Abstract
The heterotrimeric globular head (gC1 q) domain of human Cl q is made up of the C-terminal ends of the three individual chains, ghA, ghB, and ghC. A candidate receptor for the gClq domain is a multi-functional pattern recognition protein, gC1 qR. Since understanding of gC1 qR and gClq interaction could provide an insight into the pleiotropic functions of gCl qR, this study was undertaken to identify the gCl qR-binding site on the gClq domain, using the recombinant ghA, ghB, and ghC modules and their substitution mutants. Our results show that ghA, ghB, and ghC modules can interact with gClqR independently, thus reinforcing the notion of modularity within the gClq domain of human Cl q. Mutational analysis revealed that while Arg162 in the ghA module is central to interaction between gClqR and Clq, a single amino acid substitution (arginine to glutamate) in residue 114 of the ghB module resulted in enhanced binding. Expression of gClqR and Cl q in adherent monocytes with or without pro-inflammatory stimuli was also analyzed by qPCR; it showed an autocrine/paracrine basis of Cl q and gClqR interaction. Microscopic studies revealed that Clq and gC1 qR are colocalized on PBMCs. Cell proliferation assays indicated that ghA, ghB, and ghC modules were able to attenuate phytohemagglutinin-stimulated proliferation of PBMCs. Addition of gClqR had an additive effect on the anti-proliferative effect of globular head modules. In summary, our results identify residues involved in Clq-gClqR interaction and explain, to a certain level, their involvement on the immune cell surface, which is relevant for Clq-induced functions including inflammation, infection, and immunity.