FHR-1 binds to C-reactive protein and enhances rather than inhibits complement activation

Factor H (FH)-related protein 1 (FHR-1) is one of the five human factor H-related proteins, which share sequence and structural homology with the alternative pathway complement inhibitor FH. Genetic studies on disease associations and functional analyses indicate that FHR-1 enhances complement activation by co mpetitive inhibition of FH binding to some surfaces and immune proteins. We have recen tly shown that FHR-1 binds to pentraxin 3. Here, our aim was to investigate whether FH R-1 binds to another pentraxin, C-reactive protein (CRP), analyze the functional relevance of this interaction and study the role of FHR- 1 in complement activation and regulation. FHR- 1 did not bind to native, pentameric CRP but it bound strongly to monomeric CRP via its C-term inal domains. FHR-1 at high concentration competed with FH for CRP binding, indicating possible complement de -regulation also on this ligand. FHR-1 did not inhibi t regulation of solid phase C3 convertase by FH and did not inhibit terminal complement complex forma tion induced by zymosan. On the contrary, by binding C3b, FHR-1 allowed C3 convertase form ation and thereby enhanced complement activation. FHR-1/CRP interacti ons increased complement activ ation via the classical and alternative pathways on surfaces such as th e extracellular matrix and necrotic cells. Altogether, these results identify CRP as a lig and for FHR-1 and suggest that FHR-1 enhances rather than inhibits complement activation, wh ich may explain the protective effect of FHR-1 deficiency in age-related macular degeneration

FHR-5 Serum Levels and CFHR5 Genetic Variations in Patients With Immune Complex-Mediated Membranoproliferative Glomerulonephritis and C3-Glomerulopathy

Factor H-related protein 5 (FHR-5) is a member of the complement Factor H protein family. Due to the homology to Factor H, the main complement regulator of the alternative pathway, it may also be implicated in the pathomechanism of kidney diseases where Factor H and alternative pathway dysregulation play a role. Here, we report the first observational study on CFHR5 variations along with serum FHR-5 levels in immune complex-mediated membranoproliferative glomerulonephritis (IC-MPGN) and C3 glomerulopathy (C3G) patients together with the clinical, genetic, complement, and follow-up data.A total of 120 patients with a histologically proven diagnosis of IC-MPGN/C3G were enrolled in the study. FHR-5 serum levels were measured in ELISA, the CFHR5 gene was analyzed by Sanger sequencing, and selected variants were studied as recombinant proteins in ELISA and surface plasmon resonance (SPR).Eight exonic CFHR5 variations in 14 patients (12.6%) were observed. Serum FHR-5 levels were lower in patients compared to controls. Low serum FHR-5 concentration at presentation associated with better renal survival during the follow-up period; furthermore, it showed clear association with signs of complement overactivation and clinically meaningful clusters.Our observations raise the possibility that the FHR-5 protein plays a fine-tuning role in the pathogenesis of IC-MPGN/C3G

Suppression of innate and adaptive B cell activation pathways by antibody coengagement of FcgammaRIIb and CD19.

The Fc receptor (FcgammaRIIb) inhibits B cell responses when coengaged with B cell receptor (BCR), and has become a target for new autoimmune disease therapeutics. For example, BCR and FcgammaRIIb coengagement via the Fc-engineered anti-CD19 XmAb5871 suppresses humoral immune responses. We now assess effects of XmAb5871 on other activation pathways, including the pathogen-associated molecular pattern receptor, TLR9. Since TLR9 signaling is implicated in autoimmune diseases, we asked if XmAb5871 could inhibit TLR9 costimulation. We show that XmAb5871 decreases ERK and AKT activation, cell proliferation, cytokine, and IgG production induced by BCR and/or TLR9 signals. XmAb5871 also inhibited differentiation of citrullinated peptide-specific plasma cells from rheumatoid arthritis patients. XmAb5871 may therefore have potential to suppress pathogenic B cells in autoimmune diseases

The Murine Factor H-Related Protein FHR-B Promotes Complement Activation

Factor H-related (FHR) proteins consist of varying number of complement control protein domains that display various degrees of sequence identity to respective domains of the alternative pathway complement inhibitor factor H (FH). While such FHR proteins are described in several species, only human FHRs were functionally investigated. Their biological role is still poorly understood and in part controversial. Recent studies on some of the human FHRs strongly suggest a role for FHRs in enhancing complement activation via competing with FH for binding to certain ligands and surfaces. The aim of the current study was the functional characterization of a murine FHR, FHR-B. To this end, FHR-B was expressed in recombinant form. Recombinant FHR-B bound to human C3b and was able to compete with human FH for C3b binding. FHR-B supported the assembly of functionally active C3bBb alternative pathway C3 convertase via its interaction with C3b. This activity was confirmed by demonstrating C3 activation in murine serum. In addition, FHR-B bound to murine pentraxin 3 (PTX3), and this interaction resulted in murine C3 fragment deposition due to enhanced complement activation in mouse serum. FHR-B also induced C3 deposition on C-reactive protein, the extracellular matrix (ECM) extract Matrigel, and endothelial cell-derived ECM when exposed to mouse serum. Moreover, mouse C3 deposition was strongly enhanced on necrotic Jurkat T cells and the mouse B cell line A20 by FHR-B. FHR-B also induced lysis of sheep erythrocytes when incubated in mouse serum with FHR-B added in excess. Altogether, these data demonstrate that, similar to human FHR-1 and FHR-5, mouse FHR-B modulates complement activity by promoting complement activation via interaction with C3b and via competition with murine FH

Complement Factor H-Related Proteins FHR1 and FHR5 Interact With Extracellular Matrix Ligands, Reduce Factor H Regulatory Activity and Enhance Complement Activation

Components of the extracellular matrix (ECM), when exposed to body fluids may promote local complement activation and inflammation. Pathologic complement activation at the glomerular basement membrane and at the Bruch’s membrane is implicated in renal and eye diseases, respectively. Binding of soluble complement inhibitors to the ECM, including factor H (FH), is important to prevent excessive complement activation. Since the FH-related (FHR) proteins FHR1 and FHR5 are also implicated in these diseases, our aim was to study whether these FHRs can also bind to ECM components and affect local FH activity and complement activation. Both FH and the FHRs showed variable binding to ECM components. We identified laminin, fibromodulin, osteoadherin and PRELP as ligands of FHR1 and FHR5, and found that FHR1 bound to these ECM components through its C-terminal complement control protein (CCP) domains 4-5, whereas FHR5 bound via its middle region, CCPs 3-7. Aggrecan, biglycan and decorin did not bind FH, FHR1 and FHR5. FHR5 also bound to immobilized C3b, a model of surface-deposited C3b, via CCPs 3-7. By contrast, soluble C3, C3(H2O), and the C3 fragments C3b, iC3b and C3d bound to CCPs 8-9 of FHR5. Properdin, which was previously described to bind via CCPs 1-2 to FHR5, did not bind in its physiologically occurring serum forms in our assays. FHR1 and FHR5 inhibited the binding of FH to the identified ECM proteins in a dose-dependent manner, which resulted in reduced FH cofactor activity. Moreover, both FHR1 and FHR5 enhanced alternative complement pathway activation on immobilized ECM proteins when exposed to human serum, resulting in the increased deposition of C3-fragments, factor B and C5b-9. Thus, our results identify novel ECM ligands of FH family proteins and indicate that FHR1 and FHR5 are competitive inhibitors of FH on ECM and, when bound to these ligands, they may enhance local complement activation and promote inflammation under pathological conditions