Tissue-specific distribution of the Goodpasture antigen demonstrated by 2-D electrophoresis and Western blotting

The target of autoantibodies in Goodpasture's disease, the Goodpastureantigen has recently been characterized as the NC1 domain ofthe 3 chain of type IV collagen. In order to study the Goodpastureantigen in different organs, NC1 domains were isolated frombasement membranes (BM) of human glomeruli (GBM), tubules (TBM),alveoli (ABM), placenta (PBM) and aorta (VBM). NC1 preparationswere separated by 2-D electrophoresis, and silver stained orimmunoblotted to determine the subunit structure and antigenicityof different basement membranes. All basement membranes containedmonomeric components of MW 26 kDa and 24 kDa, and associateddimers, corresponding to the 2-D location of 1(IV) and cc2(IV)chains respectively. However, GBM, ABM, and to a lesser extentTBM possessed an extra set of monomeric components of MW 28kDa and associated dimers corresponding to the proposed locationof 3(IV) and 4(IV) chains. 2-D-separated polypeptides were Westernblotted with autoantibodies from patients with Goodpasture'sdisease, a monoclonal antibody to the Goodpasture antigen (P1)and a monoclonal antibody to the bovine 3(IV) chain. The predominantbinding of all these reagents was to cationic 28 kDa monomersof GBM, ABM and TBM, corresponding to the 3(IV) chain, althoughautoantibodies and P1 also bound to neutral 28 kDa monomers,corresponding to the 4(IV) chain. Autoantibodies bound weaklyto more neutral components of PBM and VBM, but neither monoclonalantibody bound to these basement membranes. This study suggeststhat there are two separate type IV collagen networks: one containingthe l(IV) and 2(IV) chains appears to be present in all basementmembranes, and the other containing the 3(IV) and 4(IV) chainshas a tissue specific distribution. The latter network bearingthe Goodpasture antigen is expressed in the basement membranesof both kidney and lung, the organs involved in Goodpasture'sdisease.