The tubB alpha-tubulin gene is essential for sexual development in Aspergillus nidulans.

Abstract

The filamentous fungus Aspergillus nidulans has two genes encoding a-tubulin, tubA and tubB. Mutational analysis of tubA has demonstrated that the tubA gene is essential for mitosis and nuclear migration. In this study we have deleted the tubB gene by replacing it with a selectable marker and have named this new allele tubBA. The results demonstrate that the tubB gene is not required for vegetative growth or asexual reproduction, nor is it required for the initiation or early stages of sexual differentiation. Deletion of tubB, however, completely prevents ascosporogenesis, because tubBA strains produce no sexual spores when self-crossed. These strains produce viable ascospores when outcrossed to tubB + strains, indicating that the tubBA mutation is recessive. We have studied the cytology of sexual development in wild-type strains and in the tubB mutant and have observed that tubBA, strains develop normally to the stage of ascus formation. However, only a single nuclear mass is observed in the tubBA ascus, indicating that either the two zygotic haploid nuclei are blocked in karyogamy or that karyogamy occurs but the resulting diploid nucleus is subsequently blocked in meiosis I.

Novel Sexual-Cycle-Specific Gene Silencing in Aspergillus nidulans

“…tubB encodes a meiosisspecific a-tubulin. Induction of the sexual cycle and development of the fruiting body is normal in a tubBD strain, but development of ascogenous tissue is blocked premeiotically at a stage prior to karyogamy (Kirk and Morris 1991). This phenotype appears microscopically identical to that of our strains expressing a MatIS response.…”

Either alpha-tubulin isogene product is sufficient for microtubule function during all stages of growth and differentiation in Aspergillus nidulans.

“…Although some aspects of ascus maturation appear to take place in tubBA strains, free cell formation never occurs, and only a single nuclear mass is ever observed in the ascus (Fig. 4C and D), indicating that development is blocked either prior to or at meiosis I (24).…”

“…A deletion allele of the tubB gene was previously constructed by eliminating all but the 3' 20% of the tubB coding region (24). This mutation, tubBA, completely prevents the formation of sexual spores (ascosporogenesis) in a self-cross.…”

“…To upregulate expression of tubA in a tubBA strain and potentially simulate the overall level of a-tubulin normally found in wild-type cells carrying both a functional tubA and tubB gene, we used a 2.8-kb genomic fragment upstream of the tubB coding region to act as a promoter for expression of the tubA coding region. We had shown previously that this 2.8-kb fragment sufficiently promotes expression of the tubB coding region, resulting in full complementation of the tubBA allele during sexual development (24). We constructed a plasmid which contained this putative tubB regulatory region joined to the tubA coding region [designated tubB(p)tubA) and which carried the argB gene as a selectable marker; this plasmid was used to transform an A. nidulans tubBA argB2 strain to arginine prototrophy.…”

“…A 2.4-kb BamHI fragment from pDP485 (courtesy of P. Doshi [9]) containing the tubA coding region, 20 Cytological studies. The contents of immature cleistothecia previously (24). Mature ascospores were examined by allowing cleistothecia to incubate at 37°C for 2 weeks, clearing them of hulle cells and conidia, and crushing them directly in fixative between the slide and coverslip.…”