DETECTION AND BIOCHEMICAL CHARACTERISTICS OF THE RECEPTOR FOR COMPLEXES OF SOLUBLE CD14 AND BACTERIAL LIPOPOLYSACCHARIDE

Soluble CD14 (sCD14) has been found to bind LPS and mediate LPS activa tion of several cell types. It has been postulated that sCD14-LPS comp lexes induce cell responses by interacting with a cell surface structu re, which, in turn, triggers cell activation. There has been no bioche mical evidence, however, for a direct interaction of sCD14 with a cell surface structure, and the putative receptor has not been identified. To rigorously test this hypothesis, we studied the interaction of hum an rsCD14 with cells in the absence of serum and in the presence and t he absence of LPS. We found 1) there was specific and saturable bindin g of I-125-sCD14, indicative of a typical receptor-ligand interaction, to several cell types, including endothelial cells, epithelial cells, astrocytes, and human monocytes; 2) specific binding to all the cell types and IL-6 induction in membrane-bound CD14 (mCD14)-negative cells occurred only when both sCD14 and LPS were present; 3) competitive di splacement experiments of I-125-sCD14 binding to astrocytes and Scatch ard plots revealed a binding of high affinity (K-d = 3.3+/-0.4 nM) and approximately 25,000 single class binding sites/cell; 4) the steady s tate for the association of I-125-sCD14 was obtained after 180-200 min ; 5) chemical cross-linking experiments revealed the association of sC D14 with a binding structure of approximately 216 kDa; 6) binding of I -125-sCD14 to CD14-expressing cell transfectants was about 50% lower t han that to nontransfected cells. Maximal binding, however, was recove red after removing mCD14, suggesting that the sCD14-LPS receptor may a lso interact with mCD14. These results provide direct biochemical evid ence for the existence of a cell surface signal-mediating binding stru cture for LPS-bearing sCD14 and suggest that this structure may repres ent the signaling unit of the postulated multimeric LPS receptor in mC D14-bearing cells.