INDUCTION OF HEYMANN NEPHRITIS WITH A GP330 MEGALIN FUSION PROTEIN/

There is considerable evidence that glomerular deposits in Heymann nep hritis, a rat model of membranous nephritis, result from shedding of i mmune complexes formed on podocytes and that the principal antigen is part of the extracellular domain of a cell surface glycoprotein recept or called gp330 or megalin. It has also been reported that the immunog en that induces Heymann nephritis is a complex formed between gp330 an d the receptor-associated protein RAP, The recent elucidation of the p rimary structure of gp330 mates it possible to investigate the ability of defined portions of gp330, devoid of RAP, to induce Heymann nephri tis. Irt the present study we show that a gp330-glutathione-S-transfer ase fusion protein, containing 137 amino acid residues (1114 to 1250) of the ectodomain, induces active Heymann nephritis and that heterolog ous antibodies against this fusion protein produce passive Heymann nep hritis. By immunofluorescence, typical glomerular immunoglobulin depos its were found, but complement components were lacking and the rats di d not develop proteinuria. In the active model, we obtained evidence i ndicating that the deposits contained portions of the ectodomain of gp 330, including regions other than those of the fusion protein. Thus, t he deposits were stained by polyclonal antibodies to gp330 and to the gp330 fusion protein, as well as by two monoclonal antibodies reactive with portions of the ectodomain of gp330, only one of which reacted w ith the fusion protein in vitro. Antibodies against the cytoplasmic do main of gp330 did not stain. Furthermore, we found that RAP was able t o bind to gp330 in the glomerular deposits but not to the gp330 fusion protein in vitro. The results show that the region of gp330 spanning amino acid residues 1114 to 1250 contains peptides capable of inductin g pathogenic antibodies of Heymann nephritis without a contributory ro le of RAP.