Domain 2 of the urokinase receptor contains an integrin-interacting epitope with intrinsic signaling activity: generation of a new integrin inhibitor.
We investigated the interaction between the urokinase receptor (uPAR) and the integrin alphavbeta3. Vitronectin (VN) induces cell migration by binding to alphavbeta3, but expression of the uPAR boosts its efficacy. Thus, uPAR may regulate VN-induced cell migration by interacting laterally with alphavbeta3. In contrast, cells expressing a uPAR mutant lacking domain 2 do not migrate in response to VN. This effect is overcome by D2A, a synthetic peptide derived from the sequence of domain 2. In addition, D2A has chemotactic activity that requires alphavbeta3 and activates alphavbeta3-dependent signaling pathways such as the Janus kinase/Stat pathway. Moreover, D2A disrupts uPAR-alphavbeta3 and uPAR-alpha5beta1 co-immunoprecipitation, indicating that it can bind both of these integrins. We also identify the chemotactically active epitope harbored by peptide D2A. Mutating two glutamic acids into two alanines generates peptide D2A-Ala, which lacks chemotactic activity but inhibits VN-, FN-, and collagen-dependent cell migration. In fact, the GEEG peptide has potent chemotactic activity, and the GAAG sequence has inhibitory capacities. In summary, we have identified an integrin-interacting sequence located in domain 2 of uPAR, which is also a new chemotactic epitope that can activate alphavbeta3-dependent signaling pathways and stimulate cell migration. This sequence thus plays a pivotal role in the regulation of uPAR-integrin interactions. Moreover, we describe a novel, very potent inhibitor of integrin-dependent cell migration.
Degryse
Bernard
B
Department of Molecular Biology and Functional Genomics, DIBIT, Università Vita Salute San Raffaele, Via Olgettina 58, 20132 Milan, Italy.
Resnati
Massimo
M
Czekay
Ralf-Peter
RP
Loskutoff
David J
DJ
Blasi
Francesco
F
D000256:Adenoviridae / Q000235:genetics; D000409:Alanine / Q000737:chemistry; D000818:Animals; D001011:Aorta / Q000378:metabolism; D002448:Cell Adhesion; D002460:Cell Line; D002465:Cell Movement; D002633:Chemotaxis; D003599:Cytoskeleton / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D000939:Epitopes / Q000737:chemistry; D018698:Glutamic Acid / Q000737:chemistry; D006801:Humans; D047468:Immunoprecipitation; D039302:Integrin alphaVbeta3 / Q000737:chemistry; D016023:Integrins / Q000737:chemistry*; D051379:Mice; D008856:Microscopy, Fluorescence; D009154:Mutation; D032389:Myocytes, Smooth Muscle / Q000166:cytology; D041681:NIH 3T3 Cells; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D051381:Rats; D011956:Receptors, Cell Surface / Q000737:chemistry*; D055293:Receptors, Urokinase Plasminogen Activator; D015398:Signal Transduction; D013696:Temperature; D013997:Time Factors; D014162:Transfection; D020032:Tyrphostins / Q000494:pharmacology
D000939:Epitopes; D039302:Integrin alphaVbeta3; D016023:Integrins; C524898:PLAUR protein, human; D010455:Peptides; C524899:Plaur protein, mouse; C524900:Plaur protein, rat; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D020032:Tyrphostins; C095512:alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide; D018698:Glutamic Acid; D000409:Alanine