Search tips
Search criteria 


Logo of jcellbiolHomeThe Rockefeller University PressEditorsContactInstructions for AuthorsThis issue
J Cell Biol. 1985 March 1; 100(3): 897–903.
PMCID: PMC2113528

Experimental separation of pronuclei in fertilized sea urchin eggs: chromosomes do not organize a spindle in the absence of centrosomes


We tested the ability of chromosomes in a mitotic cytoplasm to organize a bipolar spindle in the absence of centrosomes. Sea urchin eggs were treated with 5 X 10(-6) colcemid for 7-9 min before fertilization to block future microtubule assembly. Fertilization events were normal except that a sperm aster was not formed and the pronuclei remained up to 70 microns apart. After nuclear envelope breakdown, individual eggs were irradiated with 366-nm light to inactivate photochemically the colcemid. A functional haploid bipolar spindle was immediately assembled in association with the male chromosomes. In contrast to the male pronucleus, the female pronucleus in most of these eggs remained as a small nonbirefringent hyaline area throughout mitosis. High- voltage electron microscopy of serial semithick sections from individual eggs, previously followed in vivo, revealed that the female chromosomes were randomly distributed within the remnants of the nuclear envelope. No microtubules were found in these pronuclear areas even though the chromosomes were well-condensed and had prominent kinetochores with well-developed coronas. In the remaining eggs, a weakly birefringent monaster was assembled in the female pronuclear area. These observations demonstrate that chromosomes in a mitotic cytoplasm cannot organize a bipolar spindle in the absence of a spindle pole or even in the presence of a monaster. In fact, chromosomes do not even assemble kinetochore microtubules in the absence of a spindle pole, and kinetochore microtubules form only on kinetochores facing the pole when a monaster is present. This study also provides direct experimental proof for the longstanding paradigm that the sperm provides the centrosomes used in the development of the sea urchin zygote.

Full Text

The Full Text of this article is available as a PDF (2.8M).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Schatten H, Schatten G, Petzelt C, Mazia D. Effects of griseofulvin on fertilization and early development of sea urchins. Independence of DNA synthesis, chromosome condensation, and cytokinesis cycles from microtubule-mediated events. Eur J Cell Biol. 1982 Apr;27(1):74–87. [PubMed]
  • Aronson JF. Nuclear membrane fusion in fertilized Lytechinus variegatus eggs. J Cell Biol. 1973 Jul;58(1):126–134. [PMC free article] [PubMed]
  • Begg DA, Ellis GW. Micromanipulation studies of chromosome movement. I. Chromosome-spindle attachment and the mechanical properties of chromosomal spindle fibers. J Cell Biol. 1979 Aug;82(2):528–541. [PMC free article] [PubMed]
  • Begg DA, Ellis GW. Micromanipulation studies of chromosome movement. II. Birefringent chromosomal fibers and the mechanical attachment of chromosomes to the spindle. J Cell Biol. 1979 Aug;82(2):542–554. [PMC free article] [PubMed]
  • Berns MW, Rattner JB, Brenner S, Meredith S. The role of the centriolar region in animal cell mitosis. A laser microbeam study. J Cell Biol. 1977 Feb;72(2):351–367. [PMC free article] [PubMed]
  • Calarco-Gillam PD, Siebert MC, Hubble R, Mitchison T, Kirschner M. Centrosome development in early mouse embryos as defined by an autoantibody against pericentriolar material. Cell. 1983 Dec;35(3 Pt 2):621–629. [PubMed]
  • De Brabander M, Geuens G, De Mey J, Joniau M. Nucleated assembly of mitotic microtubules in living PTK2 cells after release from nocodazole treatment. Cell Motil. 1981;1(4):469–483. [PubMed]
  • Euteneuer U, Ris H, Borisy GG. Polarity of kinetochore microtubules in Chinese hamster ovary cells after recovery from a colcemid block. J Cell Biol. 1983 Jul;97(1):202–208. [PMC free article] [PubMed]
  • Fuseler JW. Repetitive procurement of mature gametes from individual sea stars and sea urchins. J Cell Biol. 1973 Jun;57(3):879–881. [PMC free article] [PubMed]
  • Karsenti E, Newport J, Hubble R, Kirschner M. Interconversion of metaphase and interphase microtubule arrays, as studied by the injection of centrosomes and nuclei into Xenopus eggs. J Cell Biol. 1984 May;98(5):1730–1745. [PMC free article] [PubMed]
  • Karsenti E, Newport J, Kirschner M. Respective roles of centrosomes and chromatin in the conversion of microtubule arrays from interphase to metaphase. J Cell Biol. 1984 Jul;99(1 Pt 2):47s–54s. [PMC free article] [PubMed]
  • Mazia D, Paweletz N, Sluder G, Finze EM. Cooperation of kinetochores and pole in the establishment of monopolar mitotic apparatus. Proc Natl Acad Sci U S A. 1981 Jan;78(1):377–381. [PubMed]
  • Nicklas RB. Mitosis. Adv Cell Biol. 1971;2:225–297. [PubMed]
  • Paweletz N, Mazia D. Fine structure of the mitotic cycle of unfertilized sea urchin eggs activated by ammoniacal sea water. Eur J Cell Biol. 1979 Oct;20(1):37–44. [PubMed]
  • Pickett-Heaps JD, Tippit DH. The diatom spindle in perspective. Cell. 1978 Jul;14(3):455–467. [PubMed]
  • Pickett-Heaps JD, Tippit DH, Porter KR. Rethinking mitosis. Cell. 1982 Jul;29(3):729–744. [PubMed]
  • Rieder CL. Thick and thin serial sectioning for the three-dimensional reconstruction of biological ultrastructure. Methods Cell Biol. 1981;22:215–249. [PubMed]
  • Rieder CL. The formation, structure, and composition of the mammalian kinetochore and kinetochore fiber. Int Rev Cytol. 1982;79:1–58. [PubMed]
  • Rieder CL, Bowser SS. Factors which influence light microscopic visualization of biological material in sections prepared for electron microscopy. J Microsc. 1983 Oct;132(Pt 1):71–80. [PubMed]
  • Ring D, Hubble R, Kirschner M. Mitosis in a cell with multiple centrioles. J Cell Biol. 1982 Sep;94(3):549–556. [PMC free article] [PubMed]
  • Sluder G. Experimental manipulation of the amount of tubulin available for assembly into the spindle of dividing sea urchin eggs. J Cell Biol. 1976 Jul;70(1):75–85. [PMC free article] [PubMed]
  • Sluder G. Role of spindle microtubules in the control of cell cycle timing. J Cell Biol. 1979 Mar;80(3):674–691. [PMC free article] [PubMed]
  • Sluder G, Begg DA. Control mechanisms of the cell cycle: role of the spatial arrangement of spindle components in the timing of mitotic events. J Cell Biol. 1983 Sep;97(3):877–886. [PMC free article] [PubMed]
  • Witt PL, Ris H, Borisy GG. Origin of kinetochore microtubules in Chinese hamster ovary cells. Chromosoma. 1980;81(3):483–505. [PubMed]
  • Witt PL, Ris H, Borisy GG. Structure of kinetochore fibers: microtubule continuity and inter-microtubule bridges. Chromosoma. 1981;83(4):523–540. [PubMed]

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