Coordinated spindle assembly and orientation requires Clb5p-dependent kinase in budding yeast

The orientation of the mitotic spindle along a polarity axis is critical in asymmetric cell divisions. In the budding yeast, Saccharomyces cerevisiae, loss of the S-phase B-type cyclin Clb5p under conditions of limited cyclin -dependent kinase activity (cdc28-4 clb5 Delta cells) causes a spindle posi tioning defect that results in an undivided nucleus entering the bud. Based on time-lapse digital imaging microscopy of microtubules labeled with gree n fluorescent protein fusions to either tubulin or dynein, we observed that the asymmetric behavior of the spindle pole bodies during spindle assembly was lost in the cdc28-4 clb5 Delta cells. As soon as a spindle formed, bot h poles were equally likely to interact with the bud cell cortex. Persisten t dynamic interactions with the bud ultimately led to spindle translocation across the bud neck. Thus, the mutant failed to assign one spindle pole bo dy the task of organizing astral microtubules towards the mother cell. Our data suggest that Clb5p-associated kinase is required to confer mother-boun d behavior to one pole in order to establish correct spindle polarity. In c ontrast, B-type cyclins, Clb3p and Clb4p, though partially redundant with C lb5p for an early role in spindle morphogenesis, preferentially promote spi ndle assembly.