Ultrastructure of tubulin paracrystals from sea urchin eggs, with determination of spacings by electron and optical diffraction

Triton-treated cortical fragments of unfertilized and fertilized sea urchin eggs prepared in the presence of greater than or equal to 5 mM EGTA contain 15-30% of the total egg actin. However, actin filaments are not readily apparent by electron microscopy on the cortical fragments of unfertilized eggs but are numerous on those of fertilized eggs. The majority of the actin associated with cortical fragments of unfertilized eggs is solubilized by dialysis against a low ionic strength buffer at pH 7.5. This soluble actin preparation (less than 50% pure actin) does not form proper filaments in 0.1 M KCl and 3 mM MgCl2, whereas actin purified from this preparation does, as judged by electron microscopy. Optical diffraction analysis reveals that these purified actin filaments have helical parameters very similar to those of muscle actin. Furthermore, the properties of the purified actin with regard to activation of myosin ATPase are similar to those of actin from other cell types. The possibility that actin is maintained in a nonfilamentous form on the inner surface of the unfertilized egg plasma membrane and is induced to assemble upon fertilization is discussed.

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The effects of mitotic inhibitors on the structure of vinblastine-induced tubulin paracrystals from sea-urchin eggs

Journal of Cell Science ◽

10.1242/jcs.20.1.91 ◽

1976 ◽

Vol 20 (1) ◽

pp. 91-100

Author(s):

D. Starling

Keyword(s):

Sea Urchin ◽

Ultrastructural Level ◽

Tubulin Polymerization ◽

Sea Urchin Eggs ◽

Mitotic Inhibitors ◽

Vinblastine Sulphate ◽

Tubulin Paracrystals ◽

Careful Standardization

Vinblastine sulphate (VLB) is known to induce in vivo formation of tubulin paracrystals in sea-urchin eggs. Corresponding paracrystals have been prepared in the presence of both vinblastine sulphate and other mitoclasic agents. Careful standardization of conditions was required to restrict the formation of alternative forms of the paracrystals induced by vinblastine alone. Comparisons were made between preparations in terms of paracrystal shape, size, proportion of eggs containing paracrystals, number per egg and their relative times of first appearance. A correlation between such properties were established. Comparison of paracrystals at the ultrastructural level showed them all to be similar regardless of the drugs present during their formation. The implications of tubulin polymerization in the presence of mitoclasic agents are discussed and mechanisms for paracrystal enhancement by combinations of such drugs are suggested. Some similarities of paracrystal and microtubule seeding are discussed together with the activation of tubulin in the pool.

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Two ultrastructurally distinct tubulin paracrystals induced in sea-urchin eggs by vinblastine sulphate

Journal of Cell Science ◽

10.1242/jcs.20.1.79 ◽

1976 ◽

Vol 20 (1) ◽

pp. 79-89 ◽

Cited By ~ 2

Author(s):

D. Starling

Keyword(s):

Sea Urchin ◽

Binding Sites ◽

Mammalian Cells ◽

The Other ◽

Sea Urchin Eggs ◽

Vincristine Sulphate ◽

Vinblastine Sulphate ◽

Available Information ◽

Tubulin Paracrystals

Two types of ultrastructurally distinct tubulin paracrystals have been induced in sea-urchin eggs with vinblastine sulphate (VLB) under different sets of conditions. One type of paracrystal appears to consist of hexagonally-close packed microtubules and closely resembles paracrystals present in mammalian cells treated with vinblastine or vincristine sulphate, but not previously reported in sea-urchin eggs. The other type is also made up of tubulin subunits, but these do not seem to have polymerized into microtubules. Both types of paracrystal are induced in sea-urchin eggs in the presence of VLB at a time when tubulin subunits would not normally polymerize. Possible mechanisms for tubulin activation and the induction of paracrystal formation are discussed in respect to the available information on the binding sites of the tubulin subunits.

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Yields of tubulin paracrystals, vinblastine-crystals, induced in unfertilized and fertilized sea urchin eggs in the presence of D2O.

Cell Structure and Function ◽

10.1247/csf.9.45 ◽

1984 ◽

Vol 9 (1) ◽

pp. 45-52 ◽

Cited By ~ 1

Author(s):

T. Choku Takahashi ◽

Hidemi Sato

Keyword(s):

Sea Urchin ◽

Sea Urchin Eggs ◽

Tubulin Paracrystals

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The Organization of Actin Filaments in the Stereocilia of the Bird Cochlea

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s1431927600002452 ◽

1980 ◽

Vol 38 ◽

pp. 554-555

Author(s):

E.J. Battles ◽

D. DeRosier ◽

J.C. Saunders ◽

L.G. Tilney

Keyword(s):

Hair Cell ◽

Diffraction Pattern ◽

Sea Urchin ◽

Actin Filaments ◽

Optical Diffraction ◽

Dense Material ◽

Apical Surface ◽

Structural Rigidity ◽

High Degree ◽

Degree Of Order

Extending from the apical surface of each hair cell of the chick cochlea are from 75 to 200 microvilli or stereocllia and one true cllium, the kinocilium. The stereocllia are arranged in rows of progressively increasing length (Fig. 1). Within each tapering sterocilium is a bundle of actin filaments with over 900 filaments near the tip yet only approximately 25 at the base where filaments are enmeshed in a dense material (Fig. 1); from here some of the filaments enter the apical surface of the cell (cuticular plate) as a rootlet. Examination of longitudinal sections of the stereocilia (Fig. 2) show that the filaments are aligned parallel to each other and show considerable order. Examination of an optical diffraction pattern of this bundle (Fig. 4) reveal that the actin filaments are packed such that the crossover points of adjacent actin filaments are inregister. A prominent reflection at 125Å−1 demonstrates that the filaments are cjossbridged by a macromolecular bridge situated at an average of 125Å−1 intervals (Fig. 4) in transverse sections the filaments appear hexagonally packed although there are regions where the filaments are less ordered (Fig. 3). In images processed in the computer to remove, noise and enhance detail periodic nature of the bridge can be clearly seen (see arrows Fig. 5). This image resembles that of an actin paracrystal formed from sea urchin extract composed of bundles of actin filaments crossbridged by a second protein. Thus the actin filaments in the bird stereocilia by being cross-bridged and packed with a high degree of order and produces a structure with considerable structural rigidity. Embryos were studied at various stages in development in an attempt to determine how the stereocilia form and how does the actin packing develops. These stages will be discussed.

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