Characterization of two β-tubulin genes from Geotrichum candidum

Abstract

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.