Factor H (fH) is a plasma inhibitor of the alternative pathway (AP) complement. It is composed of 20 short consensus repeat (SCR) domains. The complement regulatory function of fH is located in the N-terminal SCR1-4, whereas the two C-terminal SCRs of fH (SCR 19 and 20) are critical for interacting with surface deposited C3b/C3d in the context of host cell-specific polyanionic constituents. Mutations in SCR19-20 of human fH impair its interaction with host cells and predispose to the development of atypical hemolytic uremic syndrome (aHUS). Among the various SCR19-20 mutations of fH that are found in aHUS patients, the R1210C mutation is of particular interest as a recent genetic study has identified this mutation to be also a high-penetrant mutation for the development of age-related macular degeneration (AMD). The R1210C variant of human fH showed reduced binding to C3b/C3d, heparin and endothelial cells, yet R1210 is not conserved across species (D1210 in mouse, G1210 in rat, P1210 in cows and pigs). We hypothesize that the change of amino acid at position 1210 of human fH to cysteine, rather than the simple loss of arginine, is critical for conferring the risk of aHUS and AMD. In this exploratory grant, we propose two specific aims to test this hypothesis.
Specific Aim 1. We will perform in vitro mutagenesis studies of human and mouse fH at residue 1210 and test the prediction that a mouse D1210C mutant will, like human R1210C mutant, have impaired binding to C3b, heparin and endothelial cells. On the other hand, we predict that mutations of R1210 in human fH to R1210D, R1210G and R1210P (to mouse, rat and cow/pig residue, respectively), and mutations of D1210 in mouse fH to D1210R, D1210G and D1210P (to human, rat, and cow/pig residue, respectively), are tolerated and produce no functional consequences for C3b-, heparin- and endothelial cell-binding.
Specific Aim 2. We will generate by gene targeting a D1210C mutant mouse and determine if this mouse develops aHUS and/or retinal degeneration resembling human AMD. These studies will shed new light on structure/function knowledge of the fH C-terminal domain and establish how specific mutations in this domain may differentially affect fH function in different tissues and species. Furthermore, they will help us understand the pathogenesis of two fH-related human diseases and provide a much- needed mouse model for testing anti-complement therapies for these pathologies.
This project uses the mouse as an animal model to study the pathogenesis and therapy of two human diseases, one a kidney disease that requires dialysis and transplantation and is often lethal, and the other is a blinding disorder affecting many elderl individuals. Both diseases are caused by defects in a blood protein of the patients that is normally involved in controlling infection.