Division of Cell and Developmental Biology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.
J Cell Sci. 2010 Mar 1;123(Pt 5):736-46. doi: 10.1242/jcs.060806. Epub 2010 Feb 9.
Mutations in the tumour suppressor Adenomatous polyposis coli (Apc) initiate most sporadic colorectal cancers. Apc is implicated in regulating microtubule (MT) dynamics in interphase and mitosis. However, little is known about the underlying mechanism or regulation of this Apc function. We identified importin-beta as a binding partner of Apc that regulates its effect on MTs. Apc binds importin-beta in vitro and in Xenopus egg extracts, and RanGTP inhibits this interaction. The armadillo-like repeat domain of importin-beta binds to the middle of Apc, where it can compete with beta-catenin. In addition, two independent sites in the C terminus of Apc bind the N-terminal region of importin-beta. Binding to importin-beta reduces the ability of Apc to assemble and bundle MTs in vitro and to promote assembly of microtubule asters in Xenopus egg extracts, but does not affect the binding of Apc to MTs or to EB1. Depletion of Apc decreases the formation of cold-stable spindles in Xenopus egg extracts. Importantly, the ability of purified Apc to rescue this phenotype was reduced when it was constitutively bound to importin-beta. Thus, importin-beta binds to Apc and negatively regulates the MT-assembly and spindle-promoting activity of Apc in a Ran-regulatable manner.
肿瘤抑制因子腺瘤性结肠息肉病（APC）的突变引发了大多数散发性结直肠癌。APC 被认为在调节有丝分裂和间期的微管（MT）动力学中起作用。然而，其作用机制或调控仍知之甚少。我们发现进口素-β是 APC 的结合伴侣，可调节其对 MT 的作用。APC 在体外和非洲爪蟾卵提取物中与进口素-β结合，RanGTP 抑制这种相互作用。进口素-β的角蛋白样重复结构域与 APC 的中部结合，在此处它可以与β-连环蛋白竞争。此外，APC 羧基末端的两个独立位点结合进口素-β的 N 端区域。与进口素-β结合降低了 APC 在体外组装和捆绑 MT 的能力，并抑制了非洲爪蟾卵提取物中微管星状体的组装，但不影响 APC 与 MT 或 EB1 的结合。APC 的耗竭减少了非洲爪蟾卵提取物中冷稳定纺锤体的形成。重要的是，当 APC 与进口素-β持续结合时，其纯化产物恢复这种表型的能力降低。因此，进口素-β以 Ran 调节的方式与 APC 结合，并负调控 APC 组装 MT 和促进纺锤体形成的活性。
