Self versus Nonself Discrimination by the Soluble Complement Regulators Factor H and FHL-1

Abstract

The plasma proteins Factor H (FH) and its alternate splice variant FH-like protein 1 (FHL-1) are the major regulators of the complement alternative pathway. The indiscriminate nature of alternative pathway activation necessitates the regulators to be host selective, but the underlying principles of selectivity remained largely elusive. By analyzing human FH and FHL-1 for protection of different host and foreign cells (rabbit and yeast), we uncovered a 2-fold discriminatory mechanism of FH in favor of self: relative to FHL-1, FH exhibits a regulatory benefit on self but importantly, also, a regulatory penalty on nonself surfaces, yielding a selectivity factor of ∼2.4 for sialylated host surfaces. We further show that FHL-1 possesses higher regulatory activity than known but is relatively unselective. The reason for this unexpected high activity of FHL-1 is the observation that the complement regulatory site in FH exceeds the established first four domains. Affinity for C3b, cofactor and decay-accelerating activities, and serum assays demonstrate that the regulatory site extends domains 1–4 and includes domains 5–7. But unlike FH, FHL-1 exhibits a fast plasma clearance in mice, occurs sparsely in human plasma (at one fortieth of the FH concentration), and resists deregulation by FH-related proteins. These physiological differences and its late phylogenetic occurrence argue that FHL-1 is crucial for local rather than systemic compartments. In conclusion, we demonstrate a 2-fold discriminatory power of FH to promote selectivity for self over foreign and show that FHL-1 is more active than known but specialized for regulation on local tissues.

Unravelling the effect of a potentiating anti-Factor H antibody on atypical hemolytic uremic syndrome associated factor H variants

“…3E and 3F ). This is consistent with data of others (12, 24, 25) and probably due to complete consumption of C3 in the FH depleted serum. Addition of physiological pdFH concentrations partly restores the C3 deposition to NPS control situation ( Fig.…”

“…Mutations R1210C, V1197A and G1194D result in destabilization of the tertiary structure (29), while V1197 is also shown to be important for binding to sialic acid (36). However, it is also noted that other ligands might have differential interactions in this binding region, such as heparin sulphates compared to sialic acid (36, 37) and is explained by the various effector functions of FH on different cells types, such as endothelial cells, glomerular membrane, retinal pigment epithelium, Bruch’s membrane in the eye, platelets or erythrocytes (24, 37, 38). A dual interaction of FH to C3b and sialic acid residues is crucial for protection of cells against unwanted complement activation on human cell surfaces (38).…”

Tuning the Functionality by Splicing: Factor H and Its Alternative Splice Variant FHL-1 Share a Gene but Not All Functions

“…On host-like sheep erythrocytes only FH CCPs 19-20 (carrying sialic acid and GAG binding properties), but not FH CCPs 6-8 (entailing only GAG binding functionality) exhibited functional competition with FH (24). A direct analysis of FH and FHL-1 regulatory capacity and surface specificity (host versus foreign) in FH/FHL-1-depleted serum revealed that both regulators protect foreign cell surfaces from complement attack at high concentrations, but FH was significantly more efficient in differentiating between foreign and host surfaces known to contain sialic acid moieties (14). This is in agreement with previous reports showing increased FH binding to sialylated surfaces (30,31).…”

“…Similar to C3b binding, the successive addition of CCP domains to CCP1-4 enhanced the activity, while the addition of CCP6 to CCP1-5 had the largest impact on overall regulatory function (14,35,36). However, already in the fluid phase, when sialic acid binding does not even have a role in direct comparison to FH, FHL-1 appears to be a significantly weaker cofactor for Factor Imediated C3b degradation (13,14). Remarkably, this is also observed for the engineered versions of FH like miniFH.…”

“…In the first report, the assay was performed on convertase pre-coated cells rather than in more defined (but less physiological) conditions on the SPR chip surface. However, because the solubility of FHL-1 in aqueous buffer at physiological pH and salt concentrations is rather limited, assays that require higher concentrations of FHL-1 stock solutions to be diluted into the assay conditions may need to be optimised for FHL-1 solubility (13,14). In addition to specialized assays that investigate CA or DAA, recently the activities of FH and FHL-1 were directly compared in several serum assays.…”