Molecular Architecture of the Goodpasture Autoantigen in Anti-GBM Nephritis (original) (raw)
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Journal of the American Society of Nephrology, 2005
Alport posttransplantation anti-glomerular basement membrane (GBM) nephritis is mediated by alloantibodies against the noncollagenous (NC1) domains of the ␣3␣4␣5(IV) collagen network, which is present in the GBM of the allograft but absent from Alport kidneys. The specificity of kidney-bound anti-GBM alloantibodies from a patient who had autosomal recessive Alport syndrome (ARAS) and developed posttransplantation nephritis was compared with that of Goodpasture autoantibodies from patients with autoimmune anti-GBM disease. Allograft-eluted alloantibodies reacted specifically with ␣3␣4␣5 NC1 hexamers, targeting their ␣3NC1 and ␣4NC1 subunits, and recognized a noncontiguous alloepitope formed jointly by the E A and E B regions of ␣3NC1 domain. In contrast, human Goodpasture autoantibodies recognized the separate E A and E B autoepitopes of ␣3NC1 but not the composite alloepitope. Molecular modeling of ␣3NC1 revealed that the alloepitope is more accessible within the NC1 hexamers than the partially sequestered Goodpasture autoepitopes. Overall, the specificity of alloantibodies indicated a selective lack of immune tolerance toward the ␣3 and ␣4(IV) collagen chains not expressed in patients with ARAS. Using COL4A3 knockout mice, a model of ARAS, it was shown further that acid-dissociated rather than native ␣3␣4␣5 NC1 hexamers elicited murine anti-GBM antibodies most closely resembling human ARAS alloantibodies. In contrast, ␣3NC1 monomers elicited Goodpasture-like murine antibodies, targeting the E A and E B autoepitopes. Thus, the identity of ␣3NC1 epitopes targeted by anti-GBM antibodies is strongly influenced by the molecular organization of the immunogen. These findings suggest that different isoforms of ␣3(IV) collagen may be implicated in the pathogenesis of ARAS posttransplantation anti-GBM nephritis and Goodpasture disease.
2005
Alport posttransplantation anti-glomerular basement membrane (GBM) nephritis is mediated by alloantibodies against the noncollagenous (NC1) domains of the ␣3␣4␣5(IV) collagen network, which is present in the GBM of the allograft but absent from Alport kidneys. The specificity of kidney-bound anti-GBM alloantibodies from a patient who had autosomal recessive Alport syndrome (ARAS) and developed posttransplantation nephritis was compared with that of Goodpasture autoantibodies from patients with autoimmune anti-GBM disease. Allograft-eluted alloantibodies reacted specifically with ␣3␣4␣5 NC1 hexamers, targeting their ␣3NC1 and ␣4NC1 subunits, and recognized a noncontiguous alloepitope formed jointly by the E A and E B regions of ␣3NC1 domain. In contrast, human Goodpasture autoantibodies recognized the separate E A and E B autoepitopes of ␣3NC1 but not the composite alloepitope. Molecular modeling of ␣3NC1 revealed that the alloepitope is more accessible within the NC1 hexamers than the partially sequestered Goodpasture autoepitopes. Overall, the specificity of alloantibodies indicated a selective lack of immune tolerance toward the ␣3 and ␣4(IV) collagen chains not expressed in patients with ARAS. Using COL4A3 knockout mice, a model of ARAS, it was shown further that acid-dissociated rather than native ␣3␣4␣5 NC1 hexamers elicited murine anti-GBM antibodies most closely resembling human ARAS alloantibodies. In contrast, ␣3NC1 monomers elicited Goodpasture-like murine antibodies, targeting the E A and E B autoepitopes. Thus, the identity of ␣3NC1 epitopes targeted by anti-GBM antibodies is strongly influenced by the molecular organization of the immunogen. These findings suggest that different isoforms of ␣3(IV) collagen may be implicated in the pathogenesis of ARAS posttransplantation anti-GBM nephritis and Goodpasture disease.
Molecular mapping of the Goodpasture's epitope for glomerulonephritis
Transactions of the American Clinical and Climatological Association, 2005
Goodpasture's syndrome is an autoimmune disease characterized by pulmonary hemorrhage, glomerulonephritis, and antiglomerular basement membrane (GBM) antibodies. We have studied a rat model with chimeric proteins (CPs) consisting of portions of the nephritogenic non-collagenous domain of alpha3 type IV collagen (alpha3(IV)NC1) and non-nephritogenic alpha1(IV)NC1. CPs with aminoterminal alpha3 that contains the major epitope for Goodpasture antibody binding induced EAG. We next immunized with D3, an alpha1(IV)NC1 CP with 69AA of alpha3(IV)NC1 (binds Goodpasture sera), D4, the D3 construct shortened by 4 AA (nonbinding), P9 and P10, single AA mutants (nonbinding) and S2, an alpha1(IV)NC1 with nine AA of alpha3(IV)NC1 (binding). GBM, S2 and D3 induced EAG. GBM immunized rats had intense IgG deposits but S2 and D3 rats had minimal deposits. A 13 mer rat peptide encompassing the aminoterminal site induced EAG sans antibody, while peptides not encompassing the region failed to induce ...
