INDUCTION OF PASSIVE HEYMANN NEPHRITIS WITH ANTIBODIES SPECIFIC FOR ASYNTHETIC PEPTIDE DERIVED FROM THE RECEPTOR-ASSOCIATED PROTEIN

Passive Heymann nephritis (pHN) is an experimental rat model for human membranous glomerulopathy. In pHN, the formation of subepithelial imm une deposits (ID) involves as antigenic targets the membrane glycoprot ein gp330/megalin and the 44-kD receptor-associated protein (RAP). A s ingle binding site for ID-inducing antibodies (Abs) was previously map ped to the 86 NH2-terminal amino acids of RAP (RAP(1-86)). To further narrow this epitope, Abs eluted from the glomeruli were immunoblotted on membranes that were loaded with overlapping synthetic peptides repr esenting the amino acid sequence of RAP (SPOTs system). Two adjacent A b-binding domains with the sequences PVRLAE (amino acids 39-44) and HS D-LKIQE (amino acids 46-53), which were separated by a single L residu e at amino acid 45, were detected. Rabbit Abs raised against synthetic peptides containing these domains individually (P-31-44 and P-46-53) failed to produce glomerular IDs. By contrast, Abs raised against a la rger composite peptide (P-31-53) induced IDs within 3 d that were firm ly cross-linked to the glomerular basement membrane. These data sugges t that Ab binding in vivo depends on the conformation of the antigenic target sequence that is preserved in the synthetic peptide P-31-53, w hich covers the entire Ab-binding domain of RAP but not in its subdoma ins, P-31-44, and P-46-53 Collectively, these results locate the sole ID-inducing epitope of RAP to amino acids 39-53.