Fibrinogen interaction of CHO cells expressing chimeric αⅡb/αvβ3 integrin  

Fibrinogen interaction of CHO cells expressing chimeric αⅡb/αvβ3 integrin

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作  者:Juan-juan CHEN Xiao-yu SU Xiao-dong XI Li-ping LIN Jian DING He LU 

机构地区:[1]Division of Antitumor Pharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China [2]Shanghai Institute of Hematology, Ruijin Hospital, Shanghai 200025, China [3]INSERM UMR 553, Saint Louis Hospital, Paris 75010, France

出  处:《Acta Pharmacologica Sinica》2008年第2期204-210,共7页中国药理学报(英文版)

摘  要:Aim: The molecular mechanisms of the affinity regulation of αvβ3 integrin are important in tumor development, wound repairing, and angiogenesis. It has been established that the cytoplasmic domains of αvβ3 integrin play an important role in integrin-ligand affinity regulation. However, the relationship of structure-function within these domains remains unclear. Methods: The extracellular and transmembrane domain of αⅡb was fused to the αv integrin cytoplasmic domain, and the chimeric α subunit was coexpressed in Chinese hamster ovary (CHO) cells with the wild-type β3 subunit or with 3 mutant β3 sequences bearing truncations at the positions of T741, Y747, and F754, respectively. The CHO cells expressing these recombinant integrins were tested for soluble fibrinogen binding and the cell adhesion and spreading on immobilized fibrinogen. Results: All 4 types of integrins bound soluble fibrinogen in the absence of agonist stimulation, and only the cells expressing the chimeric α subunit with the wild-type β3 subunit, but not those with truncated β3, could adhere to and spread on immobilized fibrinogen. Conclusion: The substitution αⅡb at the cytoplasmic domain with the αv cytoplasmic sequence rendered the extracellular αⅡbβ3 a constitutively activated conformation for ligands without the need of "inside-out" signals. Our results also indicated that the COOH-terminal sequence of β3 might play a key role in integrin αⅡb/αvβ3-mediated cell adhesion and spreading on immobilized fibrinogen. The cells expressing αⅡb/αvβ3 have enormous potential for facilitating drug screen- ing for antagonists either to αvβ3 intracellular interactions or to αⅡbβ3 receptor functions.Aim: The molecular mechanisms of the affinity regulation of αvβ3 integrin are important in tumor development, wound repairing, and angiogenesis. It has been established that the cytoplasmic domains of αvβ3 integrin play an important role in integrin-ligand affinity regulation. However, the relationship of structure-function within these domains remains unclear. Methods: The extracellular and transmembrane domain of αⅡb was fused to the αv integrin cytoplasmic domain, and the chimeric α subunit was coexpressed in Chinese hamster ovary (CHO) cells with the wild-type β3 subunit or with 3 mutant β3 sequences bearing truncations at the positions of T741, Y747, and F754, respectively. The CHO cells expressing these recombinant integrins were tested for soluble fibrinogen binding and the cell adhesion and spreading on immobilized fibrinogen. Results: All 4 types of integrins bound soluble fibrinogen in the absence of agonist stimulation, and only the cells expressing the chimeric α subunit with the wild-type β3 subunit, but not those with truncated β3, could adhere to and spread on immobilized fibrinogen. Conclusion: The substitution αⅡb at the cytoplasmic domain with the αv cytoplasmic sequence rendered the extracellular αⅡbβ3 a constitutively activated conformation for ligands without the need of "inside-out" signals. Our results also indicated that the COOH-terminal sequence of β3 might play a key role in integrin αⅡb/αvβ3-mediated cell adhesion and spreading on immobilized fibrinogen. The cells expressing αⅡb/αvβ3 have enormous potential for facilitating drug screen- ing for antagonists either to αvβ3 intracellular interactions or to αⅡbβ3 receptor functions.

关 键 词:INTEGRIN cell adhesion FIBRINOGEN SIGNALTRANSDUCTION 

分 类 号:R318[医药卫生—生物医学工程]

 

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