CELLULAR ADHESION MEDIATED BY FACTOR-J, A COMPLEMENT INHIBITOR - EVIDENCE FOR NUCLEOLIN INVOLVEMENT

Factor J (FJ) is a complement inhibitor that acts on the classical and the alternative pathways. We demonstrated FJ-cell interactions in flu id phase by flow cytometry experiments using the cell lines Jurkat, K5 62, JY, and peripheral blood lymphocytes. FJ bound to plastic plates w as able to induce in vitro adhesion of these cells with potency equiva lent to fibronectin. As evidence for the specificity of this reaction, the adhesion was blocked by MAJ2, an anti-FJ monoclonal antibody, and by soluble FJ. Attachment of the cells required active metabolism and cytoskeletal integrity. The glycosaminoglycans heparin, heparan sulfa te, or chondroitin sulfates A, B, and C inhibited to varying degrees t he binding of FJ to cells, as did treatment with chondroitinase ABC. I n the search for a putative receptor, a protein of 110 kDa was isolate d by affinity chromatography, and microsequence analysis identified th is protein as nucleolin. Confocal microscopy evidenced the presence of nucleolin in cell membrane by immunofluorescence with monoclonal (D3) and polyclonal. anti-nucleolin antibodies in Jurkat cells. The intera ction FJ-nucleolin was evidenced by Western blot and enzyme-linked imm unosorbent assay. Furthermore, purified nucleolin and D3 inhibited adh esion of Jurkat cells to immobilized FJ, suggesting that the interacti on was specific and that nucleolin mediated the binding.