In Vitro and in Vivo Characterization of Molecular Interactions between Calmodulin, Ezrin/Radixin/Moesin, and L-selectin.
In Vitro and in Vivo Characterization of Molecular Interactions between Calmodulin, Ezrin/Radixin/Moesin, and L-selectin.
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DOI：
10.1074/jbc.m806983200
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发表时间：
2009-03-27
期刊：
影响因子：
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通讯作者：
Ivetic A
中科院分区：
文献类型：
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作者：
Killock DJ;Parsons M;Zarrouk M;Ameer-Beg SM;Ridley AJ;Haskard DO;Zvelebil M;Ivetic A
L-选择素是一种细胞粘附分子，在炎症期间将白细胞束缚在小静脉的腔壁上，并使它们能够在血流的力量下滚动。滚动过程中L-选择素的聚集被认为促进由外向内的信号，导致整合素活化和趋化因子受体表达，最终导致白细胞停滞。一些研究强调了L-选择素胞质尾区在功能性调节粘附和信号传导中的重要性。有趣的是，L-选择素尾部仅由17个氨基酸组成，但它被认为同时与几种蛋白质结合。例如，钙调蛋白（CaM）和埃兹蛋白/根蛋白/膜突蛋白（ERM）与L-选择素的组成性结合分别赋予对蛋白水解和微绒毛定位的抗性。在本研究中，我们发现重组纯化的CaM和ERM与L-选择素的同一尾部非竞争性结合。此外，分子模拟支持的可能性，钙调素，L-选择素，和膜突蛋白可以形成一个异源三聚体复合物。最后，使用荧光寿命成像显微镜测量荧光共振能量转移，它表明，钙调素，L-选择素，和ERM可以同时在体内相互作用。此外，L-选择素成簇促进CaM/ERM顺式相互作用（即来自相邻的L-选择素尾部）。这些结果突出了一种新的细胞内事件，发生的结果，L-选择素聚类，这可能参与转导信号，促进从滚动到逮捕的过渡。
L-selectin is a cell adhesion molecule that tethers leukocytes to the luminal walls of venules during inflammation and enables them to roll under the force of blood flow. Clustering of L-selectin during rolling is thought to promote outside-in signals that lead to integrin activation and chemokine receptor expression, ultimately contributing to leukocyte arrest. Several studies have underscored the importance of the L-selectin cytoplasmic tail in functionally regulating adhesion and signaling. Interestingly, the L-selectin tail comprises only 17 amino acids, and yet it is thought to bind simultaneously to several proteins. For example, constitutive association of calmodulin (CaM) and ezrin/radixin/moesin (ERM) to L-selectin confers resistance to proteolysis and microvillar positioning, respectively. In this report we found that recombinant purified CaM and ERM bound non-competitively to the same tail of L-selectin. Furthermore, molecular modeling supported the possibility that CaM, L-selectin, and moesin could form a heterotrimeric complex. Finally, using fluorescence lifetime imaging microscopy to measure fluorescence resonance energy transfer, it was shown that CaM, L-selectin, and ERM could interact simultaneously in vivo. Moreover, L-selectin clustering promoted CaM/ERM interaction in cis (i.e. derived from neighboring L-selectin tails). These results highlight a novel intracellular event that occurs as a consequence of L-selectin clustering, which could participate in transducing signals that promote the transition from rolling to arrest.