Isolation of a sea urchin egg kinesin-related protein using peptide antibodies

1992 ◽  
Vol 101 (2) ◽  
pp. 291-301 ◽  
Author(s):  
D.G. Cole ◽  
W.Z. Cande ◽  
R.J. Baskin ◽  
D.A. Skoufias ◽  
C.J. Hogan ◽  
...  
Keyword(s):  
Cell Division ◽  
Molecular Mass ◽  
Sea Urchin ◽  
Heavy Chain ◽  
Relative Molecular Mass ◽  
Related Protein ◽  
Sea Urchin Egg ◽  
Conventional Kinesin ◽  
Kinesin Superfamily ◽  
Peptide Antibodies

To understand the roles of kinesin and its relatives in cell division, it is necessary to identify and characterize multiple members of the kinesin superfamily from mitotic cells. To this end we have raised antisera to peptides corresponding to highly conserved regions of the motor domains of several known members of the kinesin superfamily. These peptide antibodies react specifically with the motor domains of kinesin and ncd protein, as expected, and they also react with several polypeptides (including kinesin heavy chain) that cosediment with microtubules (MTs) precipitated from AMPPNP-treated sea urchin egg cytosol. Subsequent fractionation of ATP eluates of these MTs yields a protein of relative molecular mass 330 × 10(3) that behaves as a complex of three polypeptides that are distinct from conventional kinesin subunits or fragments thereof. This complex contains 85 kDa and 95 kDa polypeptides, which react with our peptide antibodies, and a 115 kDa polypeptide, which does not. This triplet of polypeptides, which we refer to as KRP(85/95), binds to purified sea urchin egg tubulin in an AMPPNP-enhanced, ATP-sensitive manner and induces the formation of microtubule bundles. We therefore propose that the triplet corresponds to a novel sea urchin egg kinesin-related protein.

scholarly journals A Quantitative Description of Protoplasmic Movement During Cleavage in the Sea-Urchin Egg

1958 ◽  
Vol 35 (2) ◽  
pp. 407-424
Author(s):  
Y. HIRAMOTO
Keyword(s):  
Cell Division ◽  
Sea Urchin ◽  
Quantitative Description ◽  
Motive Force ◽  
Mitotic Apparatus ◽  
Band Theory ◽  
Cytoplasmic Granules ◽  
Carbon Particles ◽  
Sea Urchin Egg ◽  
Protoplasmic Movement

1. Protoplasmic movements during cleavage in the eggs of the heart-urchin Clypeaster japonicus have been followed by tracing the movements of cytoplasmic granules and of carbon particles adhering to the surface. 2. These movements are quantitatively described in normal eggs and in eggs whose mitotic apparatus has been destroyed by colchicine. 3. The results obtained are qualitatively similar to those obtained by Spek and by Dan and his collaborators. 4. Endoplasmic movement and changes in the length and shape of the astral rays are readily explained by the contracting-ring (band) theory. 5. The location of the motive force of cell division is discussed.

scholarly journals Inhibition of kinesin-driven microtubule motility by monoclonal antibodies to kinesin heavy chains.

1988 ◽  
Vol 107 (6) ◽  
pp. 2657-2667 ◽  
Author(s):  
A L Ingold ◽  
S A Cohn ◽  
J M Scholey
Keyword(s):  
Monoclonal Antibodies ◽  
Sea Urchin ◽  
Heavy Chain ◽  
Bovine Brain ◽  
Drosophila Embryo ◽  
Detectable Effect ◽  
Heavy Chains ◽  
Sea Urchin Egg ◽  
Dependent Inhibition ◽  
Mgatpase Activity

