EVALUATION OF INTEGRIN MOLECULES INVOLVED IN SUBSTRATE ADHESION

被引:27
作者
ENOMOTOIWAMOTO, M [1 ]
MENKO, AS [1 ]
PHILP, N [1 ]
BOETTIGER, D [1 ]
机构
[1] UNIV PENN,DEPT MICROBIOL,PHILADELPHIA,PA 19104
关键词
INTEGRIN; CROSS-LINKING; ADHESION; CYTOSKELETON;
D O I
10.3109/15419069309097253
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Integrins were cross-linked to their extracellular matrix ligands using non-penetrating chemical cross-linkers. This procedure did not disturb the distribution of integrin in the adhesion structure and adhesion plaque integrin staining remained even when the cultures were extracted with ionic detergents. 80-90% of the beta1 integrin in the cross-linked culture was extracted with RIPA buffer and the remaining 10-20% was recovered following reversal of the cross-linking. This separated two distinct integrin pools, one which can be cross-linked to substrate bound extracellular matrix and one which is not. The specificity of this procedure for cross-linking of integrins involved in substrate adhesion was demonstrated using NIH 3T3 cells which express both alpha5beta1 and alpha6beta1 integrins. Alpha6 was cross-linked only in cells plated on laminin whereas alpha5 was cross-linked when fibronectin was present. Using antisera directed to the cytoplasmic domains of either alpha5 or beta1 integrin, it was demonstrated that these domains can be blocked in the intact cell but the blocking can be removed using ionic detergent extraction after chemical cross-linking. The extracellular matrix associated with the substrate surface but not that associated with the media exposed surface is both cross-linked and retained on the plastic dish following cross-linking.
引用
收藏
页码:191 / 202
页数:12
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