Regulation of Integrin Affinity States through an NP XY Motif in the 
Subunit Cytoplasmic Domain (*)
- From the (1) Department of Vascular Biology, Scripps Research Institute, La Jolla, California 92037
- Department of Biochemistry, University of Helsinki, 00014 Helsinki, Finland
- § Established Investigator of the American Heart Association. To whom correspondence should be addressed. Tel.: 619-554-7151; Fax: 619-554-6403; E-mail: otoole{at}scripps.edu.
Abstract
The ligand binding affinities of the integrins are regulated through their cytoplasmic domains. To identify specific residues
that are involved in this process, we have generated mutants in the β1and β3tails and coexpressed them in Chinese hamster ovary cells with constitutively active α subunits. These α subunits are chimera
of extracellular and transmembrane α
joined to the cytoplasmic domains of α5, α
, or α
and confer an energy-dependent high affinity state when expressed in Chinese hamster ovary cells. The affinity state of these
transfectants was determined by analyzing the binding of PAC1, an antibody that specifically recognizes the activated form
of the reporter group, extracellualar α
β3. We have identified point mutants in several areas of the β tails, which result in a reduced ability to bind ligand. Complete
abolition of PAC1 binding was obtained with mutants in an NP XY motif found in many integrin β subunits and implicated in the internalization of other cell surface receptors. Similar effects
on PAC1 binding were observed whether coexpression was with α chimera containing α5, α
, or α
cytoplasmic sequences. These studies identify a novel role for the NP XY motif in the regulation of integrin binding affinity.
Footnotes
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↵* This work was supported in part by University of California Tobacco Related Disease Research Program Grants 3RT-0320 and HL-48728. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
-
↵1 The abbreviations used are:
- CHO
-
Chinese hamster ovary
- DMEM
-
Dulbecco's modified Eagle's medium
- FITC
-
fluorescein isothiocyanate
- LDL
-
low density lipoprotein.
-
↵2J. Ylanne, J. Huuskonen, T. E. O'Toole, M. H. Ginsberg, I. Virtanen, and C. G. Gahmberg, manuscript in preparation.
- © 1995 by The American Society for Biochemistry and Molecular Biology, Inc.
