Binding of Cryptococcus neoformans to heterologously expressed human complement receptors

Abstract

Previously, we demonstrated that monoclonal antibodies (MAb) directed against any of the three defined complement receptors (CR) for the third component of complement (CR1, CR3, and CR4) profoundly inhibited the binding of serum-opsonized Cryptococcus neoformans to monocyte-derived macrophages. These studies suggested either that a synergistic interaction between multiple CR was required for optimal binding of C. neoformans or that the MAb were exerting nonspecific effects (such as receptor coassociation). In the present studies, we took a novel approach to dissecting out the contributions of individual receptors to binding of a microbial pathogen. Chinese hamster ovary (CHO) cells stably transfected with human CR1, CR3, or CR4 were challenged with serum-opsonized C. neoformans. We found that CHO cells transfected with any of the three receptors bound C. neoformans, with the avidity of binding to CR3 being the greatest followed in decreasing order by CR1 and CR4. Following binding of C. neoformans to transfected CHO cells, most organisms remained surface attached only, although for each receptor a significant percentage (18.5 to 27.3%) of C. neoformans was internalized. Both C. neoformans and sheep erythrocytes that were selectively opsonized with the fragments of the third component of complement, C3b and iC3b, were bound preferentially by CHO cells transfected with CR1 and CR3, respectively. These data establish CR1, CR3, and CR4 as receptors independently capable of binding C. neoformans opsonized with fragments of C3. Moreover, our study demonstrates the usefulness of transfected cell lines as a powerful tool for identifying the contribution of individual receptors to the binding of a microbial pathogen.

The efficacy of complement‐mediated phagocytosis of Cryptococcus neoformans is dependent on the location of C3 in the polysaccharide capsule and involves both direct and indirectC3‐mediated interactions

“…Although prior investigators have shown that serum‐mediated binding of C. neoformans by human phagocytes is mediated through CR 32, this process had not been investigated with murine cells. We used mAb against CR subunits to investigate the involvement of these receptors in the phagocytosis of C. neoformans strains.…”

“…CR1 was dispensable, and CR4 played a role only in some strains. This suggests a species difference between mouse and human cells, since the latter have been shown to ingest complement‐opsonized C. neoformans through CR1, CR3, and CR4 32, 34. This discrepancy could indicate that the affinity of the CR for the different forms of C3 varies between the species and cell lines.…”

Complement and fungal pathogens: an update

“…116 The complement receptors CR1, CR3 and CR4 are the main effector molecules to mediate binding and phagocytosis of cryptococci, as demonstrated by monoclonal antibodies against the three receptors that block binding of the yeast cells. 117,118 A ranking for the CRs demonstrates the dominance of CR3. 112 The complement receptors CR3 and CR4 are also involved in complement-independent, antibody-mediated phagocytosis of C. neoformans.…”

The role of complement in invasive fungal infections

“…Further studies established that CR1, CR3 and CR4 act independently of each other in their binding opsonized Cr. neoformans 70 . The complement receptors CR3 and CR4 are also involved in the complement‐dependent antibody‐mediated phagocytosis of C. neoformans .…”