A switch in microtubule dynamics at the onset of anaphase B in the mitotic spindle of Schizosaccharomyces pombe

Abstract

Microtubule dynamics have key roles in mitotic spindle assembly and chromosome movement [1]. Fast turnover of spindle microtubules at metaphase and polewards flux of microtubules (polewards movement of the microtubule lattice with depolymerization at the poles) at both metaphase and anaphase have been observed in mammalian cells [2]. Imaging spindle dynamics in genetically tractable yeasts is now possible using green fluorescent protein (GFP)-tagging of tubulin and sites on chromosomes [3–8]. We used photobleaching of GFP-labeled tubulin to observe microtubule dynamics in the fission yeast Schizosaccharomyces pombe. Photobleaching did not perturb progress through mitosis. Bleached marks made on the spindle during metaphase recovered their fluorescence rapidly, indicating fast microtubule turnover. Recovery was spatially non-uniform, but we found no evidence for polewards flux. Marks made during anaphase B did not recover fluorescence, and were observed to slide away from each other at the same rate as spindle elongation. Fast microtubule turnover at metaphase and a switch to stable microtubules at anaphase suggest the existence of a cell-cycle-regulated molecular switch that controls microtubule dynamics and that may be conserved in evolution. Unlike the situation for vertebrate spindles, microtubule depolymerization at poles and polewards flux may not occur in S. pombe mitosis. We conclude that GFP–tubulin photobleaching in conjunction with mutant cells should aid research on molecular mechanisms causing and regulating dynamics.