Tissue-specific distribution of the Goodpasture antigen demonstrated by 2-D electrophoresis and western blotting.
The target of autoantibodies in Goodpasture's disease, the Goodpasture antigen has recently been characterized as the NC1 domain of the alpha 3 chain of type IV collagen. In order to study the Goodpasture antigen in different organs, NC1 domains were isolated from basement membranes (BM) of human glomeruli (GBM), tubules (TBM), alveoli (ABM), placenta (PBM) and aorta (VBM). NC1 preparations were separated by 2-D electrophoresis, and silver stained or immunoblotted to determine the subunit structure and antigenicity of different basement membranes. All basement membranes contained monomeric components of MW 26 kDa and 24 kDa, and associated dimers, corresponding to the 2-D location of alpha 1(IV) and alpha 2(IV) chains respectively. However, GBM, ABM, and to a lesser extent TBM possessed an extra set of monomeric components of MW 28 kDa and associated dimers corresponding to the proposed location of alpha 3 (IV) and alpha 4 (IV) chains. 2-D-separated polypeptides were Western blotted with autoantibodies from patients with Goodpasture's disease, a monoclonal antibody to the Goodpasture antigen (P1) and a monoclonal antibody to the bovine alpha 3 (IV) chain. The predominant binding of all these reagents was to cationic 28 kDa monomers of GBM, ABM and TBM, corresponding to the alpha 3 (IV) chain, although autoantibodies and Pl also bound to neutral 28 kDa monomers, corresponding to the alpha 4 (IV) chain. Autoantibodies bound weakly to more neutral components of PBM and VBM, but neither monoclonal antibody bound to these basement membranes.(ABSTRACT TRUNCATED AT 250 WORDS)
Derry
C J
CJ
Department of Medicine, Royal Postgraduate Medical School, Hammersmith Hospital, London, UK.
Pusey
C D
CD
D019867:Anti-Glomerular Basement Membrane Disease / Q000276:immunology*; D001324:Autoantigens / Q000032:analysis* / Q000276:immunology; D001485:Basement Membrane / Q000737:chemistry; D015153:Blotting, Western; D003094:Collagen / Q000032:analysis* / Q000276:immunology; D024141:Collagen Type IV; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D007678:Kidney Glomerulus / Q000737:chemistry; D009928:Organ Specificity
D001324:Autoantigens; D024141:Collagen Type IV; C057112:type IV collagen alpha3 chain; D003094:Collagen