Centrosomal components immunologically related to tektins from ciliary and flagellar microtubules.

Centrosomes are critical for the nucleation and organization of the microtubule cytoskeleton during both interphase and cell division. Using antibodies raised against sea urchin sperm flagellar microtubule proteins, we characterize here the presence and behavior of certain components associated with centrosomes of the surf clam Spisula solidissima and cultured mammalian cells. A Sarkosyl detergent-resistant fraction of axonemal microtubules was isolated from sea urchin sperm flagella and used to produce monoclonal antibodies, 16 of which were specific- or cross-specific for the major polypeptides associated with this microtubule fraction: tektins A, B and C, acetylated alpha-tubulin, and 77 and 83 kDa polypeptides. By 2-D isoelectric focussing/SDS polyacrylamide gel electrophoresis the tektins separate into several polypeptide spots. Identical spots were recognized by monoclonal and polyclonal antibodies against a given tektin, indicating that the different polypeptide spots are isoforms or modified versions of the same protein. Four independently derived monoclonal anti-tektins were found to stain centrosomes of S. solidissima oocytes and CHO and HeLa cells, by immunofluorescence microscopy. In particular, the centrosome staining of one monoclonal antibody specific for tektin B (tekB3) was cell-cycle-dependent for CHO cells, i.e. staining was observed only from early prometaphase until late anaphase. By immuno-electron microscopy tekB3 specifically labeled material surrounding the centrosome, whereas a polyclonal anti-tektin B recognized centrioles as well as the centrosomal material throughout the cell cycle. Finally, by immunoblot analysis tekB3 stained polypeptides of 48-50 kDa in isolated spindles and centrosomes from CHO cells.

[1]  M. Kirschner,et al.  Pericentrin, a highly conserved centrosome protein involved in microtubule organization , 1994, Cell.

[2]  J. Finch,et al.  A spacer protein in the Saccharomyces cerevisiae spindle poly body whose transcript is cell cycle-regulated , 1993, The Journal of cell biology.

[3]  L. Amos,et al.  Tektin B1 from ciliary microtubules: primary structure as deduced from the cDNA sequence and comparison with tektin A1. , 1993, Journal of cell science.

[4]  A. Quaroni,et al.  Identification of a 102 kDa protein (cytocentrin) immunologically related to keratin 19, which is a cytoplasmically derived component of the mitotic spindle pole. , 1993, Journal of cell science.

[5]  D. Meyer,et al.  Centractin is an actin homologue associated with the centrosome , 1992, Nature.

[6]  L. Amos,et al.  Primary structure of tektin A1: comparison with intermediate-filament proteins and a model for its association with tubulin. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[7]  R. Linck,et al.  Evidence for a non-tubulin spindle matrix and for spindle components immunologically related to tektin filaments. , 1992, Journal of cell science.

[8]  C. Sellitto,et al.  Heterogeneity of microtubule organizing center components as revealed by monoclonal antibodies to mammalian centrosomes and to nucleus-associated bodies from dictyostelium. , 1992, Cell motility and the cytoskeleton.

[9]  M. Kimble,et al.  Functional components of microtubule-organizing centers. , 1992, International review of cytology.

[10]  M. Yanagida,et al.  The fission yeast gamma-tubulin is essential for mitosis and is localized at microtubule organizing centers. , 1991, Journal of cell science.

[11]  Yixian Zheng,et al.  γ-Tubulin is present in Drosophila melanogaster and homo sapiens and is associated with the centrosome , 1991, Cell.

[12]  M. Kirschner,et al.  γ-Tubulin is a highly conserved component of the centrosome , 1991, Cell.

[13]  M. Bornens,et al.  Microtubular spindle and centrosome structures during the cell cycle in a dinoflagellate Crypthecodinium cohnii B.: an immunocytochemical study. , 1991, Bio Systems.

[14]  C. Petzelt,et al.  Murine cDNAs coding for the centrosomal antigen centrosomin A. , 1991, Journal of cell science.

[15]  M. Bornens,et al.  A centrosomal antigen localized on intermediate filaments and mitotic spindle poles. , 1990, Journal of cell science.

[16]  B. Oakley,et al.  γ-tubulin is a component of the spindle pole body that is essential for microtubule function in Aspergillus nidulans , 1990, Cell.

[17]  B. Oakley,et al.  Identification of γ-tubulin, a new member of the tubulin superfamily encoded by mipA gene of Aspergillus nidulans , 1989, Nature.

[18]  R. E. Stephens,et al.  Retention of ciliary ninefold structure after removal of microtubules. , 1989, Journal of cell science.

[19]  R. Linck,et al.  Multiple immunoblot: A sensitive technique to stain proteins and detect multiple antigens on a single two‐dimensional replica , 1989, Electrophoresis.

[20]  R. Kuriyama,et al.  225-Kilodalton phosphoprotein associated with mitotic centrosomes in sea urchin eggs. , 1989, Cell motility and the cytoskeleton.

[21]  R. Linck,et al.  Relationship between tektins and intermediate filament proteins: an immunological study. , 1989, Cell motility and the cytoskeleton.

[22]  J. Salisbury,et al.  Identification and localization of a novel, cytoskeletal, centrosome- associated protein in PtK2 cells , 1988, The Journal of cell biology.

[23]  G. Schatten,et al.  Acetylated α-tubulin in microtubules during mouse fertilization and early development , 1988 .

[24]  R. Linck,et al.  Evidence for tektins in centrioles and axonemal microtubules. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[25]  G. Schatten,et al.  Microtubules are required for centrosome expansion and positioning while microfilaments are required for centrosome separation in sea urchin eggs during fertilization and mitosis. , 1988, Cell motility and the cytoskeleton.

[26]  K. Ohta,et al.  51-kd protein, a component of microtubule-organizing granules in the mitotic apparatus involved in aster formation in vitro. , 1988, Cell motility and the cytoskeleton.

[27]  C. Sellitto,et al.  Distribution of pericentriolar material in multipolar spindles induced by colcemid treatment in Chinese hamster ovary cells. , 1988, Journal of cell science.

[28]  G. Schatten,et al.  Centrosome detection in sea urchin eggs with a monoclonal antibody against Drosophila intermediate filament proteins: characterization of stages of the division cycle of centrosomes. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[29]  R. Linck,et al.  Characterization of antibodies as probes for structural and biochemical studies of tektins from ciliary and flagellar microtubules. , 1987, Journal of cell science.

[30]  O. H. Griffith,et al.  Pitfalls of immunogold labeling: analysis by light microscopy, transmission electron microscopy, and photoelectron microscopy. , 1987, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[31]  P. Matsudaira,et al.  Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. , 1987, The Journal of biological chemistry.

[32]  G. Piperno,et al.  Cross-reactivity of antibodies specific for flagellar tektin and intermediate filament subunits , 1987, The Journal of cell biology.

[33]  R. E. Stephens,et al.  Biochemical characterization of tektins from sperm flagellar doublet microtubules , 1987, The Journal of cell biology.

[34]  M. Bornens,et al.  Structural and chemical characterization of isolated centrosomes. , 1987, Cell motility and the cytoskeleton.

[35]  S. Dunn,et al.  Effects of the modification of transfer buffer composition and the renaturation of proteins in gels on the recognition of proteins on Western blots by monoclonal antibodies. , 1986, Analytical biochemistry.

[36]  M. Kirschner,et al.  [26] Isolation of mammalian centrosomes , 1986 .

[37]  G. Piperno,et al.  Monoclonal antibodies specific for an acetylated form of alpha-tubulin recognize the antigen in cilia and flagella from a variety of organisms , 1985, The Journal of cell biology.

[38]  E. Nigg,et al.  Cyclic-AMP-dependent protein kinase type II is associated with the Golgi complex and with centrosomes , 1985, Cell.

[39]  L. Amos,et al.  Localization of tektin filaments in microtubules of sea urchin sperm flagella by immunoelectron microscopy , 1985, The Journal of cell biology.

[40]  G. Borisy,et al.  Phosphoproteins are components of mitotic microtubule organizing centers. , 1984, Proceedings of the National Academy of Sciences of the United States of America.

[41]  G. Borisy,et al.  The mitotic spindle of Chinese hamster ovary cells isolated in taxol-containing medium. , 1984, Journal of cell science.

[42]  R. Ueda,et al.  Monoclonal antibody against microtubule associated protein-1 produces immunofluorescent spots in the nucleus and centrosome of cultured mammalian cells. , 1983, Cell structure and function.

[43]  M. Dym,et al.  Colchicine-induced changes in the cytoskeleton of the golden-mantled ground squirrel (Spermophilus lateralis) Sertoli cells. , 1983, The American journal of anatomy.

[44]  G. Langevin,et al.  Structure and chemical composition of insoluble filamentous components of sperm flagellar microtubules. , 1982, Journal of cell science.

[45]  J. Weatherbee,et al.  Localization of high molecular weight microtubule-associated proteins (MAP1 and MAP2) in a HeLa microtubule-organizing centre. , 1982, Cytobios.

[46]  H. Towbin,et al.  Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[47]  R. Goldman,et al.  Biochemical and immunological analysis of rapidly purified 10-nm filaments from baby hamster kidney (BHK-21) cells , 1978, The Journal of cell biology.

[48]  M. Berns,et al.  Continuation of mitosis after selective laser microbeam destruction of the centriolar region , 1977, The Journal of cell biology.

[49]  G. Borisy,et al.  The pericentriolar material in Chinese hamster ovary cells nucleates microtubule formation , 1977, The Journal of cell biology.

[50]  G. Piperno,et al.  Two-dimensional analysis of flagellar proteins from wild-type and paralyzed mutants of Chlamydomonas reinhardtii. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[51]  M. Berns,et al.  The absence of centrioles from spindle poles of rat kangaroo (PtK2) cells undergoing meiotic-like reduction division in vitro , 1977, The Journal of cell biology.

[52]  G. Borisy,et al.  Structural polarity and directional growth of microtubules of Chlamydomonas flagella. , 1974, Journal of molecular biology.

[53]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.