Current Opinion in Nephrology and Hypertension, 2011
Purpose of review-Goodpasture's (GP) disease is an autoimmune disorder characterized by the deposition of pathogenic autoantibodies in basement membranes of kidney and lung, which induces rapidly progressive glomerulonephritis and pulmonary hemorrhage. The target antigen is the α3NC1 domain of collagen IV, which is expressed in target organs as aα345 network. Recent studies of specificity and epitopes of GP autoantibodies and discovery of novel posttranslational modification of the antigen, a sulfilimine bond, provide further insight into mechanisms of initiation and progression of GP disease.
Kidney International, 2000
property likely contributes to both the fulminant nature of High affinity of anti-GBM antibodies from Goodpasture and this disease and its resistance to therapy, because persistent transplanted Alport patients to ␣3(IV)NC1 collagen. glomerular Ab deposition has the potential to produce continu-Background. Anti-glomerular basement membrane (anti-GBM) antibody-mediated diseases are characterized by rapidly ous inflammation, despite removal of circulating Abs and adeprogressive glomerulonephritis (RPGN) that often results in quate immunosuppression. irreversible loss of renal function and renal failure. Although many factors contribute to the fulminant nature and treatment resistance of this disease, we questioned whether high affinity Spontaneous production and deposition of anti-GBM autoantibody-␣3(IV) collagen interactions lead to persistent antibody deposition, thereby perpetuating inflammation. To antibodies characterize anti-GBM antibody-mediated address this hypothesis, the binding kinetics of human antidiseases. The disease spectrum includes patients with GBM antibodies (Ab) to ␣3(IV)NC1 were evaluated using an glomerulonephritis and pulmonary hemorrhage [Goodoptical biosensor interaction analysis. pasture syndrome (GS)], rapidly progressive glomerulo-Methods. Polyclonal anti-GBM Abs were purified by ␣3(IV)NC1 affinity chromatography from the sera of patients nephritis (RPGN) without clinical evidence of lung with anti-GBM AB-mediated diseases, including individuals involvement (type I), and isolated pulmonary hemorwith Goodpasture syndrome (GS), idiopathic RPGN (N ϭ 7), rhage [1, 2]. The mortality in patients with GS is high and Alport syndrome (AL) following kidney transplantation (11%), and 40 to 70% of patients with anti-GBM anti-(N ϭ 4). The affinity-binding characteristics of the autoantibodbody-mediated nephritis develop end-stage renal disease ies were determined using a biosensor analysis system, with immobilized bovine ␣3(IV)NC1 dimers. [3-6]. Of potential pathogenic relevance, anti-GBM anti-Results. All of the autoantibody preparations bound to bodies are frequently produced in patients with Alport's ␣3(IV)NC1, whereas none bound to ␣1(IV)NC1 (control). syndrome following renal transplantation, and a signifi-Purified, normal serum IgG did not bind to either antigen. cant fraction of these patients develops crescentic glo-Estimated dissociation constants (K d) for the purified autoantibodies were 1.39E-04 Ϯ 7.30E-05 s-l (GS) and 8.90E-05 Ϯ merulonephritis that results in the loss of graft function 2.80E-05 s-l (AL). Their estimated association constants (K a) [7, 8]. were 2.67Eϩ04 Ϯ 1.8Eϩ04 (M-ls-l) and 2.76Eϩ04 Ϯ 1.70Eϩ04 In the spontaneous form of the disease, the factors (M-ls-l) for GS and AL patients, respectively. By comparison leading to autoantibody production are unclear, howwith other Ab interactions, these Abs demonstrated high affinever, exposure of "hidden" basement membrane antiity, with relatively high on (binding) rates and slow off (dissociation) rates. gens and molecular mimicry have been postulated to be Conclusions. The results suggest that anti-GBM Abs bind involved in disease induction [9]. Nevertheless, it has rapidly and remain tightly bound to the GBM in vivo. This been conclusively demonstrated that the NC1 domain of ␣3 chain of type IV collagen [␣3(IV)NC1] is the major autoantigenic target [9-12].
