Binding of urokinase-type plasminogen activator receptor (uPAR) to the mannose 6-phosphate/insulin-like growth factor II receptor: contrasting interactions of full-length and soluble forms of uPAR.
Urokinase-type plasminogen activator receptor (uPAR) binding by the mannose 6-phosphate/insulin-like growth factor II receptor (Man-6-P/IGF2R) is considered important to Man-6-P/IGF2R tumor suppressor function via regulation of cell surface proteolytic activity. Our goal was to map the uPAR binding site of the Man-6-P/IGF2R by analyzing the uPAR binding characteristics of a panel of minireceptors containing different regions of the Man-6-P/IGF2R extracytoplasmic domain. Coimmunoprecipitation assays revealed that soluble recombinant uPAR (suPAR) bound the Man-6-P/IGF2R at two distinct sites, one localized to the amino-terminal end of the Man-6-P/IGF2R extracytoplasmic domain (repeats 1-3) and the other to the more carboxyl-terminal end (repeats 7-9). These sites correspond with the positions of the two Man-6-P binding domains of Man-6-P/IGF2R. Indeed, the suPAR-Man-6-P/IGF2R interaction was inhibited by Man-6-P, and binding-competent su-PAR species represented a minor percentage (8-30%) of the suPAR present. In contrast, Man-6-P/IGF2R binding of endogenous, full-length uPAR solubilized from plasma membranes of the prostate cancer cell line, PC-3, was not inhibited by Man-6-P. Further studies showed that very little (<5%) endogenous uPAR was Man-6-P/IGF2R binding-competent. We conclude that, contrary to previous reports, the interaction between uPAR and Man-6-P/IGF2R is a low percentage binding event and that suPAR and full-length uPAR bind the Man-6-P/IGF2R by different mechanisms.
Kreiling
Jodi L
JL
Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, Nebraska 68198-4525, USA.
Byrd
James C
JC
Deisz
Robert J
RJ
Mizukami
Ikuko F
IF
Todd
Robert F
RF
MacDonald
Richard G
RG
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D007668:Kidney / Q000166:cytology; D008297:Male; D008360:Mannosephosphates / Q000378:metabolism; D016296:Mutagenesis / Q000502:physiology; D011471:Prostatic Neoplasms; D017434:Protein Structure, Tertiary; D017527:Receptor, IGF Type 2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism*; D055293:Receptors, Urokinase Plasminogen Activator; D012995:Solubility; D014407:Tumor Cells, Cultured
D008360:Mannosephosphates; C524898:PLAUR protein, human; D017527:Receptor, IGF Type 2; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; C027693:mannose-6-phosphate