A Conserved Sequence Motif in the Integrin β3Cytoplasmic Domain Is Required for Its Specific Interaction with β3-Endonexin

Martin Eigenthaler, Liane Höfferer, Sanford J. Shattil, Mark H. Ginsberg
1997 Journal of Biological Chemistry  
Integrin signaling is mediated by interaction of integrin cytoplasmic domains with intracellular signaling molecules. Recently, we identified a novel 111-amino acid polypeptide, termed ␤ 3 -endonexin, which interacts selectively with the integrin ␤ 3 cytoplasmic domain. In the present study we conducted a systematic mutational analysis of both the integrin ␤ 3 cytoplasmic domain and ␤ 3 -endonexin to map sites required for interaction. The interaction of the full-length ␤ 3 integrin subunit
more » ... ␤ 3 -endonexin in vitro required the ␤ 3 cytoplasmic domain. In a yeast two-hybrid system, both membraneproximal and membrane-distal residues of the ␤ 3 cytoplasmic domain were necessary for interaction with ␤ 3endonexin. In particular, the membrane-distal NITY motif at ␤ 3 756 -759 was critical for the interaction. Exchange of ␤ 3 residues 756 -759 (NITY) for the corresponding residues in ␤ 1 (NPKY) endowed the ␤ 1 cytoplasmic domain with the ability to interact with ␤ 3endonexin. Conversely, exchange of the NPKY motif at ␤ 1 772-775 for the NITY motif in ␤ 3 abolished interaction of this chimeric cytoplasmic domain with ␤ 3 -endonexin. Because the NITY motif is present in the ␤ 3 but not the ␤ 1 cytoplasmic domain, these results explain the selective interaction of this cytoplasmic domain with ␤ 3 -endonexin. In addition, deletional analysis suggested that a core 91-residue sequence of ␤ 3 -endonexin is sufficient for specific binding to the ␤ 3 cytoplasmic domain. These studies have identified a cytoplasmic domain sequence motif that specifies an integrin-specific protein-protein interaction. Integrins are heterodimeric adhesion receptors composed of ␣ and ␤ transmembrane subunits (1). The ␤ 3 integrin subfamily includes ␣ IIb ␤ 3 and ␣ v ␤ 3 . ␣ IIb ␤ 3 is largely specific for cells of the megakaryocytic lineage and is required for platelet aggregation (2). ␣ v ␤ 3 is found in a number of cell types, including endothelial cells, vascular smooth muscle cells, and monocytes, ␤ 3 wild type KLLITI HDRKE FAKFEEERARAKWDTAN NPLY KEATSTFT NITY RGT ␤ 3 ⌬759 KLLITI HDRKE FAKFEEERARAKWDTAN NPLY KEATSTFT NIT ␤ 3 ⌬755 KLLITI HDRKE FAKFEEERARAKWDTAN NPLY KEATSTF ␤ 3 717-755/␤ 1 772-778 KLLITI HDRKE FAKFEEERARAKWDTAN NPLY KEATSTFT NPKY EGK ␤ 3 I757P KLLITI HDRKE FAKFEEERARAKWDTAN NPLY KEATSTFT NPTY RGT ␤ 1 732 ␤ 1 778 Խ Խ ␤ 1 wild type KLLMII HDRRE FAKFEKEKMNAKWDTGE NPIY KSAVTTVV NPKY EGK ␤ 1 732-771/␤ 3 756-762 KLLMII HDRRE FAKFEKEKMNAKWDTGE NPIY KSAVTTVV NITY RGK ␤ 1 P7731
doi:10.1074/jbc.272.12.7693 pmid:9065427 fatcat:6kqdu4svjjh57jxgoba6ih6juy