaphase transition, CENP-E destruction defines a fourth point in a mitotic cascade of timed proteolysis.

Full Text

The Full Text of this article is available as a PDF (1.8M).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Blose SH, Meltzer DI, Feramisco JR. 10-nm filaments are induced to collapse in living cells microinjected with monoclonal and polyclonal antibodies against tubulin. J Cell Biol. 1984 Mar;98(3):847–858. [PMC free article] [PubMed]
  • Carter SB. Effects of cytochalasins on mammalian cells. Nature. 1967 Jan 21;213(5073):261–264. [PubMed]
  • Cooke CA, Heck MM, Earnshaw WC. The inner centromere protein (INCENP) antigens: movement from inner centromere to midbody during mitosis. J Cell Biol. 1987 Nov;105(5):2053–2067. [PMC free article] [PubMed]
  • Edgar BA, Sprenger F, Duronio RJ, Leopold P, O'Farrell PH. Distinct molecular mechanism regulate cell cycle timing at successive stages of Drosophila embryogenesis. Genes Dev. 1994 Feb 15;8(4):440–452. [PubMed]
  • Endow SA. The emerging kinesin family of microtubule motor proteins. Trends Biochem Sci. 1991 Jun;16(6):221–225. [PubMed]
  • Euteneuer U, McIntosh JR. Structural polarity of kinetochore microtubules in PtK1 cells. J Cell Biol. 1981 May;89(2):338–345. [PMC free article] [PubMed]
  • Evans T, Rosenthal ET, Youngblom J, Distel D, Hunt T. Cyclin: a protein specified by maternal mRNA in sea urchin eggs that is destroyed at each cleavage division. Cell. 1983 Jun;33(2):389–396. [PubMed]
  • Gallant P, Nigg EA. Cyclin B2 undergoes cell cycle-dependent nuclear translocation and, when expressed as a non-destructible mutant, causes mitotic arrest in HeLa cells. J Cell Biol. 1992 Apr;117(1):213–224. [PMC free article] [PubMed]
  • Glotzer M, Murray AW, Kirschner MW. Cyclin is degraded by the ubiquitin pathway. Nature. 1991 Jan 10;349(6305):132–138. [PubMed]
  • Gorbsky GJ, Sammak PJ, Borisy GG. Chromosomes move poleward in anaphase along stationary microtubules that coordinately disassemble from their kinetochore ends. J Cell Biol. 1987 Jan;104(1):9–18. [PMC free article] [PubMed]
  • Heck MM, Hittelman WN, Earnshaw WC. Differential expression of DNA topoisomerases I and II during the eukaryotic cell cycle. Proc Natl Acad Sci U S A. 1988 Feb;85(4):1086–1090. [PMC free article] [PubMed]
  • Hershko A, Ganoth D, Pehrson J, Palazzo RE, Cohen LH. Methylated ubiquitin inhibits cyclin degradation in clam embryo extracts. J Biol Chem. 1991 Sep 5;266(25):16376–16379. [PubMed]
  • Holloway SL, Glotzer M, King RW, Murray AW. Anaphase is initiated by proteolysis rather than by the inactivation of maturation-promoting factor. Cell. 1993 Jul 2;73(7):1393–1402. [PubMed]
  • Hunt T, Luca FC, Ruderman JV. The requirements for protein synthesis and degradation, and the control of destruction of cyclins A and B in the meiotic and mitotic cell cycles of the clam embryo. J Cell Biol. 1992 Feb;116(3):707–724. [PMC free article] [PubMed]
  • Hyman AA, Mitchison TJ. Two different microtubule-based motor activities with opposite polarities in kinetochores. Nature. 1991 May 16;351(6323):206–211. [PubMed]
  • Kauffman MG, Noga SJ, Kelly TJ, Donnenberg AD. Isolation of cell cycle fractions by counterflow centrifugal elutriation. Anal Biochem. 1990 Nov 15;191(1):41–46. [PubMed]
  • Koshland DE, Mitchison TJ, Kirschner MW. Polewards chromosome movement driven by microtubule depolymerization in vitro. Nature. 1988 Feb 11;331(6156):499–504. [PubMed]
  • Lehner CF, O'Farrell PH. The roles of Drosophila cyclins A and B in mitotic control. Cell. 1990 May 4;61(3):535–547. [PMC free article] [PubMed]
  • Luca FC, Ruderman JV. Control of programmed cyclin destruction in a cell-free system. J Cell Biol. 1989 Nov;109(5):1895–1909. [PMC free article] [PubMed]
  • Luca FC, Shibuya EK, Dohrmann CE, Ruderman JV. Both cyclin A delta 60 and B delta 97 are stable and arrest cells in M-phase, but only cyclin B delta 97 turns on cyclin destruction. EMBO J. 1991 Dec;10(13):4311–4320. [PMC free article] [PubMed]
  • Minshull J, Blow JJ, Hunt T. Translation of cyclin mRNA is necessary for extracts of activated xenopus eggs to enter mitosis. Cell. 1989 Mar 24;56(6):947–956. [PubMed]
  • Minshull J, Pines J, Golsteyn R, Standart N, Mackie S, Colman A, Blow J, Ruderman JV, Wu M, Hunt T. The role of cyclin synthesis, modification and destruction in the control of cell division. J Cell Sci Suppl. 1989;12:77–97. [PubMed]
  • Minshull J, Golsteyn R, Hill CS, Hunt T. The A- and B-type cyclin associated cdc2 kinases in Xenopus turn on and off at different times in the cell cycle. EMBO J. 1990 Sep;9(9):2865–2875. [PMC free article] [PubMed]
