Kiely Jeanne-Marie, Hu Yenya, García-Cardeña Guillermo, Gimbrone Michael A
Department of Pathology, Center for Excellence in Vascular Biology, Brigham and Women's Hospital, and Harvard Medical School, Boston, MA 02115, USA.
J Immunol. 2003 Sep 15;171(6):3216-24. doi: 10.4049/jimmunol.171.6.3216.
E-selectin, an endothelial cell surface adhesion receptor for leukocytes, also acts as a signaling receptor. Upon multivalent ligation, E-selectin transduces outside-in signals into the endothelium leading to changes in intracellular Ca(2+) concentration and activation of the mitogen-activated protein kinase signaling pathway. In addition, following leukocyte engagement, E-selectin associates via its cytoplasmic domain with components of the actin cytoskeleton and undergoes alterations in phosphorylation state that result in changes in gene expression. In this study, we show that E-selectin is localized in cholesterol-rich lipid rafts at the cell surface, and that upon ligation E-selectin clusters and redistributes in the plasma membrane colocalizing with a fraction of caveolin-1-containing rafts. In addition, we demonstrate that leukocyte adhesion via E-selectin results in association with and activation of phospholipase Cgamma (PLCgamma). Moreover, we show that disruption of lipid rafts with the cholesterol-depleting drug methyl-beta-cyclodextrin disrupts the raft localization of E-selectin as well as the ligation-induced association of E-selectin with PLCgamma, and subsequent tyrosine phosphorylation of PLCgamma. In contrast, cholesterol depletion has no effect on E-selectin-dependent mitogen-activated protein kinase activation. Thus, these findings demonstrate that the presence of E-selectin in lipid rafts is necessary for its association with, and activation of, PLCgamma, and suggest that this subcellular localization of E-selectin is related to its signaling function(s) during leukocyte-endothelial interactions.
E选择素是白细胞的内皮细胞表面黏附受体，也作为信号受体发挥作用。在多价连接后，E选择素将外向信号转导至内皮细胞，导致细胞内Ca(2+)浓度变化以及丝裂原活化蛋白激酶信号通路的激活。此外，在白细胞结合后，E选择素通过其胞质结构域与肌动蛋白细胞骨架成分结合，并经历磷酸化状态的改变，从而导致基因表达的变化。在本研究中，我们表明E选择素定位于细胞表面富含胆固醇的脂筏中，并且在连接后，E选择素聚集并在质膜中重新分布，与一部分含有小窝蛋白-1的脂筏共定位。此外，我们证明通过E选择素的白细胞黏附导致与磷脂酶Cγ（PLCγ）的结合和激活。而且，我们表明用胆固醇消耗药物甲基-β-环糊精破坏脂筏会破坏E选择素的脂筏定位以及E选择素与PLCγ的连接诱导结合，以及随后PLCγ的酪氨酸磷酸化。相反，胆固醇消耗对E选择素依赖性丝裂原活化蛋白激酶激活没有影响。因此，这些发现表明E选择素在脂筏中的存在对于其与PLCγ的结合和激活是必要的，并表明E选择素的这种亚细胞定位与其在白细胞-内皮细胞相互作用期间的信号功能有关。
