Skip to main content
NCBI home page
The Plant Cell logoLink to The Plant Cell
. 1994 Feb;6(2):303–314. doi: 10.1105/tpc.6.2.303

gamma-Tubulin in Arabidopsis: gene sequence, immunoblot, and immunofluorescence studies.

B Liu 1, H C Joshi 1, T J Wilson 1, C D Silflow 1, B A Palevitz 1, D P Snustad 1
PMCID: PMC160435  PMID: 8148650

Abstract

gamma-Tubulin is a protein associated with microtubule (Mt)-organizing centers in a variety of eukaryotic cells. Unfortunately, little is known about such centers in plants. Genomic and partial cDNA clones encoding two gamma-tubulins of Arabidopsis were isolated and sequenced. Comparisons of genomic and cDNA sequences showed that both genes, TubG1 and TubG2, contain nine introns at conserved locations. The sequences of the two genes both predict proteins containing 474 amino acids, with molecular masses of 53,250 and 53,280 D, respectively. The predicted gamma 1- and gamma 2-tubulins exhibit 98% amino acid identity with each other and approximately 70% amino acid identity with the gamma-tubulins of animals and fungi. RNA gel blot results demonstrated that both genes are transcribed in suspension culture cells, seedlings, and roots and flowers of mature plants. Immunoblots of Arabidopsis proteins using an antibody specific to a conserved peptide of gamma-tubulin showed a major cross-reacting polypeptide with an M(r) of 58,000. The same antibody stained all Mt arrays in tissue and suspension culture cells of this species. Binding was inhibited by the homologous oligopeptide in the gamma-tubulins predicted by the two Arabidopsis gene sequences. Antibody staining avoided the plus ends of Mts at the kinetochores and cell plate, but unlike the case in animal cells, seemed to be localized over broad stretches of the kinetochore fibers and phragmoplast toward the minus ends. We concluded that at least two gamma-tubulin protein homologs are present in Arabidopsis and that at least one of them is localized along Mt arrays. Its distribution is correlated with and may help explain unique characteristics of Mt organization in plants.

