Functional Roles of α1-, α2-, β1-, and β2-Tubulins in Vegetative Growth, Microtubule Assembly, and Sexual Reproduction of Fusarium graminearum.

The plant pathogen Fusarium graminearum contains two α-tubulin isotypes (α1 and α2) and two β-tubulin isotypes (β1 and β2). The functional roles of these tubulins in microtubule assembly are not clear. Previous studies reported that α1- and β2-tubulin deletion mutants showed severe growth defects and hypersensitivity to carbendazim, which have not been well explained. Here, we investigated the interaction between α- and β-tubulin of F. graminearum. Colocalization experiments demonstrated that β1- and β2-tubulin are colocalized. Coimmunoprecipitation experiments suggested that β1-tubulin binds to both α1- and α2-tubulin and that β2-tubulin can also bind to α1- or α2-tubulin. Interestingly, deletion of α1-tubulin increased the interaction between β2-tubulin and α2-tubulin. Microtubule observation assays showed that deletion of α1-tubulin completely disrupted β1-tubulin-containing microtubules and significantly decreased β2-tubulin-containing microtubules. Deletion of α2-, β1-, or β2-tubulin had no obvious effect on the microtubule cytoskeleton. However, microtubules in α1- and β2-tubulin deletion mutants were easily depolymerized in the presence of carbendazim. The sexual reproduction assay indicates that α1- and β1-tubulin deletion mutants could not produce asci and ascospores. These results implied that α1-tubulin may be essential for the microtubule cytoskeleton. However, our Δα1-2×α2 mutant (α1-tubulin deletion mutant containing two copies of α2-tubulin) exhibited normal microtubule network, growth, and sexual reproduction. Interestingly, the Δα1-2×α2 mutant was still hypersensitive to carbendazim. In addition, both β1-tubulin and β2-tubulin were found to bind the mitochondrial outer membrane voltage-dependent anion channel (VDAC), indicating that they could regulate the function of VDAC. IMPORTANCE In this study, we found that F. graminearum contains four different α-/β-tubulin heterodimers (α1-/β1-, α1-/β2-, α2-/β1-, and α2-/β2-tubulin heterodimers), and they assemble together into a single microtubule. Moreover, α1- and α2-tubulins are functionally interchangeable in microtubule assembly, vegetative growth, and sexual reproduction. These results provide more insights into the functional roles of different tubulins of F. graminearum, which could be helpful for purification of tubulin heterodimers and development of new tubulin-binding agents.