Complement component C3 fixes selectively to the major outer membrane protein (MOMP) of Legionella pneumophila and mediates phagocytosis of liposome-MOMP complexes by human monocytes

Abstract

Legionella pneumophila is a facultative intracellular bacterial pathogen that parasitizes human monocytes and alveolar macrophages. Previous studies from this laboratory have shown that monocyte complement receptors CR1 and CR3 and complement component C3 in serum mediate L. pneumophila phagocytosis. In this study, we have explored C3 fixation to L. pneumophila. We developed a whole-cell enzyme-linked immunosorbent assay (ELISA) to measure C3 fixation to the bacterial surface. By this assay, C3 fixes to L. pneumophila that are opsonized in fresh nonimmune serum, and C3 fixation takes place via the alternative pathway of complement activation. Immunoblot analysis of opsonized L. pneumophila indicated that C3 fixes selectively to specific acceptor molecules of L. pneumophila. Consistent with this, when nitrocellulose blots of whole L. pneumophila or bacterial components are incubated in fresh nonimmune serum, C3 fixes exclusively to the major outer membrane protein (MOMP) of L. pneumophila, a porin; C3 does not fix to L. pneumophila LPS on these blots. To further explore the role of MOMP in C3 fixation and phagocytosis, we reconstituted purified MOMP into liposomes. By the ELISA, MOMP-liposomes, but not plain liposomes lacking MOMP, avidly fix C3. Consistent with a dominant role for MOMP in C3 fixation, MOMP-liposomes form a C3 complex of the same apparent molecular weight as whole L. pneumophila in nonimmune serum. Opsonized radioiodinated MOMP-liposomes avidly adhere to monocytes, and adherence is dose dependent upon serum. By electron microscopy, opsonized MOMP-liposomes are efficiently phagocytized by human monocytes, and phagocytosis takes place by a conventional appearing form of phagocytosis. This study demonstrates that C3 fixes selectively to the MOMP of L. pneumophila, and that, in the presence of nonimmune serum, MOMP can mediate phagocytosis of liposomes and, potentially, phagocytosis of intact L. pneumophila by human monocytes.