Reinacher-Schick A, Gumbiner B M
Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
J Cell Biol. 2001 Feb 5;152(3):491-502. doi: 10.1083/jcb.152.3.491.
The adenomatous polyposis coli (APC) protein is implicated in the majority of hereditary and sporadic colon cancers. APC is known to function as a tumor suppressor through downregulation of beta-catenin as part of a high molecular weight complex known as the beta-catenin destruction complex. The molecular composition of the intact complex and its site of action in the cell are still not well understood. Reports on the subcellular localization of APC in various cell systems have differed significantly and have been consistent with an association with a cytosolic complex, with microtubules, with the nucleus, or with the cortical actin cytoskeleton. To better understand the role of APC and the destruction complex in colorectal cancer, we have begun to characterize and isolate these complexes from confluent polarized human colon epithelial cell monolayers and other epithelial cell types. Subcellular fractionation and immunofluorescence microscopy reveal that a predominant fraction of APC associates tightly with the apical plasma membrane in a variety of epithelial cell types. This apical membrane association is not dependent on the mutational status of either APC or beta-catenin. An additional pool of APC is cytosolic and fractionates into two distinct high molecular weight complexes, 20S and 60S in size. Only the 20S fraction contains an appreciable portion of the cellular axin and small but detectable amounts of glycogen synthase kinase 3beta and beta-catenin. Therefore, it is likely to correspond to the previously characterized beta-catenin destruction complex. Dishevelled is almost entirely cytosolic, but does not significantly cofractionate with the 20S complex. The disproportionate amount of APC in the apical membrane and the lack of other destruction complex components in the 60S fraction of APC raise questions about whether these pools of APC take part in the degradation of beta-catenin, or alternatively, whether they could be involved in other functions of the protein that still must be determined.
腺瘤性结肠息肉病蛋白（APC）与大多数遗传性和散发性结肠癌有关。已知APC作为一种肿瘤抑制因子发挥作用，通过下调β-连环蛋白，作为一种称为β-连环蛋白破坏复合体的高分子量复合物的一部分。完整复合体的分子组成及其在细胞中的作用位点仍未完全清楚。关于APC在各种细胞系统中的亚细胞定位的报道差异很大，并且与与胞质复合体、微管、细胞核或皮质肌动蛋白细胞骨架的关联一致。为了更好地理解APC和破坏复合体在结直肠癌中的作用，我们开始从汇合极化的人结肠上皮细胞单层和其他上皮细胞类型中表征和分离这些复合体。亚细胞分级分离和免疫荧光显微镜显示，在各种上皮细胞类型中，APC的主要部分与顶端质膜紧密结合。这种顶端膜关联不依赖于APC或β-连环蛋白的突变状态。APC的另一部分位于胞质中，并分为两个不同的高分子量复合体，大小分别为20S和60S。只有20S部分含有细胞中可观比例的轴抑制蛋白以及少量但可检测到的糖原合酶激酶3β和β-连环蛋白。因此，它可能对应于先前表征的β-连环蛋白破坏复合体。无序蛋白几乎完全位于胞质中，但与20S复合体没有明显的共分级分离。APC在顶端膜中的不成比例数量以及APC的60S部分中缺乏其他破坏复合体成分，引发了关于这些APC池是否参与β-连环蛋白降解的问题，或者它们是否可能参与该蛋白仍有待确定的其他功能。
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