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Changes in protein association with intracellular membranes of Xenopus laevis oocytes during maturation and activation

Published online by Cambridge University Press:  26 September 2008

N.S. Duesbery  and
Y. Masui
N.S. Duesbery*
Affiliation:
University of Toronto, Toronto, Canada.
Y. Masui
Affiliation:
University of Toronto, Toronto, Canada.
*
N.S. Duesbery, Department of Zoology, University of Tronto, Toronto, CanadaM5S 1A1.
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Extract

Intracellular membranes isolated from fully grown immature oocytes, mature oocytes (eggs) and activated eggs of Xenopus laevis were fractionated through a discontinuous sucrose density gradient into light, intermediate and heavy fractions. Electron microscopy showed that the light and intermediate fractions consisted mainly of smooth membranes, while the heavy fraction consisted mainly of rough membranes and mitochondria. Variations in the proteins associate with samples taken at different stages were observed by SDS-PAGE. The following differences were consistently observed: a 200 kDa protein was present only in the intermediate fraction of activated eggs, 29 and 44 kDa proteins were present only in the intermediate fractions of immature oocytes and activated eggs, and 120 and 145 kDa proteins were present only in the heavy fractions of mature oocytes and activated eggs. Examination of Western blots showed that cyclins A and B2 did not associate with membrane fractions at any stage of meiosis. Instead, cyclin A was present in the cytosols of mature oocytes and cyclin B2 was present in the cytosols of immature and mature oocytes. c-mos protein was detected in the cytosols and occasionally in the light fractions of mature oocytes and activated eggs. While α- and β-tubulins were detected in the light and intermediate fractions at all the stages of meiosis examined, only β-tubulin was present in the heavy fraction. βtubulin present in the heavy fraction was detected only at interphase, i.e. in immature oocutes and activated eggs, and not in mature oocytes. Using immunogold labelling we confirmed these results and found evidence to suggest that β-tubulin associates with the rough endoplasmic reticulum of interphase cells by a linking protein.

Keywords

Cell cycleEndoplasmic reticulumOocyte maturationTubulinXenopus laevis

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Type
Article
Information
Zygote , Volume 1 , Issue 2 , May 1993 , pp. 129 - 141
Copyright
Copyright © Cambridge University Press 1993

