The role of astral microtubules in conjugate division in the dikaryon of Coprinus cinereus

We used fluorescence microscopy to examine the positioning of the two nuclei, the configuration of the mitotic apparatus, and the site of septation during conjugate division in eight dikaryons homozygous for either one α-or one of the seven β-tubulin mutations as well as the parental wild-type dikaryon of Coprinus cinereus. In the wild-type dikaryon, more than 90% of the mitotic apparatus had distinct asters, whereas in the mutants the frequency of the apparatus with distinct asters was more or less lowered. In one of the β-tubulin mutants, BEN 193, the defect of astral microtubules was most remarkable and 75% of the mitotic apparatus lacked astral microtubules, although spindle formation and elongation occurred normally. In BEN 193, the two nuclei were positioned normally during conjugate division in the majority of hyphal cells examined, despite the defect of astral microtubules; the leading nucleus divided in the clamp and the second nucleus in the main hypha just beneath the clamp. This result provided evidence against a direct involvement of astral microtubules in the positioning of the two nuclei during conjugate division. In BEN 193, however, septation in the clamp was disturbed in spite of the fact that a mitotic nucleus was positioned normally in the clamp. In the mutant, the actin ring, which forms prior to septation, was also disturbed in its angle in the clamp. These results strongly suggested that astral microtubules control septation in the clamp.