Potential role for a novel AP180-related protein during endocytosis in MDCK cells.
Potential role for a novel AP180-related protein during endocytosis in MDCK cells.
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MDCK 细胞内吞过程中新型 AP180 相关蛋白的潜在作用。
DOI：
10.1152/ajpcell.00079.2003
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发表时间：
2003
期刊：
影响因子：
--
通讯作者：
Carlin,Cathleen
中科院分区：
文献类型：
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作者：
Kusner,Linda;Carlin,Cathleen
网格蛋白组装蛋白AP 180最初被鉴定为定位于突触前连接的脑特异性蛋白。AP 180的作用是限制囊泡的大小，并在快速循环过程中维持可释放的突触囊泡池。在这项研究中，我们发现极化的上皮Madin-Darby犬肾（MDCK）细胞表达两种AP 180相关蛋白：普遍表达的62 kDa网格蛋白组装淋巴样髓性白血病（CALM，AP 180 -2）蛋白和一种新的高分子量同系物，我们命名为AP 180 -3。对MDCK细胞中表达的AP 180 -3进行序列分析，结果显示其与大鼠脑组织中的AP 180具有高度同源性。AP 180 -3含有脑特异性AP 180中发现的保守基序，包括epsin NH 2-末端同源（ENTH）结构域、AP-2 α亚基的结合位点和DLL重复序列。我们的研究表明，来自MDCK细胞的AP 180 -3与AP-2和网格蛋白形成复合物，并且这些复合物的膜募集受磷酸化调节。我们通过免疫组织化学证实，AP 180 -3定位于MDCK细胞的胞质囊泡，并且也存在于小鼠肾脏的小管上皮细胞中。我们通过免疫检测观察到，除了MDCK细胞外，高分子量的AP 180相关蛋白在许多细胞中表达。
Clathrin assembly protein, AP180, was originally identified as a brain-specific protein localized to the presynaptic junction. AP180 acts to limit vesicle size and maintain a pool of releasable synaptic vesicles during rapid recycling. In this study, we show that polarized epithelial Madin-Darby canine kidney (MDCK) cells express two AP180-related proteins: the ubiquitously expressed 62-kDa clathrin assembly lymphoid myeloid leukemia (CALM, AP180-2) protein and a novel high-molecular-weight homolog that we have named AP180-3. Sequence analysis of AP180-3 expressed in MDCK cells shows high homology to AP180 from rat brain. AP180-3 contains conserved motifs found in brain-specific AP180, including the epsin NH2-terminal homology (ENTH) domain, the binding site for the α-subunit of AP-2, and DLL repeats. Our studies show that AP180-3 from MDCK cells forms complexes with AP-2 and clathrin and that membrane recruitment of these complexes is modulated by phosphorylation. We demonstrate by immunohistochemistry that AP180-3 is localized to cytoplasmic vesicles in MDCK cells and is also present in tubule epithelial cells from mouse kidney. We observed by immunodetection that a high-molecular-weight AP180-related protein is expressed in numerous cells in addition to MDCK cells.