Full Text

The Full Text of this article is available as a PDF (2.6 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bajer A. S., Molè-Bajer J. Reorganization of microtubules in endosperm cells and cell fragments of the higher plant Haemanthus in vivo. J Cell Biol. 1986 Jan;102(1):263–281. doi: 10.1083/jcb.102.1.263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Berry J. O., Nikolau B. J., Carr J. P., Klessig D. F. Transcriptional and post-transcriptional regulation of ribulose 1,5-bisphosphate carboxylase gene expression in light- and dark-grown amaranth cotyledons. Mol Cell Biol. 1985 Sep;5(9):2238–2246. doi: 10.1128/mcb.5.9.2238. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cande W. Z. Centrosomes: composition and reproduction. Curr Opin Cell Biol. 1990 Apr;2(2):301–305. doi: 10.1016/0955-0674(90)90023-8. [DOI] [PubMed] [Google Scholar]
  4. Carpenter J. L., Ploense S. E., Snustad D. P., Silflow C. D. Preferential expression of an alpha-tubulin gene of Arabidopsis in pollen. Plant Cell. 1992 May;4(5):557–571. doi: 10.1105/tpc.4.5.557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chevrier V., Komesli S., Schmit A. C., Vantard M., Lambert A. M., Job D. A monoclonal antibody, raised against mammalian centrosomes and screened by recognition of plant microtubule organizing centers, identifies a pericentriolar component in different cell types. J Cell Sci. 1992 Apr;101(Pt 4):823–835. doi: 10.1242/jcs.101.4.823. [DOI] [PubMed] [Google Scholar]
  6. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  7. Fuchs U., Moepps B., Maucher H. P., Schraudolf H. Isolation, characterization and sequence of a cDNA encoding gamma-tubulin protein from the fern Anemia phyllitidis L. Sw. Plant Mol Biol. 1993 Nov;23(3):595–603. doi: 10.1007/BF00019306. [DOI] [PubMed] [Google Scholar]
  8. Gueth-Hallonet C., Antony C., Aghion J., Santa-Maria A., Lajoie-Mazenc I., Wright M., Maro B. gamma-Tubulin is present in acentriolar MTOCs during early mouse development. J Cell Sci. 1993 May;105(Pt 1):157–166. doi: 10.1242/jcs.105.1.157. [DOI] [PubMed] [Google Scholar]
  9. Hu N., Messing J. The making of strand-specific M13 probes. Gene. 1982 Mar;17(3):271–277. doi: 10.1016/0378-1119(82)90143-3. [DOI] [PubMed] [Google Scholar]
  10. Joshi H. C., Palacios M. J., McNamara L., Cleveland D. W. Gamma-tubulin is a centrosomal protein required for cell cycle-dependent microtubule nucleation. Nature. 1992 Mar 5;356(6364):80–83. doi: 10.1038/356080a0. [DOI] [PubMed] [Google Scholar]
  11. Kalt A., Schliwa M. Molecular components of the centrosome. Trends Cell Biol. 1993 Apr;3(4):118–128. doi: 10.1016/0962-8924(93)90174-y. [DOI] [PubMed] [Google Scholar]
  12. Kimble M., Kuriyama R. Functional components of microtubule-organizing centers. Int Rev Cytol. 1992;136:1–50. doi: 10.1016/s0074-7696(08)62049-5. [DOI] [PubMed] [Google Scholar]
  13. Kopczak S. D., Haas N. A., Hussey P. J., Silflow C. D., Snustad D. P. The small genome of Arabidopsis contains at least six expressed alpha-tubulin genes. Plant Cell. 1992 May;4(5):539–547. doi: 10.1105/tpc.4.5.539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  15. Lambert A. M. Microtubule-organizing centers in higher plants. Curr Opin Cell Biol. 1993 Feb;5(1):116–122. doi: 10.1016/s0955-0674(05)80016-x. [DOI] [PubMed] [Google Scholar]
  16. Liu B., Marc J., Joshi H. C., Palevitz B. A. A gamma-tubulin-related protein associated with the microtubule arrays of higher plants in a cell cycle-dependent manner. J Cell Sci. 1993 Apr;104(Pt 4):1217–1228. doi: 10.1242/jcs.104.4.1217. [DOI] [PubMed] [Google Scholar]
  17. McDonald A. R., Liu B., Joshi H. C., Palevitz B. A. Gamma-tubulin is associated with a cortical-microtubule-organizing zone in the developing guard cells of Allium cepa L. Planta. 1993;191(3):357–361. doi: 10.1007/BF00195693. [DOI] [PubMed] [Google Scholar]
  18. Melki R., Vainberg I. E., Chow R. L., Cowan N. J. Chaperonin-mediated folding of vertebrate actin-related protein and gamma-tubulin. J Cell Biol. 1993 Sep;122(6):1301–1310. doi: 10.1083/jcb.122.6.1301. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Oakley C. E., Oakley B. R. Identification of gamma-tubulin, a new member of the tubulin superfamily encoded by mipA gene of Aspergillus nidulans. Nature. 1989 Apr 20;338(6217):662–664. doi: 10.1038/338662a0. [DOI] [PubMed] [Google Scholar]
  20. Olszewski N. E., Martin F. B., Ausubel F. M. Specialized binary vector for plant transformation: expression of the Arabidopsis thaliana AHAS gene in Nicotiana tabacum. Nucleic Acids Res. 1988 Nov 25;16(22):10765–10782. doi: 10.1093/nar/16.22.10765. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Oppenheimer D. G., Haas N., Silflow C. D., Snustad D. P. The beta-tubulin gene family of Arabidopsis thaliana: preferential accumulation of the beta 1 transcript in roots. Gene. 1988;63(1):87–102. doi: 10.1016/0378-1119(88)90548-3. [DOI] [PubMed] [Google Scholar]
  22. Palacios M. J., Joshi H. C., Simerly C., Schatten G. Gamma-tubulin reorganization during mouse fertilization and early development. J Cell Sci. 1993 Feb;104(Pt 2):383–389. doi: 10.1242/jcs.104.2.383. [DOI] [PubMed] [Google Scholar]
  23. Palevitz B. A. Morphological Plasticity of the Mitotic Apparatus in Plants and Its Developmental Consequences. Plant Cell. 1993 Sep;5(9):1001–1009. doi: 10.1105/tpc.5.9.1001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Raff J. W., Kellogg D. R., Alberts B. M. Drosophila gamma-tubulin is part of a complex containing two previously identified centrosomal MAPs. J Cell Biol. 1993 May;121(4):823–835. doi: 10.1083/jcb.121.4.823. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Rose M. D., Biggins S., Satterwhite L. L. Unravelling the tangled web at the microtubule-organizing center. Curr Opin Cell Biol. 1993 Feb;5(1):105–115. doi: 10.1016/s0955-0674(05)80015-8. [DOI] [PubMed] [Google Scholar]
  26. Smirnova E. A., Bajer A. S. Spindle poles in higher plant mitosis. Cell Motil Cytoskeleton. 1992;23(1):1–7. doi: 10.1002/cm.970230102. [DOI] [PubMed] [Google Scholar]
  27. Snustad D. P., Haas N. A., Kopczak S. D., Silflow C. D. The small genome of Arabidopsis contains at least nine expressed beta-tubulin genes. Plant Cell. 1992 May;4(5):549–556. doi: 10.1105/tpc.4.5.549. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Stearns T., Evans L., Kirschner M. Gamma-tubulin is a highly conserved component of the centrosome. Cell. 1991 May 31;65(5):825–836. doi: 10.1016/0092-8674(91)90390-k. [DOI] [PubMed] [Google Scholar]
  29. Valvekens D., Van Montagu M., Van Lijsebettens M. Agrobacterium tumefaciens-mediated transformation of Arabidopsis thaliana root explants by using kanamycin selection. Proc Natl Acad Sci U S A. 1988 Aug;85(15):5536–5540. doi: 10.1073/pnas.85.15.5536. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Vieira J., Messing J. Production of single-stranded plasmid DNA. Methods Enzymol. 1987;153:3–11. doi: 10.1016/0076-6879(87)53044-0. [DOI] [PubMed] [Google Scholar]
  31. Villemur R., Joyce C. M., Haas N. A., Goddard R. H., Kopczak S. D., Hussey P. J., Snustad D. P., Silflow C. D. Alpha-tubulin gene family of maize (Zea mays L.). Evidence for two ancient alpha-tubulin genes in plants. J Mol Biol. 1992 Sep 5;227(1):81–96. doi: 10.1016/0022-2836(92)90683-b. [DOI] [PubMed] [Google Scholar]
  32. Winey M., Hoyt M. A., Chan C., Goetsch L., Botstein D., Byers B. NDC1: a nuclear periphery component required for yeast spindle pole body duplication. J Cell Biol. 1993 Aug;122(4):743–751. doi: 10.1083/jcb.122.4.743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Zheng Y., Jung M. K., Oakley B. R. Gamma-tubulin is present in Drosophila melanogaster and Homo sapiens and is associated with the centrosome. Cell. 1991 May 31;65(5):817–823. doi: 10.1016/0092-8674(91)90389-g. [DOI] [PubMed] [Google Scholar]

Articles from The Plant Cell are provided here courtesy of Oxford University Press

RESOURCES