Involvement of a particular species of beta-tubulin (beta3) in conidial development in aspergillus nidulans

Strains of Aspergillus containing the benA22 mutation are resistant to benomyl for vegetative growth but do not produce conidia. To test whether conidiation involved an additional benomyl-sensitive tubulin (i.e., was mediated by a tubulin other than the tubulins coded for by the benA locus), a collection of mutants was produced that formed conidia in the presence of benomyl, i.e., were conidiation-resistant (CR⁻) mutants. We analyzed the tubulins of these CR⁻ mutants using two-dimensional gel electrophoresis and found that the mutants lacked one species of beta-tubulin (designated beta3). We have examined two of these mutants in detail. In crosses with strains containing wild-type tubulins, we found that the absence of the beta3-tubulin co-segregated perfectly with the CR⁻ phenotype. In diploids containing both the benA22 and CR⁻ mutations, we found that the CR⁻ phenotype was recessive and that beta3-tubulin was present on two-dimensional gels of tubulins prepared from these diploids. In another set of crosses, these two CR⁻ strains and seven others were first made auxotrophic for uridine and then crossed against strains that had homologously integrated a plasmid containing an incomplete internal fragment of the beta3-tubulin gene and the pyr4 gene of Neurospora crassa (which confers uridine prototrophy on transformants). If the CR⁻ phenotype were produced by a mutation in a gene distinct from the structural gene for beta3-tubulin (designated the tubC gene), then crossing over should have produced some CR⁺ segregants among the uridine auxotrophic progeny of the second cross. All of the uridine auxotrophs from this type of cross, however, showed the CR⁻ phenotype, suggesting that the mutation in these strains is at or closely linked to the tubC locus. The most obvious explanation of these results is that beta3-tubulin is oridinarily used during conidiation and the presence of this species of beta-tubulin renders conidiation sensitive to benomyl. In the CR⁻ mutants, beta3-tubulin is absent, and in the presence of the benA22 mutation the benomyl-resistant beta1- and/or beta2-tubulin substitutes for beta3 to make conidiation benomyl resistant. We discuss these results and give two models to explain the interactions between these betatubulin species.