SENSITIVE RESPONSES OF LEUKOCYTES TO LIPOPOLYSACCHARIDE REQUIRE A PROTEIN DISTINCT FROM CD14 AT THE CELL-SURFACE

Responses of leukocytes to low concentrations of LPS require the expre ssion of membrane-associated CD14 (mCD14) on the cell surface; mCD14 i s, however, a glycosyl-phosphatidylinositol-anchored protein and, ther efore, a poor candidate for transducing signals across the plasma memb rane. The role of other cell surface proteins in response of leukocyte s to LPS was tested by measuring IL-6 secretion of cultured human mono cytes and adhesion of PMN in response to LPS after treatment of the ce lls with trypsin. Trypsin abolished leukocyte integrin-mediated adhesi on of PMN in response to LPS, but the trypsimized cells remained respo nsive to the alternate agonists TNF, formyl peptide, and PMA. Similarl y, trypsin treatment of monocytes inhibited IL-6 production in respons e to LPS, but not to formyl peptide or PMA. No change in cell surface expression of mCD14 was observed by cytofluorometry, and no proteolysi s of mCD14 was detected by immunoblot analysis. These results suggest that digestion of a cell surface protein distinct from mCD14 must acco unt for the loss of responsiveness to LPS. The uptake by monocytes of [H-3]LPS presented in LPS-soluble CD14 complexes was also inhibited by trypsin treatment. Monomeric LPS was taken up superstoichiometrically to mCD14 with a ratio of up to 15:1, and trypsin treatment decreased this uptake by more than half. This suggests that a cell surface prote in may function in accepting LPS from mCD14. Together the results sugg est that one or more trypsin-sensitive cell surface proteins distinct from CD14 participate in both the uptake of LPS by leukocytes and the initiation of the signaling process.