  • Mullins JM, Biesele JJ. Cytokinetic activities in a human cell line: the midbody and intercellular bridge. Tissue Cell. 1973;5(1):47–61. [PubMed]
  • Mullins JM, Biesele JJ. Terminal phase of cytokinesis in D-98s cells. J Cell Biol. 1977 Jun;73(3):672–684. [PMC free article] [PubMed]
  • Murray AW, Kirschner MW. Dominoes and clocks: the union of two views of the cell cycle. Science. 1989 Nov 3;246(4930):614–621. [PubMed]
  • Murray AW, Kirschner MW. Cyclin synthesis drives the early embryonic cell cycle. Nature. 1989 May 25;339(6222):275–280. [PubMed]
  • Murray AW, Solomon MJ, Kirschner MW. The role of cyclin synthesis and degradation in the control of maturation promoting factor activity. Nature. 1989 May 25;339(6222):280–286. [PubMed]
  • Muschel RJ, Zhang HB, McKenna WG. Differential effect of ionizing radiation on the expression of cyclin A and cyclin B in HeLa cells. Cancer Res. 1993 Mar 1;53(5):1128–1135. [PubMed]
  • Nicklas RB. The motor for poleward chromosome movement in anaphase is in or near the kinetochore. J Cell Biol. 1989 Nov;109(5):2245–2255. [PMC free article] [PubMed]
  • Norbury C, Nurse P. Animal cell cycles and their control. Annu Rev Biochem. 1992;61:441–470. [PubMed]
  • Pelham HR, Jackson RJ. An efficient mRNA-dependent translation system from reticulocyte lysates. Eur J Biochem. 1976 Aug 1;67(1):247–256. [PubMed]
  • Pines J, Hunter T. Isolation of a human cyclin cDNA: evidence for cyclin mRNA and protein regulation in the cell cycle and for interaction with p34cdc2. Cell. 1989 Sep 8;58(5):833–846. [PubMed]
  • Pines J, Hunter T. Human cyclin A is adenovirus E1A-associated protein p60 and behaves differently from cyclin B. Nature. 1990 Aug 23;346(6286):760–763. [PubMed]
  • Rechsteiner M. PEST sequences are signals for rapid intracellular proteolysis. Semin Cell Biol. 1990 Dec;1(6):433–440. [PubMed]
  • Rieder CL, Alexander SP. Kinetochores are transported poleward along a single astral microtubule during chromosome attachment to the spindle in newt lung cells. J Cell Biol. 1990 Jan;110(1):81–95. [PMC free article] [PubMed]
  • Rogers S, Wells R, Rechsteiner M. Amino acid sequences common to rapidly degraded proteins: the PEST hypothesis. Science. 1986 Oct 17;234(4774):364–368. [PubMed]
  • Schroeder TE. The contractile ring. I. Fine structure of dividing mammalian (HeLa) cells and the effects of cytochalasin B. Z Zellforsch Mikrosk Anat. 1970;109(4):431–449. [PubMed]
  • Sherley JL, Kelly TJ. Regulation of human thymidine kinase during the cell cycle. J Biol Chem. 1988 Jun 15;263(17):8350–8358. [PubMed]
  • Skoufias DA, Scholey JM. Cytoplasmic microtubule-based motor proteins. Curr Opin Cell Biol. 1993 Feb;5(1):95–104. [PubMed]
  • Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC. Measurement of protein using bicinchoninic acid. Anal Biochem. 1985 Oct;150(1):76–85. [PubMed]
  • Standart N, Minshull J, Pines J, Hunt T. Cyclin synthesis, modification and destruction during meiotic maturation of the starfish oocyte. Dev Biol. 1987 Nov;124(1):248–258. [PubMed]
  • Surana U, Amon A, Dowzer C, McGrew J, Byers B, Nasmyth K. Destruction of the CDC28/CLB mitotic kinase is not required for the metaphase to anaphase transition in budding yeast. EMBO J. 1993 May;12(5):1969–1978. [PMC free article] [PubMed]
  • Swenson KI, Farrell KM, Ruderman JV. The clam embryo protein cyclin A induces entry into M phase and the resumption of meiosis in Xenopus oocytes. Cell. 1986 Dec 26;47(6):861–870. [PubMed]
  • Towbin H, Staehelin T, Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. [PMC free article] [PubMed]
  • van der Velden HM, Lohka MJ. Mitotic arrest caused by the amino terminus of Xenopus cyclin B2. Mol Cell Biol. 1993 Mar;13(3):1480–1488. [PMC free article] [PubMed]
  • Westendorf JM, Swenson KI, Ruderman JV. The role of cyclin B in meiosis I. J Cell Biol. 1989 Apr;108(4):1431–1444. [PMC free article] [PubMed]
  • Yen TJ, Gay DA, Pachter JS, Cleveland DW. Autoregulated changes in stability of polyribosome-bound beta-tubulin mRNAs are specified by the first 13 translated nucleotides. Mol Cell Biol. 1988 Mar;8(3):1224–1235. [PMC free article] [PubMed]
  • Yen TJ, Compton DA, Wise D, Zinkowski RP, Brinkley BR, Earnshaw WC, Cleveland DW. CENP-E, a novel human centromere-associated protein required for progression from metaphase to anaphase. EMBO J. 1991 May;10(5):1245–1254. [PMC free article] [PubMed]
  • Yen TJ, Li G, Schaar BT, Szilak I, Cleveland DW. CENP-E is a putative kinetochore motor that accumulates just before mitosis. Nature. 1992 Oct 8;359(6395):536–539. [PubMed]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • Compound
    Compound
    PubChem Compound links
  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...