A conserved sequence motif in the integrin beta3 cytoplasmic domain is required for its specific interaction with beta3-endonexin.
Integrin signaling is mediated by interaction of integrin cytoplasmic domains with intracellular signaling molecules. Recently, we identified a novel 111-amino acid polypeptide, termed beta3-endonexin, which interacts selectively with the integrin beta3 cytoplasmic domain. In the present study we conducted a systematic mutational analysis of both the integrin beta3 cytoplasmic domain and beta3-endonexin to map sites required for interaction. The interaction of the full-length beta3 integrin subunit with beta3-endonexin in vitro required the beta3 cytoplasmic domain. In a yeast two-hybrid system, both membrane-proximal and membrane-distal residues of the beta3 cytoplasmic domain were necessary for interaction with beta3-endonexin. In particular, the membrane-distal NITY motif at beta3 756-759 was critical for the interaction. Exchange of beta3 residues 756-759 (NITY) for the corresponding residues in beta1 (NPKY) endowed the beta1 cytoplasmic domain with the ability to interact with beta3-endonexin. Conversely, exchange of the NPKY motif at beta1 772-775 for the NITY motif in beta3 abolished interaction of this chimeric cytoplasmic domain with beta3-endonexin. Because the NITY motif is present in the beta3 but not the beta1 cytoplasmic domain, these results explain the selective interaction of this cytoplasmic domain with beta3-endonexin. In addition, deletional analysis suggested that a core 91-residue sequence of beta3-endonexin is sufficient for specific binding to the beta3 cytoplasmic domain. These studies have identified a cytoplasmic domain sequence motif that specifies an integrin-specific protein-protein interaction.
Eigenthaler
M
M
Department of Vascular Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
Höfferer
L
L
Shattil
S J
SJ
Ginsberg
M H
MH
D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000737:chemistry / Q000378:metabolism*; D016466:CHO Cells; D017124:Conserved Sequence; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism*; D006801:Humans; D039661:Integrin beta3; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism*; D016415:Sequence Alignment
D015703:Antigens, CD; C403295:ITGB3BP protein, human; D039661:Integrin beta3; D009687:Nuclear Proteins; D010980:Platelet Membrane Glycoproteins; D011506:Proteins