Abstract. Goodpasture's disease is a severe nephritis characterized by autoantibodies to the ␣3 chain of type IV collagen, ␣3(IV)NC1, in the glomerular basement membrane. The disease is very strongly associated with HLA-DR15, the affinities of ␣3(IV)NC1 peptides for DR15 are known, and elution experiments have identified major naturally processed sequences. Here, the fine specificity and cytokine profile of ␣3(IV)NC1-reactive T cells from patients with Goodpasture's disease is defined. Peripheral blood mononuclear cells from patients at diagnosis proliferated in response to significantly more peptides ( 2 ϭ 8.6, P ϭ 0.004) from a panel spanning the sequence of ␣3(IV)NC1 than did those from control DR15-positive donors and were highly focused (P ϭ 0.0002, binomial distribution) on two peptides, ␣3 71-90 and ␣3 131-150 .

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The mechanisms underlying apparently spontaneous autoimmune diseases, such as autoimmune hemolytic anemia (AIHA) in New Zealand Black (NZB) mice, are unknown. Here, we determine the contribution of the dominant red blood cell (RBC) autoantigen, the anion exchanger protein Band 3, to the development of NZB autoimmune responses. The approach was to prevent Band 3 expression in NZB mice by disrupting the AE1 gene. AE1 ؊/؊ NZB mice produced RBC autoantibodies at the same levels as the wild-type strain, but they differed in recognizing antigens that correspond to glycophorins, rather than Band 3. Splenic T-helper (Th) cells from wild-type NZB mice proliferated strongly against multiple Band 3 peptides, particularly the dominant epitope within aa861-874. This helper response was severely attenuated in AE1 ؊/؊ animals, leaving only weak proliferation to peptide aa861-874. The results demonstrate that the defect in self-tolerance in NZB AIHA is directed to the RBC type, and is not specific for, or dependent on, Band 3. However, the predisposition to RBC autoimmunity may be focused onto Band 3 by weak Th cell cross-reactivity between the helper dominant epitope and an exogenous antigen. The redundancy of the major autoantigen illustrates the requirement for specific therapy to induce dominant forms of tolerance, such as T-cell regulation. IntroductionThe underlying mechanisms that drive damaging responses in autoimmune disease are poorly understood. Valuable insights have been obtained from the study of murine models. For example, genetic analyses have shown that different loci govern susceptibility to autoimmunity and determine which organs are affected, 1 while functional studies have identified particular autoantigens and highlighted the role of T-cell subsets in pathogenesis. 2 However, the stimuli that initiate and perpetuate apparently spontaneous autoaggression remain the subject of controversy.Autoimmune hemolytic anemia (AIHA) is a classic example of an antibody-mediated autoimmune disease, and provides an opportunity to study the development of specific pathogenic responses, since the major human 3-5 and murine 6,7 autoantigens have been identified. Efforts to identify possible mechanisms of autoaggression often focus on spontaneous AIHA in the New Zealand Black (NZB) strain of mouse. The mice develop signs of anemia by 5 to 6 months of age due to the production autoantibodies that bind red blood cells (RBCs) and cause erythrophagocytosis. 8 Band 3, the RBC anion exchanger protein (AE1), is the dominant autoantigen. 6,7,9,10 Eluted autoantibodies exclusively immunoprecipitate Band 3 from murine RBCs, 6 and 4 of 5 IgG anti-RBC monoclonal autoantibodies (mAbs) derived from NZB mice react with this antigen. 9,10 The autoantibody response is helper dependent 11,12 and Band 3 is also the major target for autoreactive T-helper (Th) cells. 7,[13][14][15][16] Splenic CD4 ϩ T cells from NZB mice, but not H-2-matched healthy strains, proliferate in vitro in response to Band 3, 7,13 and initial epitope mapping studies id...