Barth Angela I M, Siemers Kathleen A, Nelson W James
Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, CA 94305-5435, USA.
J Cell Sci. 2002 Apr 15;115(Pt 8):1583-90. doi: 10.1242/jcs.115.8.1583.
End-binding protein (EB) 1 binds to the C-terminus of adenomatous polyposis coli (APC) protein and to the plus ends of microtubules (MT) and has been implicated in the regulation of APC accumulation in cortical clusters at the tip of extending membranes. We investigated which APC domains are involved in cluster localization and whether binding to EB1 or MTs is essential for APC cluster localization. Armadillo repeats of APC that lack EB1- and MT-binding domains are necessary and sufficient for APC localization in cortical clusters; an APC fragment lacking the armadillo repeats, but containing MT- and EB1-binding domains, does not localize to the cortical clusters but instead co-aligns with MTs throughout the cell. Significantly, analysis of endogenous proteins reveals that EB1 does not accumulate in the APC clusters. However, overexpressed EB1 does accumulate in APC clusters; the APC-binding domain in EB1 is located in the C-terminal region of EB1 between amino acids 134 and 268. Overexpressed APC- or MT-binding domains of EB1 localize to APC cortical clusters and MT, respectively, without affecting APC cluster formation itself. These results show that localization of APC in cortical clusters is different from that of EB1 at MT plus ends and appears to be independent of EB1.
末端结合蛋白（EB）1与腺瘤性息肉病大肠杆菌（APC）蛋白的C末端以及微管（MT）的正端结合，并参与调节延伸膜尖端皮质簇中APC的积累。我们研究了哪些APC结构域参与簇定位，以及与EB1或MT的结合对于APC簇定位是否至关重要。缺乏EB1和MT结合结构域的APC犰狳重复序列对于APC在皮质簇中的定位是必要且充分的；一个缺乏犰狳重复序列但含有MT和EB1结合结构域的APC片段并不定位于皮质簇，而是在整个细胞中与MT共排列。值得注意的是，对内源蛋白的分析表明EB1不会在APC簇中积累。然而，过表达的EB1确实会在APC簇中积累；EB1中的APC结合结构域位于EB1的C末端区域，氨基酸位置在134至268之间。过表达的EB1的APC或MT结合结构域分别定位于APC皮质簇和MT，而不影响APC簇的形成本身。这些结果表明，APC在皮质簇中的定位与EB1在MT正端的定位不同，并且似乎独立于EB1。
