Filamin binds to the cytoplasmic domain of the beta1-integrin. Identification of amino acids responsible for this interaction.
Integrins play an important role in regulating cell adhesion, motility, and activation. In an effort to identify intracellular proteins expressed by activated T cells that interact with the cytoplasmic domain of beta1-integrin (CD29), we used the beta1-integrin cytoplasmic domain as bait in the yeast two-hybrid system. Here we report that the cytoplasmic domain of beta1-integrin specifically interacts with the cytoskeletal protein filamin. This interaction required all but the most carboxyl-terminal three residues of the cytoplasmic domain of beta1, and the carboxyl-terminal 477 residues of filamin containing the terminal 4. 5 approximately 96-residue tandem repeats of filamin. To verify this interaction in vivo, we showed that filamin specifically coprecipitated with beta1 in mammalian cells. We also showed that recombinant filamin chimeric proteins were able to bind to the beta1 cytoplasmic domain in vitro. We observed that a subset of single point mutations in the cytoplasmic domain of beta1, which had been previously reported to impair its function, disrupt the interaction between beta1 and filamin. Taken together, these findings suggest that the interaction between beta1 and filamin, which in turn can bind actin, provides a mechanism for the interaction of this cell surface receptor with cytoskeletal proteins and that this interaction plays a role in normal receptor function.
Bristol-Myers Squibb Pharmaceutical Research Institute, Princeton, New Jersey 08543, USA. lood@bms.com
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D000595:Amino Acid Sequence; D001665:Binding Sites; D016764:Cell Polarity; D003285:Contractile Proteins / Q000235:genetics / Q000378:metabolism*; D003593:Cytoplasm; D064448:Filamins; D006801:Humans; D019012:Integrin beta1 / Q000235:genetics / Q000378:metabolism*; D019169:Jurkat Cells; D008213:Lymphocyte Activation; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D011233:Precipitin Tests; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013601:T-Lymphocytes / Q000378:metabolism
D003285:Contractile Proteins; D064448:Filamins; D019012:Integrin beta1; D008840:Microfilament Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins