2 MODES OF HOMOLOGOUS C3 DEPOSITION ON RAMOS BURKITTS-LYMPHOMA CELL SUBSTRAINS CO-EXPRESSING DAF (CD55), CD59, AND CR-2 (CD21), AND ON CELLS LACKING THEM

We established decay-accelerating factor (DAF)/CD59-positive and -negative substrains of a human B cell line, Ramos, R(DAF+/CD59+) and R(DAF-/CD59-) respectively. Unexpectedly, treatment of R(DAF+/CD59+) cells with Mg2+-EGTA-serum resulted in efficient C3 deposition, while treatment of R(DAF-/CD59-) cells did not. All six substrains of R(DAF-/CD59-) cells were CR2-negative, and treatment of the cells with M177 [a membrane cofactor protein (MCP) cofactor-blocking antibody] and/or acidic buffer only minimally affected the extent of C3 deposition. However, when R(DAF-/CD59-) cells were pretreated with M177 followed by incubation with low conductivity (3 mS) Mg2+-EGTA-serum, C3 deposition leading to effective cytolysis was provoked. On the other hand, all seven R(DAF+/CD59+) substrains were CR2-positive and could potentially induce C3 autoactivation without cytolysis under physiological conditions. Both M177 and pH again minimally affected the extent of C3 deposition. However, conductivity altered the sensitivity to C3: at under 3.0 mS, R(DAF+/CD59+) cells became almost insensitive to alternative pathway-mediated C3 deposition. Anti-CR2 partially inhibited C3 deposition on R(DAF+/CD59+) cells and C3 deposition was abrogated on the CR2-lacking R(DAF-/CD59-) cells, suggesting that CR2 was associated with the deposition of C3. These results, together with the finding that fluid phase activation of complement did not enhance C3 deposition, suggest that there are two distinct modes of spontaneous homologous C3 deposition on human lymphoma cells. In one case, CR2 or its related molecules participates in C3 deposition overcoming the protective function of DAF/MCP and this type of C3 deposition is maximized under physiological conditions. In the other case, C3 deposition is induced by another homologous C3 activator that becomes functional under low conductivity conditions and the absence of DAF/MCP. These two modes of homologous alternative pathway activation would explain the reported instances of spontaneous C3 deposition on human B lymphoid cell lines (under physiological conditions in the presence of DAF/MCP) and on paroxysmal nocturnal hemoglobinuria erythrocytes (under low conductivity conditions in the absence of DAF/MCP).