Ro 60 functions as a receptor for beta(2)-glycoprotein I on apoptotic cells.
OBJECTIVE
The autoantigens 60-kd Ro/SSA (Ro 60) and beta(2)-glycoprotein I (beta(2)GPI) are both displayed on the surface membrane of apoptotic cells. Epitope-spreading experiments have suggested that these autoantigens may be present as a complex on the apoptotic cell surface. This study was undertaken to investigate whether beta(2)GPI interacts with Ro 60 on apoptotic cells and alters the binding of anti-Ro 60 IgG.


METHODS
The interaction between soluble recombinant Ro 60 fragments and beta(2)GPI was investigated in vitro by direct and saturation binding assays using native human beta(2)GPI and recombinant domain deletion mutants. Binding of beta(2)GPI to early and late apoptotic cells was assessed by multiparameter flow cytometry, and specificity of binding was determined by competitive inhibition with soluble recombinant Ro 60 and anti-Ro 60 IgG.


RESULTS
The Ro 60 fragment expressing a surface-exposed epitope (apotope) bound with high affinity (K(d) = approximately 15 nM) to domain V of beta(2)GPI in vitro. Beta(2)-glycoprotein I bound to the surface of apoptotic cells in a dose-dependent manner and was blocked by the Ro 60 apotope fragment. In reciprocal competitive inhibition studies, beta(2)GPI blocked the binding of anti-Ro 60 autoantibodies to apoptotic cells in a dose-dependent manner, and anti-Ro 60 IgG inhibited the binding of beta(2)GPI. Moreover, beta(2)GPI showed a 2-fold increase in binding to apoptotic cells that overexpress Ro 60 on the surface.


CONCLUSIONS
These results demonstrate that Ro 60 functions as a novel receptor for beta(2)GPI on the surface of apoptotic cells. The formation of Ro 60-beta(2)GPI complexes may protect against anti-Ro 60 autoantibody-mediated tissue injury.
Reed
Joanne H
JH
Flinders University of South Australia, Adelaide, and Flinders Medical Centre, Bedford Park, South Australia, Australia.
Giannakopoulos
Bill
B
Jackson
Michael W
MW
Krilis
Steven A
SA
Gordon
Tom P
TP
D000936:Antigen-Antibody Complex / Q000276:immunology; D017209:Apoptosis / Q000276:immunology / Q000502:physiology*; D001323:Autoantibodies / Q000276:immunology / Q000378:metabolism; D001324:Autoantigens / Q000276:immunology / Q000378:metabolism*; D002460:Cell Line; D004847:Epithelial Cells / Q000378:metabolism* / Q000473:pathology*; D000939:Epitopes / Q000276:immunology / Q000378:metabolism; D006801:Humans; D019169:Jurkat Cells; D011485:Protein Binding / Q000276:immunology / Q000502:physiology; D020733:RNA, Small Cytoplasmic / Q000276:immunology / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000276:immunology / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000276:immunology / Q000378:metabolism*; D053482:beta 2-Glycoprotein I / Q000276:immunology / Q000378:metabolism*; D000099207:SS-A Antigen
D000936:Antigen-Antibody Complex; D001323:Autoantibodies; D001324:Autoantigens; D000939:Epitopes; D020733:RNA, Small Cytoplasmic; D011956:Receptors, Cell Surface; D012261:Ribonucleoproteins; D053482:beta 2-Glycoprotein I; C116006:RO60 protein, human; D000099207:SS-A Antigen