Journal of the American Society of Nephrology, 2007
Goodpasture disease (GP) is defined by the presence of anti-glomerular basement membrane (anti-GBM) antibodies and rapidly progressive glomerulonephritis. Besides anti-GBM, many patients with GP produce anti-neutrophil cytoplasmic antibodies (ANCA). For elucidation of the pathophysiologic significance of ANCA in this setting, epitope and antigen specificity of the anti-GBM antibodies and antigen specificity of ANCA were studied. Bovine testis ␣(IV)NC1 (tNC1); recombinant human ␣1, ␣3, ␣4, and ␣5(IV)NC1 (r␣1 through r␣5); and three chimeric proteins that contain previously defined epitope regions designated E A , E B , and S2 were used to examine the anti-GBM antibodies by ELISA in 205 Chinese patients with GP with or without ANCA. In the 205 anti-GBM antibody-positive sera, 63 (30.7%) were also ANCA positive (61 myeloperoxidase-ANCA and six proteinase 3-ANCA, four being triple positive). All 205 sera recognized tNC1 and r␣3(IV)NC1. In the double-positive group, 54.0, 66.7, 71.4% of the sera could recognize r␣1, r␣4, and r␣5, respectively, compared with 49.3, 60.6, and 55.6% for patients with anti-GBM antibodies alone. The levels of the antibodies to r␣3, tNC1, and the ␣3/␣1 ratio were lower in the double-positive group than that in patients with anti-GBM antibody alone (P < 0.05). Most of the sera could recognize the epitope regions E A , E B , and S2, but the absorbance values to E A , E B , and S2 were lower in double-positive group (P < 0.05). Double-positive patients had a broader spectrum of anti-GBM antibodies and lower levels of antibodies against ␣3(IV)NC1 compared with that of patients with anti-GBM antibodies alone.
Goodpasture's epitope in development of experimental autoimmune glomerulonephritis in rats
Kidney International, 1996
Goodpasture's epitope in development of experimental autoimmune glomerulonephritis in rats. The Goodpasture's epitope (GP) has recently been localized to the last 36 AA of the non-collagenous (NC1) domain of the a3 chain of type IV collagen [a3(IV)]. Since a3(IV) induces glomerulonephritis (GN) in rats and rabbits, the purpose of the present study was to determine if the GP epitope itself could induce GN. We immunized rats with synthetic peptides of GP epitope, 36-mer, alone or as protein conjugates. Rats immunized with bovine GBM served as positive controls. Peptide immunized rats developed high titer antibodies to peptides, but only unconjugated 36-mer induced antibody against human and bovine GBM, but not to rat GBM. Acidic residues and the full length 36-mer were important in production of GBM reactive antibodies. Positive controls developed antibody to GBM without reactivity against 36-mer, had IgG and fibrin on the basement membrane, GN and proteinuria. Kidney eluted antibody was reactive with rat, bovine, and human GBM but not 36-mer. GN rat lymphocytes underwent blast transformation to GBM but not peptide, and peptide immunized animals responded only to the respective peptides. None of the animals immunized with GP peptide epitope, despite the development of anti-peptide antibodies or anti-GBM antibodies, developed any in vivo fixation of antibody to the GBM, abnormal proteinuria, or GN. The present study shows that the GP epitope is sufficient to induce an immune response to the epitope, but it is not sufficient to induce GN. This demonstrates that other factors or epitopes are important in the pathogenicity of GBM induced GN in this model. These remain to be delineated.
Journal of Biological Chemistry, 2000
Goodpasture (GP) disease is an autoimmune disorder in which autoantibodies against the ␣3(IV) chain of type IV collagen bind to the glomerular and alveolar basement membranes, causing progressive glomerulonephritis and pulmonary hemorrhage. Two major conformational epitope regions have been identified on the noncollagenous domain of type IV collagen (NC1 domain) of the ␣3(IV) chain as residues 17-31 (E A) and 127-141 (E B) (Netzer, K.-O. et al. (1999) J. Biol. Chem. 274, 11267-11274). To determine whether these regions are two distinct epitopes or form a single epitope, three GP sera were fractionated by affinity chromatography on immobilized NC1 chimeras containing the E A and/or the E B region. Four subpopulations of GP antibodies with distinct epitope specificity for the ␣3(IV)NC1 domain were thus separated and characterized. They were designated GP A , GP B , GP AB , and GP X , to reflect their reactivity with E A only, E B only, both regions, and neither, respectively. Hence, regions E A and E B encompass critical amino acids that constitute three distinct epitopes for GP A , GP B , and GP AB antibodies, respectively, whereas the epitope for GP X antibodies is located in a different unknown region. The GP A antibodies were consistently immunodominant, accounting for 60-65% of the total immunoreactivity to ␣3(IV)NC1; thus, they probably play a major role in pathogenesis. Regions E A and E B are held in close proximity because they jointly form the epitope for Mab3, a monoclonal antibody that competes for binding with GP autoantibodies. All GP epitopes are sequestered in the hexamer configuration of the NC1 domain found in tissues and are inaccessible for antibody binding unless dissociation of the hexamer occurs, suggesting a possible mechanism for etiology of GP disease. GP antibodies have the capacity to extract ␣3(IV)NC1 monomers, but not dimers, from native human glomerular basement membrane hexamers, a property that may be of fundamental importance for the pathogenesis of the disease.