title: HLA-B27 in transgenic rats forms disulfide-linked heavy chain oligomers and multimers that bind to the chaperone BiP.
abstract: To test the hypothesis that HLA-B27 predisposes to disease by forming disulfide-linked homodimers, we examined rats transgenic for HLA-B27, mutant Cys(67)Ser HLA-B27, or HLA-B7. In splenic Con A blasts from high transgene copy B27 lines that develop inflammatory disease, the anti-H chain mAb HC10 precipitated four bands of molecular mass 78-105 kDa and additional higher molecular mass material, seen by nonreducing SDS-PAGE. Upon reduction, all except one 78-kDa band resolved to 44 kDa, the size of the H chain monomer. The 78-kDa band was found to be BiP/Grp78, and the other high molecular mass material was identified as B27 H chain. Analysis of a disease-resistant low copy B27 line showed qualitatively similar high molecular mass bands that were less abundant relative to H chain monomer. Disease-prone rats with a Cys(67)Ser B27 mutant showed B27 H chain bands at 95 and 115 kDa and a BiP band at 78 kDa, whereas only scant high molecular mass bands were found in cells from control HLA-B7 rats. (125)I-surface labeled B27 oligomers were immunoprecipitated with HC10, but not with a mAb to folded B27-beta(2)-microglobulin-peptide complexes. Immunoprecipitation of BiP with anti-BiP Abs coprecipitated B27 H chain multimers. Folding and maturation of B27 were slow compared with B7. These data indicate that disulfide-linked intracellular H chain complexes are more prone to form and bind BiP in disease-prone wild-type B27 and B27-C67S rats than in disease-resistant HLA-B7 rats. The data support the hypothesis that accumulation of misfolded B27 participates in the pathogenesis of B27-associated disease.
authors.affiliation: Harold C. Simmons Arthritis Research Center and Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Tri Minh
authors.initials: TM
authors.lastname: Tran
authors.forename: Nimman
authors.initials: N
authors.lastname: Satumtira
authors.forename: Martha L
authors.initials: ML
authors.lastname: Dorris
authors.forename: Ekkehard
authors.initials: E
authors.lastname: May
authors.forename: Andrew
authors.initials: A
authors.lastname: Wang
authors.forename: Eiichi
authors.initials: E
authors.lastname: Furuta
authors.forename: Joel D
authors.initials: JD
authors.lastname: Taurog
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001666:Binding Sites, Antibody; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000276:immunology / Q000378:metabolism; D000091342:Endoplasmic Reticulum Chaperone BiP; D015796:HLA-B27 Antigen / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D015793:HLA-B7 Antigen / Q000378:metabolism; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D011485:Protein Binding / Q000276:immunology; D011499:Protein Processing, Post-Translational / Q000276:immunology; D021122:Protein Subunits / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D051381:Rats; D011917:Rats, Inbred Lew; D013154:Spleen / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D019076:Transgenes / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D004220:Disulfides; D000091342:Endoplasmic Reticulum Chaperone BiP; D015796:HLA-B27 Antigen; D015793:HLA-B7 Antigen; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D021122:Protein Subunits