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References

Adelman, M.R., Sabatini, D.D. & Blobel, G. (1973). Ribosome-membrane interaction: nondestructive disassembly of rat liver microsomes into ribosomal and membranous components. J. Cell Biol. 56, 206–29.Google Scholar
Allan, V.J. & Vale, R.D. (1991). Cell cycle control of microtubule-based membrane transport and tubule formation in vitro. J. Cell Biol. 113, 347–59.CrossRefGoogle ScholarPubMed
Amos, L.A. & Klug, A. (1974). Arrangement of subunits in flagellar microtubules. J. Cell Sci. 14, 523–49.CrossRefGoogle ScholarPubMed
Balinsky, B.I. & Devis, R.J. (1963). Origin and differentiation of cytoplasmic structures in the oocytes of Xenopus laevis. Acta Embryol. Exp. Morphol. 6, 55–108.Google Scholar
Bole, D.G., Hendershot, L.M. & Kearney, J.F. (1986). Post-translational association of immunoglobulin heavy chain binding protein with nascent heavy chains in nonsecreting and secreting hybridomas. J. Cell Biol. 102, 1558–66.Google Scholar
Bordier, C. (1981). Phase separation of integral membrane proteins in Triton X-114 solution. J. Biol. Chem. 256, 1604–7.CrossRefGoogle ScholarPubMed
Brachet, J., Hanocq, F. & Van Gansen, P. (1970). A cytochemical and ultrastructural analysis of in vitro maturation in amphibian oocytes. Dev. Biol. 21, 157–95.Google Scholar
Ceriotti, A. & Colman, A. (1989). Protein transport from endoplasmic reticulum to the golgi complex can occur during meiotic metaphase in Xenopus oocytes. J. Cell Biol. 109, 1439–44.Google Scholar
Colman, A., Jones, E.A. & Heasman, J. (1985). Meiotic maturation in Xenopus oocytes: a link between the cessation of protein secretion and the polarized disappearance of Golgi apparati. J. Cell Biol. 101, 313–18.Google Scholar
Dabora, S.L. & Sheetz, M.P. (1989). The microtubule-dependent formation of a tubulovesicular network with characteristics of the ER from cultured cell extracts. Cell 54, 27–35.CrossRefGoogle Scholar
De Sousa, P.A. & Masui, Y. (1990). The effect of cytochalasin D on protein synthesis in Xenopus laevis oocytes. Mol. Reprod. Dev. 26, 248–52.CrossRefGoogle ScholarPubMed
Detrich, W.W. III. & Williams, R.C. Jr. (1978). Reversible dissociation of the αβ dimer of tubulin from bovine brain. Biochemistry 17, 3900–7.Google Scholar
Dominguez, J.M. & Paiement, J. (1989). Reconstitution of endoplasmic reticulum in rapidly dividing cells of early Xenopus embryos. Am. J. Anat. 186, 99–113.CrossRefGoogle Scholar
Dumont, J.N. (1972). Oogenesis in Xenopus laevis (Daudin). I.Stages of oocyte development in laboratory maintained animals. J. Morphol. 136, 153–79.CrossRefGoogle ScholarPubMed
Elinson, R.P. (1985). Changes in levels of polymeric tubulin associated with activation and dorsoventral polarization of the frog egg. Dev. Biol. 109, 224–33.CrossRefGoogle ScholarPubMed
Gautier, J. & Maller, J.L. (1991). Cyclin B in Xenopus oocytes: implications for the mechanism of pre-MPF activation. EMBO J. 10, 177–82.Google Scholar
Gautier, J., Minshull, J., Lohka, M., Glotzer, M., Hunt, T. & Maller, J.L. (1990). Cyclin is a component of maturation-promoting factor from Xenopus. Cell 60, 487–94.CrossRefGoogle ScholarPubMed
Geuens, G., Gundersen, G.G., Nuydens, R., Cornelissen, F., Bulinski, J.C. & DeBrabander, M. (1986). Ultrastructural colocalization of tyrosinated and detyrosinated α-tubulin in interphase and mitotic cells. J. Cell Biol. 103, 1883–93.Google Scholar
Hirokawa, N. (1982). Cross-linker system between neuro-filaments, microtubules, and membrane organelles in frog axons revealed by the quick-freeze, deep-etching method. J. Cell Biol. 94, 129–42.Google Scholar
Hirokawa, N., Bloom, G.S. and Vallee, R.B. (1985). Cytoskeletal architecture and immunocytochemical localization of microtubule-associated proteins in regions of axons associated with rapid axonal transport: the β,β′-iminodipropionitrile-intoxicated axon as a model system. J. Cell Biol. 101, 227–39.Google Scholar
Houliston, E. & Elinson, R.P. (1991). Evidence for the involvement of microtubules, ER, and kinesin in the cortical rotation of fertilized frog eggs. J. Cell Biol. 114, 1014–28.CrossRefGoogle ScholarPubMed
Houliston, E. & Elinson, R.P. (1992). Microtubules and cytoplasmic reorganization in the frog egg. Curr. Top. Dev. Biol. 26, 53–70.CrossRefGoogle ScholarPubMed
