Identification of a functionally important sequence in the cytoplasmic tail of integrin beta 3 by using cell-permeable peptide analogs.
Integrins are major two-way signaling receptors responsible for the attachment of cells to the extracellular matrix and for cell-cell interactions that underlie immune responses, tumor metastasis, and progression of atherosclerosis and thrombosis. We report the structure-function analysis of the cytoplasmic tail of integrin beta 3 (glycoprotein IIla) based on the cellular import of synthetic peptide analogs of this region. Among the four overlapping cell-permeable peptides, only the peptide carrying residues 747-762 of the carboxyl-terminal segment of integrin beta 3 inhibited adhesion of human erythroleukemia (HEL) cells and of human endothelial cells (ECV) 304 to immobilized fibrinogen mediated by integrin beta 3 heterodimers, alpha IIb beta 3, and alpha v beta 3, respectively. Inhibition of adhesion was integrin-specific because the cell-permeable beta 3 peptide (residues 747-762) did not inhibit adhesion of human fibroblasts mediated by integrin beta 1 heterodimers. Conversely, a cell-permeable peptide representing homologous portion of the integrin beta 1 cytoplasmic tail (residues 788-803) inhibited adhesion of human fibroblasts, whereas it was without effect on adhesion of HEL or ECV 304 cells. The cell-permeable integrin beta 3 peptide (residues 747-762) carrying a known loss-of-function mutation (Ser752Pro) responsible for the genetic disorder Glanzmann thrombasthenia Paris I did not inhibit cell adhesion of HEL or ECV 304 cells, whereas the beta 3 peptide carrying a Ser752Ala mutation was inhibitory. Although Ser752 is not essential, Tyr747 and Tyr759 form a functionally active tandem because conservative mutations Tyr747Phe or Tyr759Phe resulted in a nonfunctional cell permeable integrin beta 3 peptide. We propose that the carboxyl-terminal segment of the integrin beta 3 cytoplasmic tail spanning residues 747-762 constitutes a major intracellular cell adhesion regulatory domain (CARD) that modulates the interaction of integrin beta 3-expressing cells with immobilized fibrinogen. Import of cell-permeable peptides carrying this domain results in inhibition "from within" of the adhesive function of these integrins.
Liu
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Department of Microbiology and Immunology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
Timmons
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S
Lin
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YZ
Hawiger
J
J
D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000737:chemistry* / Q000502:physiology*; D001692:Biological Transport; D002448:Cell Adhesion / Q000187:drug effects; D002460:Cell Line; D004730:Endothelium, Vascular / Q000502:physiology*; D006801:Humans; D019012:Integrin beta1 / Q000737:chemistry / Q000502:physiology; D039661:Integrin beta3; D004915:Leukemia, Erythroblastic, Acute; D008297:Male; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000737:chemistry* / Q000502:physiology*; D012867:Skin; D014407:Tumor Cells, Cultured
D015703:Antigens, CD; D019012:Integrin beta1; D039661:Integrin beta3; D010446:Peptide Fragments; D010455:Peptides; D010980:Platelet Membrane Glycoproteins