The calcium-binding protein cell division cycle 31 of Saccharomyces cerevisiae is a component of the half bridge of the spindle pole body

Cdc31 mutants of Saccharomyces cerevisiae arrest at the nonpermissive temperature with large buds, G2 DNA content and, a single, abnormally large spindle pole body (SPB) (Byers, B. 1981. Molecular Genetics in Yeast. Alfred Benzon Symposium. 16:119-133). In this report, we show that the CDC31 gene product is essential for cell viability. We demonstrate that purified CDC31 protein binds Ca2+ and that this binding is highly specific. Taken together, three lines of evidence indicate that CDC31 is a component of the SPB. First, CDC31 cofractionates with enriched preparations of SPBs. Second, immunofluorescence staining indicates that CDC31 colocalizes with a known SPB component. Third, immunoelectron microscopy with whole cells and with isolated SPBs reveals that CDC31 is localized to the half bridge of the SPB, which lies immediately adjacent to the SPB plaques. CDC31 was detected mainly at the cytoplasmic side of the half bridge and, therefore, defines a further substructure of the SPB. We suggest that CDC31 is a member of a family of calcium-binding, centrosome- associated proteins from a phylogenetically diverse group of organisms.

[1]  Gerald R. Fink,et al.  Guide to yeast genetics and molecular biology , 1993 .

[2]  P. Mirabito,et al.  BIMA, a TPR-containing protein required for mitosis, localizes to the spindle pole body in Aspergillus nidulans , 1993, The Journal of cell biology.

[3]  Y. Anraku,et al.  Mutations in yeast calmodulin cause defects in spindle pole body functions and nuclear integrity , 1992, The Journal of cell biology.

[4]  M. Snyder,et al.  CIK1: a developmentally regulated spindle pole body-associated protein important for microtubule functions in Saccharomyces cerevisiae. , 1992, Genes & development.

[5]  M. Rose,et al.  Separate domains of KAR1 mediate distinct functions in mitosis and nuclear fusion , 1992, The Journal of cell biology.

[6]  G. Fink,et al.  The yeast Ca(2+)-ATPase homologue, PMR1, is required for normal Golgi function and localizes in a novel Golgi-like distribution. , 1992, Molecular biology of the cell.

[7]  T. Roberts,et al.  Asymmetric mitotic segregation of the yeast spindle pole body , 1992, Cell.

[8]  M. Winey,et al.  7 – Spindle Pole Body of Saccharomyces cerevisiae: A Model for Genetic Analysis of the Centrosome Cycle , 1992 .

[9]  D Botstein,et al.  Structure of the yeast endoplasmic reticulum: Localization of ER proteins using immunofluorescence and immunoelectron microscopy , 1991, Yeast.

[10]  M. Bornens,et al.  A human centrosomal protein is immunologically related to basal body- associated proteins from lower eucaryotes and is involved in the nucleation of microtubules , 1991, The Journal of cell biology.

[11]  Andrew W. Murray,et al.  Feedback control of mitosis in budding yeast , 1991, Cell.

[12]  B. Roberts,et al.  S. cerevisiae genes required for cell cycle arrest in response to loss of microtubule function , 1991, Cell.

[13]  M. Winey,et al.  MPS1 and MPS2: novel yeast genes defining distinct steps of spindle pole body duplication , 1991, The Journal of cell biology.

[14]  Y. Anraku,et al.  Cell cycle control by calcium and calmodulin in Saccharomyces cerevisiae. , 1991, Biochimica et biophysica acta.

[15]  T. Davis,et al.  Can calmodulin function without binding calcium? , 1991, Cell.

[16]  M. Boguski,et al.  TPR proteins as essential components of the yeast cell cycle. , 1991, Cold Spring Harbor symposia on quantitative biology.

[17]  M. Rout,et al.  Components of the yeast spindle and spindle pole body , 1990, The Journal of cell biology.

[18]  J. Hendrick,et al.  The purified E. coli integral membrane protein SecY E is sufficient for reconstitution of SecA-dependent precursor protein translocation , 1990, Cell.

[19]  J. Hanover,et al.  Structure and function of the nuclear pore complex: New perspectives , 1990, BioEssays : news and reviews in molecular, cellular and developmental biology.

[20]  R. Sikorski,et al.  A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. , 1989, Genetics.

[21]  S. Fukui,et al.  Saccharomyces cerevisiae protein kinase dependent on Ca2+ and calmodulin , 1989, Journal of bacteriology.

[22]  R. Rothstein.  18 – One-Step Gene Disruption in Yeast , 1989 .

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

[24]  W. J. Dower,et al.  High efficiency transformation of E. coli by high voltage electroporation , 1988, Nucleic Acids Res..

[25]  B. Huang,et al.  Molecular cloning of cDNA for caltractin, a basal body-associated Ca2+- binding protein: homology in its protein sequence with calmodulin and the yeast CDC31 gene product , 1988, The Journal of cell biology.

[26]  E. Harlow,et al.  Antibodies: A Laboratory Manual , 1988 .

[27]  G. Fink,et al.  KAR1, a gene required for function of both intranuclear and extranuclear microtubules in yeast , 1987, Cell.

[28]  B. Byers,et al.  Yeast gene required for spindle pole body duplication: homology of its product with Ca2+-binding proteins. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[29]  F. Studier,et al.  Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. , 1986, Journal of molecular biology.

[30]  D. Botstein,et al.  A gene required for the separation of chromosomes on the spindle apparatus in yeast , 1986, Cell.

[31]  R. Pipe.  Light and electron microscope localization of beta-glucuronidase activity in the stomach and digestive gland of the marine gastropod Littorina littorea. , 1986, The Histochemical journal.

[32]  C. Richardson,et al.  A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[33]  K. Hancock,et al.  Enzyme-Linked Immunoelectrotransfer Blot (EITB) , 1985 .

[34]  M. Melkonian,et al.  Striated flagellar roots: isolation and partial characterization of a calcium-modulated contractile organelle , 1984, The Journal of cell biology.

[35]  J. Pringle,et al.  Relationship of actin and tubulin distribution to bud growth in wild- type and morphogenetic-mutant Saccharomyces cerevisiae , 1984, The Journal of cell biology.

[36]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[37]  R. Rothstein.  One-step gene disruption in yeast. , 1983, Methods in enzymology.

[38]  R. Mortimer,et al.  An endomitotic effect of a cell cycle mutation of Saccharomyces cerevisiae. , 1981, Genetics.

[39]  J. Broach,et al.  The Molecular biology of the yeast saccharomyces, life cycle and inheritance , 1981 .

[40]  B. Byers.  Cytology of the Yeast Life Cycle , 1981 .

[41]  F. Sanger,et al.  DNA sequencing with chain-terminating inhibitors. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[42]  G. Fink,et al.  A mutant of Saccharomyces cerevisiae defective for nuclear fusion. , 1976, Proceedings of the National Academy of Sciences of the United States of America.

[43]  M. M. Bradford.  A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. , 1976, Analytical biochemistry.

[44]  B. Byers,et al.  Behavior of spindles and spindle plaques in the cell cycle and conjugation of Saccharomyces cerevisiae , 1975, Journal of bacteriology.

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