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© The Rockefeller University Press, 0021-9525/2000//441 $5.00
The Journal of Cell Biology, Volume 148, Number 3, , 2000 441-452

Original Article

Coordinated Spindle Assembly and Orientation Requires Clb5p-Dependent Kinase in Budding Yeast



Marisa Segala, Duncan J. Clarkea, Paul Maddoxb, E.D. Salmonb, Kerry Bloomb, and Steven I. Reeda

a Department of Molecular Biology, MB7, The Scripps Research Institute, La Jolla, California, 92037
b Department of Biology, CB3280, University of North Carolina, Chapel Hill, North Carolina, 27599
Department of Molecular Biology, MB7, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, CA 92037.(858) 784-2781(858) 784-9836

sreed{at}scripps.edu

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 positioning defect that results in an undivided nucleus entering the bud. Based on time-lapse digital imaging microscopy of microtubules labeled with green 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, both poles were equally likely to interact with the bud cell cortex. Persistent dynamic interactions with the bud ultimately led to spindle translocation across the bud neck. Thus, the mutant failed to assign one spindle pole body the task of organizing astral microtubules towards the mother cell. Our data suggest that Clb5p-associated kinase is required to confer mother-bound behavior to one pole in order to establish correct spindle polarity. In contrast, B-type cyclins, Clb3p and Clb4p, though partially redundant with Clb5p for an early role in spindle morphogenesis, preferentially promote spindle assembly.

Key Words: spindle polarity • astral microtubules • cell cycle • cyclin • Saccharomyces cerevisiae

© 2000 The Rockefeller University Press

Abbreviations used in this paper: DIC, differential interference contrast; GFP, green fluorescent protein; SPB, spindle pole body.


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