Summary
The β-tubulin genes Gβ1 and Gβ2 from the phytopathogenic hemiascomycete Geotrichum candidum were found to be highly diverged in amino acid sequence from those of other filamentous fungi. Gβ1 and Gβ2 were also divergent from each other, with the coding regions sharing only 66% nucleotide sequence homology and 64% amino acid identity. However, the proteins shared 82% similarity and only 25 of the 161 non-identical amino acid substitutions were non-conservative. The organization of Gβ1 is similar to other fungal β-tubulin genes, but Gβ2 has several unusual features; it has 2 amino acid additions in the N-terminal 40 residues and must employ an uncommon 5′ splice junction sequence in preference to an overlapping perfect consensus. The amino acid change found to confer benomyl resistance in Neurospora crassa was also present in Gβ2. Gβ1 has four introns which are located similarly to those of β-tubulin genes in other fungi. Gβ2, however, has a single intron in a unique location. Translational fusions employing the 5′ non-coding regions of the two Geotrichum β-tubulin genes were made with the hygromycin phosphotransferase gene and shown to function in Schizosaccharomyces pombe and Trichoderma hamatum. However, G. candidum could not be transformed with these or other tested plasmids commonly used for fungal transformation.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (1987) Current protocols in molecular biology. Greene Publishing Associates and Wiley-Interscience, New York
Ballance DJ (1986) Sequences important for gene expression in filamentous fungi. Yeast 2:229–236
Beach D, Nurse P (1981) High-frequency transformation of the fission yeast Schizosaccharomyces pombe. Nature 290:140–142
Butler E, Fogle D (1988) Galactomyces citri-aurantii a newly found teleomorph of Geotrichum citri-aurantii the cause of sour rot of citrus fruit. Mycotaxon 33:197–212
Crouse GF, Frischauf A, Lehrach H (1983) An integrated and simplified approach to cloning into plasmids and singlestranded phages. Methods Enzymol 101:78–89
Davidse LC, Flach W (1977) Differential binding of methyl benzimidazol-2-yl carbamate to fungal tubulin as a mechanism of resistance to this antimitotic agent in mutant strains of Aspergillus nidulans. J Cell Biol 72:174–193
Devereux J, Haeberli P, Smithies O (1984) A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res 12:387–395
Elwood HJ, Olson GJ, Sogin ML (1985) The small subunit ribosomal RNA gene structure from the hypotrichous ciliates Oxytricha nova and Stylonchia pustulata. Mol Biol Evol 2:399–410
Frischauf A-M, Lehrach H, Poustka A, Murray N (1983) Lambda replacement vectors carrying polylinker sequences. J Mol Biol 170:827–842
Gallwitz D, Halfter H, Mertins P (1987) Splicing of mRNA precursors in yeast. In: Kinghorn JR (ed) Gene structure in eukaryotic microbes. Special Publications of the Society for General Microbiology, Oxford, Washington DC, pp 27–40
Gurr SJ, Unkles SE, Kinghorn JR (1987) The structure and organization of nuclear genes of filamentous fungi. In: Kinghorn JR (ed) Gene structure in eukaryotic microbes. Special Publications of the Society for General Microbiology, Oxford, Washington DC, pp 93–139
Harper JF, Mages W (1988) Organization and structure of Volvox β-tubulin genes. Mol Gen Genet 213:315–324
Henikoff S (1984) Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. Gene 28:351–359
Hiraoka Y, Toda T, Yanagida M (1984) The NDA3 gene of fission yeast encodes β-tubulin: a cold-sensitive nda3 mutation reversibly blocks spindle formation and chromosome movement in mitosis. Cell 39:349–358
