Segal Marisa, Bloom Kerry, Reed Steven I
Department of Molecular Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
Mol Biol Cell. 2002 Dec;13(12):4141-55. doi: 10.1091/mbc.02-05-0067.
Spindle orientation is critical for accurate chromosomal segregation in eukaryotic cells. In the yeast Saccharomyces cerevisiae, orientation of the mitotic spindle is achieved by a program of microtubule-cortex interactions coupled to spindle morphogenesis. We previously implicated Bud6p in directing microtubule capture throughout this program. Herein, we have analyzed cells coexpressing GFP:Bud6 and GFP:Tub1 fusions, providing a kinetic view of Bud6p-microtubule interactions in live cells. Surprisingly, even during the G1 phase, microtubule capture at the recent division site and the incipient bud is dictated by Bud6p. These contacts are eliminated in bud6 delta cells but are proficient in kar9 delta cells. Thus, Bud6p cues microtubule capture, as soon as a new cell polarity axis is established independent of Kar9p. Bud6p increases the duration of interactions and promotes distinct modes of cortical association within the bud and neck regions. In particular, microtubule shrinkage and growth at the cortex rarely occur away from Bud6p sites. These are the interactions selectively impaired at the bud cortex in bud6 delta cells. Finally, interactions away from Bud6p sites within the bud differ from those occurring at the mother cell cortex, pointing to the existence of an independent factor controlling cortical contacts in mother cells after bud emergence.
纺锤体定向对于真核细胞中染色体的精确分离至关重要。在酿酒酵母中，有丝分裂纺锤体的定向是通过与纺锤体形态发生相关的微管 - 皮质相互作用程序实现的。我们之前认为Bud6p在整个程序中指导微管捕获。在此，我们分析了共表达GFP:Bud6和GFP:Tub1融合蛋白的细胞，提供了活细胞中Bud6p - 微管相互作用的动力学视图。令人惊讶的是，即使在G1期，最近分裂位点和起始芽处的微管捕获也由Bud6p决定。这些接触在bud6Δ细胞中消失，但在kar9Δ细胞中仍然存在。因此，一旦建立了独立于Kar9p的新细胞极性轴，Bud6p就会提示微管捕获。Bud6p增加了相互作用的持续时间，并促进了芽和颈部区域内不同的皮质结合模式。特别是，皮质处微管的收缩和生长很少发生在远离Bud6p位点的地方。这些是bud6Δ细胞中芽皮质处选择性受损的相互作用。最后，芽内远离Bud6p位点的相互作用与母细胞皮质处的相互作用不同，这表明在芽出现后母细胞中存在控制皮质接触的独立因子。
