Supplemental Material for Vicars et al., 2024

Chromosome congression and alignment on the metaphase plate involves lateral and microtubule plus-end interactions with the kinetochore. Here we take advantage of our ability to efficiently generate a GFP-marked acentric X chromosome fragment in Drosophila neuroblasts to identify forces acting on chromosome arms that drive congression and alignment. We find acentrics efficiently congress and align on the metaphase plate, often more rapidly than kinetochore-bearing chromosomes. Unlike intact chromosomes, the paired sister acentrics oscillate as they move to and reside on the metaphase plate in a plane distinct and significantly further from the main mass of intact chromosomes. Consequently, at anaphase onset acentrics are oriented either parallel or perpendicular to the spindle. Parallel-oriented sisters separate by sliding while those oriented perpendicularly separate via unzipping. This oscillation, together with the fact that in the presence of spindles with disrupted interpolar microtubules acentrics are rapidly shunted away from the poles, indicates that distributed plus-end directed forces are primarily responsible for acentric migration. This conclusion is supported by the observation that reduction of EB1 preferentially disrupts acentric alignment. Taken together these studies suggest that plus-end forces mediated by the outer interpolar microtubules contribute significantly to acentric congression and alignment. Surprisingly, we observe disrupted telomere pairing and alignment of sister acentrics indicating that the kinetochore is required to ensure proper gene-to-gene alignment of sister chromatids. Finally, we demonstrate that like mammalian cells, the Drosophila congressed chromosomes on occasion exhibit a toroid configuration.

染色质在减数分裂中期板上的凝聚与对齐过程，涉及侧向以及微管加端与着丝粒的相互作用。本研究利用我们在果蝇神经母细胞中高效生成GFP标记的着丝粒染色体片段的能力，旨在识别驱动染色体臂凝聚和对齐的力。我们发现，非着丝粒染色体片段能够高效地在中期板上凝聚和对齐，其速度往往超过携带着丝粒的染色体。与完整染色体不同，成对的姐妹非着丝粒染色体在移动至并驻留在中期板上时，会在一个与完整染色体主体显著更远的平面内发生振荡。因此，在后期开始时，非着丝粒染色体要么与纺锤体平行，要么垂直。平行方向的姐妹染色体通过滑动分离，而垂直方向的染色体则通过解旋分离。这种振荡现象，以及在有纺锤体极间微管中断的情况下，非着丝粒染色体迅速从两极移开的事实，表明分布的加端指向力是着丝粒迁移的主要责任者。这一结论得到了EB1减少优先破坏非着丝粒对齐的观察结果的支持。综合这些研究结果表明，由外极间微管介导的加端力对非着丝粒的凝聚和对齐具有显著的贡献。令人惊讶的是，我们观察到端粒配对和姐妹非着丝粒的对齐受到破坏，这表明着丝粒是确保姐妹染色单体基因到基因正确对齐所必需的。最后，我们证明，与哺乳动物细胞类似，果蝇的凝聚染色体有时也会呈现出环状结构。