Imoh, H., Okamoto, M. & Eguchi, G. (1983). Accumulation of annulate lamellae in the subcortical layer during progesterone-induced oocyte maturation in Xenopus laevis. Dev., Growth and Differ. 25, 1–10.Google Scholar
Kessel, R.G. & Subtelney, S. (1981). Alteration of annulate lamellae in the in vitro progesterone-treated, full-grown Rana pipiens oocyte. J. Exp. Zool. 217, 119–35.CrossRefGoogle ScholarPubMed
Kobayashi, H., Minshull, J., Ford, C., Golsteyn, R., Poon, R. and Hunt, T. (1991). On the synthesis and destruction of A- and B-type cyclins during oogenesis and meiotic maturation in Xenopus laevis. J. Cell Biol. 114, 755–65.CrossRefGoogle ScholarPubMed
Laemmli, U.K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–5.Google Scholar
Leaf, D.S., Roberts, S.J., Gerhart, J.C. & Moore, Hsiao-P. (1990). The secretory pathway is blocked between the trans-golgi and the plasma membrane during meiotic maturation in Xenopus oocytes. Dev. Biol. 141, 1–12.CrossRefGoogle ScholarPubMed
Lohka, M.J. & Maller, J.L. (1985). Induction of nuclear envelope breakdown, chromosome condensation, and spindle formation in cell-free extracts. J. Cell Biol. 101, 518–23.Google Scholar
Masui, Y. & Markert, C.L. (1971). Cytoplasmic control of nuclear behaviour during meiotic maturation of frog oocytes. J. Exp. Zool. 177, 129–6.Google Scholar
Miller, R.H. & Lasek, R.J. (1985). Cross-bridges mediate anterograde and retrograde vesicle transport along microtubules in squid axoplasm. J. Cell Biol. 101, 2181–93.CrossRefGoogle ScholarPubMed
Newport, J.W. & Kirschner, M.W. (1984). Regulation of the cell cycle during early Xenopus development. Cell 37, 731–42.Google Scholar
Reynolds, E.S. (1963). The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J. Cell Biol. 17, 208–12.Google Scholar
Sagata, N., Watanabe, N., Vande Woude, G.F. & Ikawa, Y. (1989). The c-mos proto-oncogene product is a cytostatic factor (CSF) responsible for meiotic arrest in vertebrate eggs. Nature 342, 505–11.CrossRefGoogle ScholarPubMed
Smith, P.K., Krohn, R.I., Hermanson, G.T., Mallia, A.K., Gartner, F.H., Provenzano, M.D., Fujimoto, E.K., Goeke, N.M., Olson, B.J. & Klenk, D.C. (1985). Measurement of protein using bicinchoninic acid. Anal. Biochem. 150, 76–85.Google Scholar
Suprynowicz, F.A. & Mazia, D. (1985). Fluctuation of the Ca2+ -sequestering activity of permeabilized sea urchin embryos during the cell cycle. Proc. Natl. Acad. Sci. USA 82, 2389–93.CrossRefGoogle ScholarPubMed
Terasaki, M., Chen, L.B. & Fujiwara, K. (1986). Microtubules and the endoplasmic reticulum are highly interdependent structures. J. Cell Biol. 103, 1557–68.Google Scholar
Thyberg, J. & Moskalewski, S. (1985). Microtubules and the organization of the Golgi complex. Exp. Cell Res. 159, 1–16.Google Scholar
Towbin, H., Staehelin, T. & Gordon, J. (1979). Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc. Natl. Acad. Sci. USA 76, 4350–4.Google Scholar
Valentine, R.C. & Green, N.M. (1967). Electron microscopy of an antibody-hapten complex. J. Mol. Biol. 27, 615–17.Google Scholar
Van Gansen, P. (1966). Ultrastructure comparée du cytoplasme périphérique des oocytes murs et des ouefs vierges de Xenopus laevis (Bactracien anoure). J. Embryol. Exp. Morphol. 15, 355–64.Google Scholar
Watanabe, N., Vande Woude, G.F., Ikawa, Y. & Sagata, N. (1989). Specific proteolysis of the c-mos proto-oncogene product by calpain on fertilization of Xenopus eggs. Nature 342, 505–11.CrossRefGoogle ScholarPubMed
Watanabe, N., Vande Woude, G.F., Ikawa, Y. & Sagata, N. (1991). Independent inactivation of MPF and cytostatic factor (Mos) upon fertilization of Xenopus eggs. Nature 352, 247–8.Google Scholar
Watson, M.L. (1958). Staining of tissue sections for electron microscopy with heavy metals. J. Biophys. Biochem. Cytol. 4, 475–8.Google Scholar
Young, G.P.H., Ding, J., Young, E., Deshpande, A.K., Goldstein, M., Koide, S.S. & Cohn, Z.A. (1984). A Ca2+ -activated channel from Xenopus laevis oocyte membranes reconstituted into planar bilayers. Proc. Natl. Acad. Sci. USA 81, 5155–9.CrossRefGoogle ScholarPubMed
Zhang, S.C. & Masui, Y. (1992). Activation of Xenopus laevis eggs in the absence of intracellular Ca activity by the protein phosphorylation inhibitor, 6-dimethylaminopurine (6-DMAP). J. Exp. Zool. 262, 317–29.Google Scholar
Zhou, R., Oskarsson, M., Paules, R.S., Schulz, N., Cleveland, D. & Vande Woude, G.F. (1991). Ability of the c-mos product to associate with and phosphorylate tubulin. Science 251, 671–5.CrossRefGoogle ScholarPubMed