Immunoscreening of phage-displayed cDNA-encoded polypeptides identifies B cell targets in autoimmune disease.
Characterisation of self-antigens can contribute to an understanding of the aetiology of autoimmune disorders as well as to the development of new therapies and diagnostic methods. The present study was undertaken to investigate the applicability of complementary DNA (cDNA) phage-display technology to the identification of autoantigens recognised by the humoral response in autoimmune disease. Using systemic lupus erythematosus (SLE) as a model system, a pool of patient immunoglobulin G (IgG) was biopanned on a fibroblast cDNA phage-display library constructed in the vector pJuFo. Following three rounds of biopanning, recovered cDNAs were sequenced and then identified using BLAST comparisons with international databases. Both previously reported SLE autoantigens, for example, alpha-enolase and U1 small nuclear ribonucleoprotein-C (U1snRNP-C), and novel autoantibody targets, including ribosomal protein S20 (RPS20), ribosomal protein S13 (RPS13), ubiquitin-like protein PIC1 (PIC1), and transcription factor-like protein MRG15 (MRG15), were recovered from the biopanning procedure. Radiobinding assays were used subsequently to confirm the reactivity of some putative autoantigens to panels of sera from SLE patients, control patient groups, and healthy individuals. SLE patient sera were positive for reactivity to: U1snRNP-C, 4/15 (27%); alpha-enolase, 1/15 (7%); RPS20, 3/15 (20%); RPS13, 1/15 (7%); PIC1, 1/15 (7%); and MRG15, 2/15 (13%). Overall, cDNA phage-display technology appears to be applicable to the identification of autoantigens in autoimmune disease.
Kemp
E Helen
EH
Division of Clinical Sciences (North), University of Sheffield, S5 7AU, Sheffield, United Kingdom. e.h.kemp@sheffield.ac.uk
Herd
Laura M
LM
Waterman
Elizabeth A
EA
Wilson
Anthony G
AG
Weetman
Anthony P
AP
Watson
Philip P
PP
D000328:Adult; D001323:Autoantibodies / Q000276:immunology; D001324:Autoantigens / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D007074:Immunoglobulin G / Q000276:immunology; D008180:Lupus Erythematosus, Systemic / Q000276:immunology*; D008297:Male; D019151:Peptide Library; D010455:Peptides / Q000276:immunology; D011863:Radioimmunoassay
D001323:Autoantibodies; D001324:Autoantigens; D018076:DNA, Complementary; D007074:Immunoglobulin G; D019151:Peptide Library; D010455:Peptides