Distinct involvement of β3 integrin cytoplasmic domain tyrosine residues 747 and 759 in integrin-mediated cytoskeletal assembly and phosphotyrosine signaling
Distinct involvement of β3 integrin cytoplasmic domain tyrosine residues 747 and 759 in integrin-mediated cytoskeletal assembly and phosphotyrosine signaling
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DOI：
10.1074/jbc.273.20.12623
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发表时间：
1998-05-15
影响因子：
4.8
通讯作者：
Kieffer, N
中科院分区：
文献类型：
--
作者：
Schaffner-Reckinger, E;Gouon, V;Kieffer, N
我们研究了在α(V)β(3)介导的细胞扩散过程中，粘着斑激酶(FAK)和帕西林酪氨酸磷酸化所必需的整合素亚单位胞浆区域的结构要求。使用不同的β(3)突变体转染的CHO细胞，我们证明了α(V)β(3)介导的细胞扩展与FAK和PXLIN的酪氨酸磷酸化密切相关，并强调了NPLY747和NITY759基序在这些信号过程中的明显参与。仅NITY759基序的缺失就足以完全阻止依赖于α(V)β(3)的焦点接触形成、细胞扩散和FAK/帕西林磷酸化，单一的Y759A突变诱导了强烈的抑制表型，而更保守但仍有磷酸化缺陷的Y759F突变恢复了野生型受体的功能。高度保守的Tyr(747)的丙氨酸替换完全取消GP、局灶性粘连斑块的依赖形成、细胞扩散和FAK/paxlin磷酸化，而Y747F替换仅部分恢复了这些事件。由于这些突变都不影响受体与配体的相互作用，我们的结果表明，NITU759基序的结构完整性，而不是Tyr(759)的磷酸化状态，对于β(3)介导的细胞骨架重组和FAK和Paxlin的酪氨酸磷酸化是重要的，而NPLY747基序中747残基上的Tyr的存在是最佳的β(3)配体结合事件所必需的。
We have investigated the structural requirements of the beta(3) integrin subunit cytoplasmic domain necessary for tyrosine phosphorylation of focal adhesion kinase (FAK) and paxillin during alpha(v) beta(3)-mediated cell spreading. Using CHO cells transfected with various beta(3) mutants, we demonstrate a close correlation between alpha(v) beta(3)-mediated cell spreading and tyrosine phosphorylation of FAK and paxillin, and highlight a distinct involvement of the NPLY747 and NITY759 motifs in these signaling processes. Deletion of the NITY759 motif alone was sufficient to completely prevent alpha(v) beta(3)-dependent focal contact formation, cell spreading, and FAK/paxillin phosphorylation, The single Y759A substitution induced a strong inhibitory phenotype, while the more conservative, but still phosphorylation-defective, Y759F mutation restored wild type receptor function. Alanine substitution of the highly conserved Tyr(747) completely abolished gp,dependent formation of focal adhesion plaques, cell spreading, and FAK/paxillin phosphorylation, whereas a Y747F substitution only partially restored these events. As none of these mutations affected receptor-ligand interaction, our results suggest that the structural integrity of the NITU759 motif, rather than the phosphorylation status of Tyr(759) is important for beta(3)-mediated cytoskeleton reorganization and tyrosine phosphorylation of FAK and paxillin, while the presence of Tyr at residue 747 within the NPLY747 motif is required for optimal beta(3) post-ligand binding events.