Kaster KR, Burgett SG, Ingolia TD (1984) Hygromycin B resistance as dominant selectable marker in yeast. Curr Genet 8:353–358
Keen NT, Tamaki S, Kobayashi D, Trollinger D (1988) Improved broad-host-range plasmids for cloning in Gram-negative bacteria. Gene 70:191–197
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York
May GS, Gambino J, Weatherbee JA, Morris NR (1985) Identification and functional analysis of beta-tubulin genes by site specific integrative transformation in Aspergillus nidulans. J Cell Biol 101:712–719
May GS, Tsang L-S, Smith H, Fidel S, Morris NR (1987) Aspergillus nidulans β-tubulin genes are unusually divergent. Gene 55:231–243
May GS, Waring RB, Morris NR (1990) Increasing tubC β-tubulin synthesis by placing it under the control of a benA β-tubulin upstream sequence causes a reduction in benA β-tubulin level but has no effect on microtubule function. Cell Motil Cytoskeleton 16:214–220
Mitchison JM (1970) Physiological and cytological methods for Schizosaccharomyces pombe. Methods Cell Physiol 4:131–165
Neff NF, Thomas JH, Grisafi P, Botstein D (1983) Isolation of the β-tubulin gene from yeast and demonstration of its essential function in vivo. Cell 33:211–219
Orbach MJ, Porro EB, Yanofsky C (1986) Cloning and characterization of the gene for β-tubulin from a benomyl-resistant mutant of Neurospora crassa and its use as a dominant selectable marker. Mol Cell Biol 6:2452–2461
Panaccione DG, Hanau RM (1990) Characterization of two divergent β-tubulin genes from Colletotrichum graminicola. Gene 86:163–170
Pustell J, Kafatos FC (1984) A convenient and adaptable package of computer programs for DNA and protein sequence management, analysis and homology determination. Nucleic Acids Res 12:643–655
Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467
Sharp PM, Li W-H (1986) An evolutionary perspective on synonymous codon usage in unicellular organisms. J Mol Evol 24:28–38
Sharp PM, Tuohy TMF, Mosurski KR (1986) Codon usage in yeast: cluster analysis clearly differentiates highly and lowly expressed genes. Nucleic Acids Res 14:5125–5143
Sherwood JE, Sommerville SC (1989) Molecular characterization of a β-tubulin gene from Erysiphe graminis. Phytopathology 79:1205 Abstract
Smith HA, Gorman JW, Koltin Y, Gorman JA (1990) Functional expression of the Candida albicans β-tubulin gene in Saccharomyces cerevisiae. Gene 90:115–123
Sogin ML, Miotto K, Miller L (1986) Primary structure of the Neurospora crassa small subunit ribosomal RNA coding region. Nucleic Acids Res 14:9540
Sorensen MA, Kurland CG, Pederson S (1989) Codon usage determines translation rate in Escherichia coli. J Mol Biol 207:365–377
Tamaki SJ, Gold S, Robeson M, Manulis S, Keen NT (1988) Structure and organization of the pel genes from Erwinia chrysanthemi EC16. J Bacteriol 170:3468–3478
Turgeon BG, Garber RC, Yoder OC (1987) Development of a fungal transformation system based on selection of sequences with promoter activity. Mol Cell Biol 7:3297–3305
Vieira J, Messing J (1987) Production of single-stranded plasmid DNA. Methods Enzymol 153:3–11
Vollmer SJ, Yanofsky C (1986) Efficient cloning of genes of Neurospora crassa. Proc Natl Acad Sci USA 83:4869–4873
Weatherbee JA, Morris NR (1984) Aspergillus contains multiple tubulin genes. J Biol Chem 259:15452–15459
Yoder OC (1988) Cochhobolus heterostrophus, cause of corn leaf blight. Adv Plant Pathol 6:93–112
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Gold, S.E., Casale, W.L. & Keen, N.T. Characterization of two β-tubulin genes from Geotrichum candidum . Molec. Gen. Genet. 230, 104–112 (1991). https://doi.org/10.1007/BF00290657
Received:
Revised:
Issue Date:
DOI: https://doi.org/10.1007/BF00290657