Boon-Leong Lim

gC1q-R/p33, a member of a new class of multifunctional and multicompartmental cellular proteins, is involved in inflammation and infection.

Human gC1q‐R (p33, p32, C1qBP, TAP) is a ubiquitously expressed, multiligand‐binding, multicompartmental cellular protein involved in various ligand‐mediated cellular responses. Although expressed on the surface of cells, an intriguing feature of the membrane‐associated form of gC1q‐R is that its translated amino acid sequence does not predict the presence of either a sequence motif compatible with a transmembrane segment or a consensus site for a glycosylphosphatidylinositol anchor. Moreover, the N‐terminal sequence of the pre‐pro‐protein of gC1q‐R contains a motif that targets the molecule to the mitochondria and as such was deemed unlikely to be expressed on the surface. However, several lines of experimental evidence clearly show that gC1q‐R is present in all compartments of the cell, including the extracellular cell surface. First, surface labeling of B lymphocytes with the membrane‐impermeable reagent sulfosuccinimidyl 6‐(biotinamido)hexanoate shows specific biotin incorporation into the surface‐expressed but not the intracellular form of gC1q‐R. Second, FACS and confocal laser scanning microscopic analyses using anti‐gC1q‐R IgG mAb 60.11 or 74.5.2, and the fluorophore Alexa 488‐conjugated F(ab′)2 goat anti‐mouse IgG as a probe, demonstrated specific staining of Raji cells (>95% viable). Three‐dimensional analyses of the same cells by confocal microscopy showed staining distribution that was consistent with surface expression. Third, endothelial gC1q‐R, which is associated with the urokinase plasminogen activator receptor, and cytokeratin 1 bind 125I‐high molecular weight kininogen in a specific manner, and the binding is inhibited dose‐dependently by mAb 74.5.2 recognizing gC1q‐R residues 204–218. Fourth, native gC1q‐R purified from Raji cell membranes but not intracellular gC1q‐R is glycosylated, as evidenced by a positive periodic acid Schiff stain as well as sensitivity to digestion with endoglycosidase H and F. Finally, cross‐linking experiments using C1q as a ligand indicate that both cC1q‐R and gC1q‐R are co‐immunoprecipitated with anti‐C1q. Taken together, the evidence accumulated to date supports the concept that in addition to its intracellular localization, gC1q‐R is expressed on the cell surface and can serve as a binding site for plasma and microbial proteins, but also challenges the existing paradigm that mitochondrial proteins never leave their designated compartment. It is therefore proposed that gC1q‐R belongs to a growing list of a class of proteins initially targeted to the mitochondria but then exported to different compartments of the cell through specific mechanisms which have yet to be identified. The designation ‘multifunctional and multicompartmental cellular proteins’ is proposed for this class of proteins.

Interaction of human lung surfactant proteins A and D with mite (Dermatophagoides pteronyssinus) allergens

Human lung surfactant proteins A (SP‐A) and D (SP‐D) are both collagenous C‐type lectins which appear to mediate antimicrobial activity by binding to carbohydrates on micro‐organisms and to receptors on phagocytic cells. Purified native SP‐A and SP‐D, isolated from human bronchoalveolar lavage fluid, were found to bind to whole mite extracts (Dermatophagoides pteronyssinus) and the purified allergen Der p I, in a carbohydrate‐specific and calcium‐dependent manner. Binding was inhibited by ethylenediamine tetra‐acetic acid (EDTA) as well as by maltose in the case of SP‐D, or mannose in the case of SP‐A. A recombinant polypeptide, which trimerized to form the neck region and carbohydrate recognition domains of SP‐D, also inhibited the binding of native SP‐D to the whole mite extract and Der p I. Both SP‐A and SP‐D did not bind to deglycosylated whole mite extracts or to recombinant Der p proteins, which lacked carbohydrate residues. These results suggest that the ability of surfactant proteins to bind certain allergens is mediated through their carbohydrate‐recognition domains (CRDs) interacting with carbohydrate residues on the allergens. Moreover, SP‐A and SP‐D were found to inhibit allergen‐specific IgE binding to the mite extracts either via steric hindrance or competitive binding. It is therefore possible that SP‐A and SP‐D may be involved in the modulation of allergen sensitization and/or the development of allergic reactions.