Faculty of Life Sciences, University of Manchester, Michael Smith Building, Oxford Road, Manchester M13 9PT, UK.
J Cell Sci. 2010 Mar 1;123(Pt 5):653-9. doi: 10.1242/jcs.059501. Epub 2010 Feb 2.
Bub1 was one of the first protein kinases identified as a component of the spindle-assembly checkpoint, a surveillance mechanism that delays anaphase onset until all chromosomes are stably attached to spindle microtubules. Whereas the kinase activity of Bub1 is not required for checkpoint function in yeast, its requirement in mammalian cells is still unclear. Using a complementation assay with bona fide BUB1-null mouse embryonic fibroblasts, we show that the kinase activity of Bub1 is not required for checkpoint function or chromosome alignment. Its activity is, however, required for centromeric localisation of Sgo1, a known protector of centromeric cohesion. Despite the absence of Sgo1 from mitotic centromeres in cells devoid of Bub1 activity, centromeric cohesion is still maintained until anaphase. An explanation for this comes from observations showing that Sgo1 is first recruited to centromeric heterochromatin in G2, but then becomes diffusely localised throughout the nucleus in early prophase, before returning to centromeres later in prophase. Importantly, whereas centromeric localisation of Sgo1 in prophase is dependent on the kinase activity of Bub1, its recruitment to centromeric heterochromatin in G2 is not. Rather, the localisation of Sgo1 in G2 is abolished when heterochromatin protein 1 is not bound to centromeric heterochromatin. Thus, it seems that Sgo1 sets up the centromeric protection mechanism in G2, but that its Bub1-dependent localisation to centromeres during mitosis is not required to maintain cohesion.
Bub1 是最早被鉴定为纺锤体组装检查点（spindle-assembly checkpoint）组成部分的蛋白激酶之一，该检查点是一种监控机制，可延迟后期起始，直到所有染色体都稳定连接到纺锤体微管上。尽管 Bub1 的激酶活性在酵母中对于检查点功能不是必需的，但在哺乳动物细胞中仍然不清楚其是否必需。使用与真正的 BUB1 缺失的小鼠胚胎成纤维细胞进行互补测定，我们表明 Bub1 的激酶活性对于检查点功能或染色体排列不是必需的。然而，它的活性对于 Sgo1 的着丝粒定位是必需的，Sgo1 是已知的着丝粒凝聚保护蛋白。尽管在缺乏 Bub1 活性的细胞中，Sgo1 从有丝分裂着丝粒中缺失，但直到后期，着丝粒凝聚仍然保持。一个解释是，观察到 Sgo1 首先在 G2 中被招募到着丝粒异染色质，但随后在早前期中弥散定位到核内，然后在前期后期再返回着丝粒。重要的是，虽然 Sgo1 在前期的着丝粒定位依赖于 Bub1 的激酶活性，但它在 G2 中向着丝粒异染色质的募集不是。相反，当异染色质蛋白 1 未结合到着丝粒异染色质时，Sgo1 在 G2 中的定位就会被消除。因此，似乎 Sgo1 在 G2 中建立了着丝粒保护机制，但在有丝分裂期间 Bub1 依赖性的向着丝粒的定位对于维持凝聚不是必需的。