We have prepared and characterized seven mouse monoclonal antibodies (SUK 1-7) to the 130-kD heavy chain of sea urchin egg kinesin. On immunoblots, SUK 3 and SUK 4 cross-reacted with Drosophila embryo 116-kD heavy chains, and SUK 4, SUK 5, SUK 6, and SUK 7 bound to the 120-kD heavy chains of bovine brain kinesin. Three out of seven monoclonal antikinesins (SUK 4, SUK 6, and SUK 7) caused a dose-dependent inhibition of sea urchin egg kinesin-induced microtubule translocation, whereas the other four monoclonal antibodies had no detectable effect on this motility. The inhibitory monoclonal antibodies (SUK 4, SUK 6, and SUK 7) appear to bind to spatially related sites on an ATP-sensitive microtubule binding 45-kD chymotryptic fragment of the 130-kD heavy chain, whereas SUK 2 binds to a spatially distinct site. None of the monoclonal antikinesins inhibited the microtubule activated MgATPase activity of kinesin, suggesting that SUK 4, SUK 6, and SUK 7 uncouple this MgATPase activity from motility.

scholarly journals STUDIES ON SULFHYDRYL GROUPS DURING CELL DIVISION OF SEA URCHIN EGG

1960 ◽  
Vol 8 (3) ◽  
pp. 603-607 ◽  
Author(s):  
Hikoichi Sakai
Keyword(s):  
Cell Division ◽  
Sea Urchin ◽  
Sulfhydryl Groups ◽  
Cell Stage ◽  
Sea Urchin Eggs ◽  
Maximum Value ◽  
Sea Urchin Egg ◽  
Egg Cortex

Masses of cortices of both unfertilized and fertilized sea urchin eggs can be isolated by crushing eggs in hypotonic MaCl2 (0.1 M) solution. The amount of cortical material in terms of protein-N increases steadily after fertilization until the monaster stage and thereafter remains almost constant until well into the two-cell stage. The amount of bound—SH per protein-N of the egg cortex also increases after fertilization, reaches a maximum value at the amphiaster stage and thereafter decreases rapidly as the cleavage of the cell proceeds.

scholarly journals Roles of kinesin and kinesin-like proteins in sea urchin embryonic cell division: evaluation using antibody microinjection.

1993 ◽  
Vol 123 (3) ◽  
pp. 681-689 ◽  
Author(s):  
B D Wright ◽  
M Terasaki ◽  
J M Scholey
Keyword(s):  
Cell Division ◽  
Sea Urchin ◽  
Heavy Chain ◽  
Mammalian Cells ◽  
Embryonic Cell ◽  
Mitotic Apparatus ◽  
Specific Antibodies ◽  
Sea Urchin Embryos ◽  
Mitotic Figures

Previous studies suggest that kinesin heavy chain (KHC) is associated with ER-derived membranes that accumulate in the mitotic apparatus in cells of early sea urchin embryos (Wright, B. D., J. H. Henson, K. P. Wedaman, P. J. Willy, J. N. Morand, and J. M. Scholey. 1991. J. Cell Biol. 113:817-833). Here, we report that the microinjection of KHC-specific antibodies into these cells has no effect on mitosis or ER membrane organization, even though one such antibody, SUK4, blocks kinesin-driven motility in vitro and in mammalian cells. Microinjected SUK4 was localized to early mitotic figures, suggesting that it is able to access kinesin in spindles. In contrast to KHC-specific antibodies, two antibodies that react with kinesin-like proteins (KLPs), namely CHO1 and HD, disrupted mitosis and prevented subsequent cell division. CHO1 is thought to exert this effect by blocking the activity of a 110-kD KLP. The relevant target of HD, which was raised against the KHC motor domain, is unknown; HD may disrupt mitosis by interfering with an essential spindle KLP but not with KHC itself, as preabsorption of HD with KHC did not alter its ability to block mitosis. These data indicate that some KLPs have essential mitotic functions in early sea urchin embryos but KHC itself does not.

scholarly journals THE EFFECT OF PENICILLIN ON EGGS OF THE SEA URCHIN, ARBACIA PUNCTULATA

