title: Biomolecular characterization of CD44-fibrin(ogen) binding: distinct molecular requirements mediate binding of standard and variant isoforms of CD44 to immobilized fibrin(ogen).
abstract: CD44 and fibrin(ogen) play critical roles in the hematogenous dissemination of tumor cells, including colon carcinomas. We recently reported that CD44 is the primary fibrin, but not fibrinogen, receptor on LS174T colon carcinomas. However, the biochemical nature of this interaction and the roles of CD44 standard (CD44s) versus CD44 variant (CD44v) isoforms in fibrin(ogen) recognition have yet to be delineated. Microspheres, coated with CD44 immunopurified from LS174T or T84 colon carcinoma cells, which express primarily CD44v, effectively bind to immobilized fibrin, but not fibrinogen, in shear flow. In contrast, CD44s from HL-60 cells binds to both immobilized fibrin and fibrinogen under flow. Use of highly specific enzymes and metabolic inhibitors reveals that LS174T CD44 binding to fibrin is dependent on O-glycosylation of CD44, whereas CD44s-fibrin(ogen) interaction has an absolute requirement for N-, but not O-, linked glycans. The presence of chondroitin and dermatan sulfate on CD44 standard and variant isoforms facilitates fibrin recognition. Use of the anti-CD44 function-blocking monoclonal antibody Hermes-1 nearly abolishes binding of LS174T CD44 to fibrin, although it has no effect on CD44s-fibrin(ogen) interaction. The CD44-binding site is localized within the N-terminal portion of the fibrin beta chains, including amino acid residues (beta15-66). Surface plasmon resonance experiments revealed high affinity binding of immobilized CD44 with solubilized fibrin but not fibrinogen. Collectively, these data suggest that immobilization of fibrinogen exposes a cryptic site that mediates binding to CD44s but not CD44v. Our findings may provide a rational basis for designing novel therapeutic strategies to combat metastasis.
authors.affiliation: Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, Baltimore, Maryland 21218, USA.
authors.forename: Christina S
authors.initials: CS
authors.lastname: Alves
authors.forename: Sergiy
authors.initials: S
authors.lastname: Yakovlev
authors.forename: Leonid
authors.initials: L
authors.lastname: Medved
authors.forename: Konstantinos
authors.initials: K
authors.lastname: Konstantopoulos
mesh_terms: D000268:Adhesiveness; D001665:Binding Sites; D002460:Cell Line; D005337:Fibrin / Q000378:metabolism*; D005340:Fibrinogen / Q000378:metabolism*; D006025:Glycosaminoglycans / Q000378:metabolism; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms / Q000378:metabolism; D018000:Receptors, Peptide / Q000378:metabolism; D013455:Sulfur / Q000378:metabolism; D020349:Surface Plasmon Resonance
chemical_list: D006025:Glycosaminoglycans; D018960:Hyaluronan Receptors; D011134:Polysaccharides; D020033:Protein Isoforms; D018000:Receptors, Peptide; C035637:fibrin receptor; D013455:Sulfur; D005337:Fibrin; D005340:Fibrinogen