Moh Mei Chung, Zhang Chunli, Luo Chunli, Lee Lay Hoon, Shen Shali
Department of Physiology, Faculty of Medicine, National University of Singapore, 2 Medical Drive, Singapore 117597, Republic of Singapore.
J Biol Chem. 2005 Jul 22;280(29):27366-74. doi: 10.1074/jbc.M500852200. Epub 2005 May 25.
We have recently identified a novel gene, hepaCAM, in liver that encodes a cell adhesion molecule of the immunoglobulin superfamily. In this study, we examined the characteristics of hepaCAM protein and the relationship between its structure and function, in particular its adhesive properties. The wild-type and the cytoplasmic domain-truncated mutants of hepaCAM were transfected into the human breast carcinoma MCF7 cells, and the physiological and biological properties were assessed. Biochemical analyses revealed that hepaCAM is an N-linked glycoprotein phosphorylated in the cytoplasmic domain and that it forms homodimers through cis-interaction on the cell surface. The subcellular localization of hepaCAM appears density-dependent; in well spread cells, hepaCAM is distributed to cell protrusions, whereas in confluent cells, hepaCAM is predominantly accumulated at the sites of cell-cell contacts on the cell membrane. In polarized cells, hepaCAM is recruited to the lateral and basal membranes, and lacking physical interaction, hepaCAM is shown to co-localize with E-cadherin at the lateral membrane. Cell adhesion and motility assays demonstrated that hepaCAM increased cell spreading on the matrices fibronectin and matrigel, delayed cell detachment, and enhanced wound healing. Furthermore, when the cytoplasmic domain was deleted, hepaCAM mutants did not affect cell surface localization and dimer formation. Cell-matrix adhesion, however, was less significantly increased, and cell motility was almost unchanged when compared with the effect of the wild-type hepaCAM. Taken together, the cytoplasmic domain of hepaCAM is essential to its function on cell-matrix interaction and cell motility.
我们最近在肝脏中鉴定出一个新基因hepaCAM，它编码免疫球蛋白超家族的一种细胞粘附分子。在本研究中，我们检测了hepaCAM蛋白的特性及其结构与功能之间的关系，特别是其粘附特性。将hepaCAM的野生型和胞质结构域截短突变体转染到人乳腺癌MCF7细胞中，并评估其生理和生物学特性。生化分析表明，hepaCAM是一种在胞质结构域磷酸化的N-连接糖蛋白，它通过细胞表面的顺式相互作用形成同二聚体。hepaCAM的亚细胞定位似乎依赖于密度；在铺展良好的细胞中，hepaCAM分布于细胞突起处，而在汇合细胞中，hepaCAM主要聚集在细胞膜上细胞-细胞接触的部位。在极化细胞中，hepaCAM被募集到侧膜和基底膜，并且在缺乏物理相互作用的情况下，hepaCAM显示与E-钙粘蛋白在侧膜处共定位。细胞粘附和运动分析表明，hepaCAM增加了细胞在纤连蛋白和基质胶基质上的铺展，延迟了细胞脱离，并促进了伤口愈合。此外，当胞质结构域缺失时，hepaCAM突变体不影响细胞表面定位和二聚体形成。然而，与野生型hepaCAM的作用相比，细胞-基质粘附增加的幅度较小，细胞运动几乎没有变化。综上所述，hepaCAM的胞质结构域对其在细胞-基质相互作用和细胞运动中的功能至关重要。
