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American Journal of Pathology, Vol 148, 1613-1623, Copyright © 1996 by American Society for Investigative Pathology

REGULAR ARTICLES

Induction of Heymann nephritis with a gp330/megalin fusion protein

R Raychowdhury, G Zheng, D Brown and RT McCluskey
Department of Pathology, Massachusetts General Hospital, Charlestown 02129, USA.

There is considerable evidence that glomerular deposits in Heymann nephritis, a rat model of membranous nephritis, result from shedding of immune complexes formed on podocytes and that the principal antigen is part of the extracellular domain of a cell surface glycoprotein receptor called gp330 or megalin. It has also been reported that the immunogen that induces Heymann nephritis is a complex formed between gp330 and the receptor-associated protein RAP. The recent elucidation of the primary structure of gp330 makes it possible to investigate the ability of defined portions of gp330, devoid of RAP, to induce Heymann nephritis. In the present study we show that a gp330-glutathione-S- transferase fusion protein, containing 137 amino acid residues (1114 to 1250) of the ectodomain, induces active Heymann nephritis and that heterologous antibodies against this fusion protein produce passive Heymann nephritis. By immunofluorescence, typical glomerular immunoglobulin deposits were found, but complement components were lacking and the rats did not develop proteinuria. In the active model, we obtained evidence indicating that the deposits contained portions of the ectodomain of gp330, including regions other than those of the fusion protein. Thus, the 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 with the fusion protein in vitro. Antibodies against the cytoplasmic domain of gp330 did not stain. Furthermore, we found that RAP was able to 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 inducing pathogenic antibodies of Heymann nephritis without a contributory role of RAP.

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