1945 ◽  
Vol 28 (5) ◽  
pp. 405-413 ◽  
Author(s):  
Richard J. Henry ◽  
Maryon D. Henry
Keyword(s):  
Oxygen Consumption ◽  
Cell Division ◽  
Sea Urchin ◽  
Inhibitory Action ◽  
Initial Concentration ◽  
Arbacia Punctulata ◽  
Sea Urchin Egg

1. Penicillin in the range of concentration from 250 U/ml. to approximately 2650 U/ml. inhibits the rate of cell division of the fertilized sea urchin egg from 0 to 100 per cent. 2. Penicillin in the same range of concentrations has no effect on the oxygen consumption of the unfertilized or the fertilized eggs. 3. Penicillin is bound by some component of the sea urchin egg in amounts sufficiently large to lower the initial concentration, this binding apparently not being related to the inhibitory action.

The relation between ionized calcium and cortical granule exocytosis in eggs of the sea urchin Echinus esculentus

1980 ◽  
Vol 207 (1167) ◽  
pp. 149-161 ◽  
Keyword(s):  
Molecular Mass ◽  
Electric Fields ◽  
Sea Urchin ◽  
Cortical Granule ◽  
Free Calcium ◽  
Relative Molecular Mass ◽  
Cortical Granules ◽  
Full Complement ◽  
Free Calcium Concentration ◽  
Cortical Granule Exocytosis

By subjecting sea urchin eggs to intense, short-duration, electric fields the permeability to low relative molecular mass substances is markedly increased. After such treatment, the extracellular space markers 22 Na + and [ 14 C]mannitol penetrate into the interior of the egg and localized destruction of the oolemma is apparent. The technique permits the rapid introduction of low relative molecular mass substances into the interior of the egg. We have employed it to investigate the efficacy of various buffered calcium concentrations in bringing about exocytosis of cortical granules of the egg. Eggs rendered permeable in the presence of EGTA (free Ca < 10 -8 M) retain a full complement of cortical granules and appear little different in cortical ultrastructure from unfertilized eggs, as judged by scanning and transmission electron microscopy. The proportion of cortical granules remaining in the egg cortex 30 s after application of an electric field in the presence of higher concentrations of calcium decreases pro­gressively as the free calcium concentration introduced into the egg interior is increased from 0.5 to 6 μM. The disappearance of the cortical granules is attributed to their having undergone exocytosis, since the changes in cortical ultrastructure that result from treatment with micro­-molar calcium concentrations are demonstrated to be similar to the changes that result from exocytosis of the cortical granules in intact eggs after fertilization.

scholarly journals Studies on Sulfhydryl Groups during Cell Division of Sea Urchin Egg

1962 ◽  
Vol 45 (3) ◽  
pp. 427-438 ◽  
Author(s):  
Hikoichi Sakai
Keyword(s):  
Cell Division ◽  
Sea Urchin ◽  
Soluble Fraction ◽  
Water Soluble ◽  
The Other ◽  
Original Sample ◽  
Water Soluble Fraction ◽  
Sea Urchin Egg ◽  
Soluble Fractions

The contractility of the thread model prepared from the KCl-soluble proteins of the egg and in vivo factors for the contraction are investigated in Hemicentrotus, Anthocidaris, and Pseudocentrotus eggs. The contractility of the thread model induced by metal ions or cystine changes during development in the characteristic pattern of high at the metaphase and low at the monaster and the interkinetic stages. The change in contractility is paralleled by the change in the —SH content of the protein. The water-soluble fraction of the eggs has activity in causing contraction of the thread model. This activity changes during development in the same way as the contractility itself. The contraction of the thread induced by the water-soluble fractions is accompanied by a decrease in the —SH content of the thread. The activity of the water-soluble fraction in inducing the contraction is proportional to its ability to decrease the number of —SH groups. On boiling, the activity is largely destroyed. The activity is due to two components, one being non-dialyzable and the other dialyzable. Separately each component has little effect, but when mixed, the activity of the original sample is completely restored.

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