A novel approach to identify antigens recognized by CD4 T cells using complement-opsonized bacteria expressing a cDNA library.
In patients with hematological malignancies receiving HLA-matched stem cell transplantation, T cells specific for minor histocompatibility antigens play a major role in graft rejection, induction of graft-versus-host disease and beneficial graft-versus-leukemia reactivity. Several human minor histocompatibility antigens recognized by T cells have been identified, but only two are presented by HLA class II molecules. In search of an efficient approach to identify antigenic peptides processed through the HLA class II pathway, we constructed a cDNA library in bacteria that were induced to express proteins. Bacteria were opsonized with complement to enforce receptor-mediated uptake by Epstein-Barr virus immortalized B cells that were subsequently used as antigen-presenting cells. This approach was validated with an HLA class II-restricted antigen encoded by gene DBY. We were able to identify bacteria expressing DBY diluted into a 300-fold excess of bacteria expressing a nonrelevant gene. Screening of a bacterial library using a DBY-specific CD4 T cell clone resulted in the isolation of several DBY cDNAs. We propose this strategy for a rapid identification of HLA class II-restricted antigenic peptides recognized by CD4 T cells.
van de Corput
L
L
Department of Hematology, Leiden University Medical Center, Leiden, The Netherlands.
Chaux
P
P
van der Meijden
E D
ED
De Plaen
E
E
Frederik Falkenburg
J H
JH
van der Bruggen
P
P
D001419:Bacteria / Q000235:genetics*; D001483:Base Sequence; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D003001:Cloning, Molecular / Q000379:methods; D003165:Complement System Proteins; D053487:DEAD-box RNA Helicases; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics*; D015723:Gene Library; D006801:Humans; D015778:Minor Histocompatibility Antigens / Q000097:blood; D011506:Proteins / Q000235:genetics
D017931:DNA Primers; D018076:DNA, Complementary; D015778:Minor Histocompatibility Antigens; D011506:Proteins; D003165:Complement System Proteins; C492034:DDX3Y protein, human; D053487:DEAD-box RNA Helicases