Fan G H, Yang W, Wang X J, Qian Q, Richmond A
Veterans Affairs Medical Center, Nashville, Tennessee 37212-2637, USA.
Biochemistry. 2001 Jan 23;40(3):791-800. doi: 10.1021/bi001661b.
Agonist treatment of cells expressing the chemokine receptor, CXCR2, induces receptor phosphorylation and internalization through a dynamin-dependent mechanism. In the present study, we demonstrate that a carboxyl terminus-truncated mutant of CXCR2 (331T), which no longer undergoes agonist-induced phosphorylation, continues to undergo ligand-induced internalization in HEK293 cells. This mutant receptor exhibits reduced association with beta-arrestin 1 but continues to exhibit association with adaptin 2 alpha and beta subunits. Replacing Leu320-321 and/or Ile323-Leu324 with Ala (LL320,321AA, IL323,324AA, and LLIL320,321,323,324AAAA) in wild-type CXCR2 or 331T causes little change in ligand binding and signaling through Ca(2+) mobilization but greatly impairs the agonist-induced receptor sequestration and ligand-mediated chemotaxis. The LL320,321AA, IL323,324AA, and LLIL320,321,323,324AAAA mutants of CXCR2 exhibit normal binding to beta-arrestin 1 but exhibit decreased binding to adaptin 2alpha and beta. These data demonstrate a role for the LLKIL motif in the carboxyl terminus of CXCR2 in receptor internalization and cell chemotaxis and imply a role for adaptin 2 in the endocytosis of CXCR2.
对表达趋化因子受体CXCR2的细胞进行激动剂处理，可通过一种依赖发动蛋白的机制诱导受体磷酸化和内化。在本研究中，我们证明CXCR2的羧基末端截短突变体（331T）不再经历激动剂诱导的磷酸化，但在HEK293细胞中仍继续经历配体诱导的内化。这种突变受体与β-抑制蛋白1的结合减少，但与衔接蛋白2的α和β亚基仍有结合。将野生型CXCR2或331T中的Leu320 - 321和/或Ile323 - Leu324替换为丙氨酸（LL320,321AA、IL323,324AA和LLIL320,321,323,324AAAA），对通过Ca(2+)动员的配体结合和信号传导影响不大，但极大地损害了激动剂诱导的受体隔离和配体介导的趋化作用。CXCR2的LL320,321AA、IL323,324AA和LLIL320,321,323,324AAAA突变体与β-抑制蛋白1的结合正常，但与衔接蛋白2α和β的结合减少。这些数据证明了CXCR2羧基末端的LLKIL基序在受体内化和细胞趋化作用中的作用，并暗示衔接蛋白2在CXCR2的内吞作用中发挥作用。
