C3b covalently associated to tetanus toxin modulates TT processing and presentation by U937 cells

Complement protein C3, like C4 and α2-macroglobulin (α2M), is a potentially bivalent ligand: (1) its proteolytic fragment, C3b, is able to interact covalently with antigens, and (2) this bound fragment is able to interact non-covalently with specific complement receptors of antigen presenting cells (APC). The formation of antigen-C3b complexes frequently occurs in vivo at inflammatory sites during the early stages of an immune response. Tetanus toxin (TT)-C3b covalent complexes, prepared from purified proteins, were used to study how C3b association influences the handling of TT by U937 cells used as APC. TT-specific T cell proliferation following TT-C3b processing was observed at a concentration when TT alone was inefficient. Whereas TT pinocytic uptake was low, TT-C3b uptake, through the help of complement receptor CR1, was three times higher. Free TT was rapidly transported to the lysosomes where it was proteolysed, whereas TT-C3b complexes were first retained in the endosomes and underwent only limited proteolysis. While the ester link of the complexes was fairly stable in the endosomes, it was gradually hydrolysed in the lysosomes with ensuing efficient proteolysis of the two proteins. This reflects the fact that associated C3b escorts TT during intracellular trafficking in the APC, and influences antigen processing. A triple role of C3b escorting antigen resides at the level of antigen uptake, routing, and proteolysis inside U937 cells, thus modulating antigen-dependent T cell proliferation.