Identification of Cell Surface Straight Chain Poly-N-Acetyl-Lactosamine Bearing Protein Ligands for VH4-34-Encoded Natural IgM Antibodies.
B cell binding and cytotoxicity by human VH4-34-encoded Abs of the IgM isotype has been well documented. A VH4-34-IgM has recently shown a favorable early response in a phase 1 trial for treatment of B cell acute lymphoblastic leukemia. Although its B cell ligand has been identified as straight chain poly-N-acetyl-lactosamine (SC-PNAL), the carrier of the sugar moiety has not been identified. Using nanoelectrospray ionization mass spectrometry, we identify the metabolic activation related protein complex of CD147-CD98 as a major carrier of poly-N-acetyl-lactosamine (SC-PNAL) on human pre-B cell line Nalm-6. Previous studies have suggested CD45 as the SC-PNAL carrier for VH4-34-encoded IgG Abs. Because Nalm-6 is CD45 negative, human peripheral blood B lymphocytes and human B cell line, Reh, with high CD45 expression, were examined for SC-PNAL carrier proteins. Western blot analysis shows that the CD147-98 complex is indeed immunoprecipitated by VH4-34-encoded IgMs from human peripheral blood B lymphocytes and human B cell lines, Reh, OCI-Ly8, and Nalm-6. However, CD45 is immunoprecipitated only from peripheral B lymphocytes, but not from Reh despite the high expression of CD45. These results suggest that human B cells retain SC-PNAL on the CD147-98 complex, but modulate the sugar moiety on CD45. Because the carbohydrate moiety may act as a selecting Ag for VH4-34 autoantibody repertoire, its differential expression on proteins may provide a clue to the intricate atypical regulation of the VH4-34 gene.
Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, Stanford University, Stanford, CA 94305; and nbhat@stanford.edu.
Neelima M
http://orcid.org/0000-0002-6472-1638
NM
Bhat
Stanford University Mass Spectrometry, Stanford University, Stanford, CA 94305.
Christopher M
http://orcid.org/0000-0002-0210-3711
CM
Adams
Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, Stanford University, Stanford, CA 94305; and.
Yi
Y
Chen
Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, Stanford University, Stanford, CA 94305; and.
Marcia M
MM
Bieber
Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, Stanford University, Stanford, CA 94305; and.
Nelson N H
NN
Teng
D000911:Antibodies, Monoclonal / Q000276:immunology; D001323:Autoantibodies / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology*; D051926:Basigin / Q000276:immunology*; D002460:Cell Line; D027261:Fusion Regulatory Protein-1 / Q000276:immunology*; D005786:Gene Expression Regulation / Q000276:immunology; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007075:Immunoglobulin M / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000235:genetics; D047468:Immunoprecipitation; D017493:Leukocyte Common Antigens / Q000378:metabolism; D011134:Polysaccharides / Q000276:immunology*; D011947:Receptors, Antigen, B-Cell / Q000235:genetics / Q000276:immunology; D021241:Spectrometry, Mass, Electrospray Ionization
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; C497886:BSG protein, human; D027261:Fusion Regulatory Protein-1; D007143:Immunoglobulin Heavy Chains; D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region; D011134:Polysaccharides; D011947:Receptors, Antigen, B-Cell; D051926:Basigin; C037199:poly-N-acetyllactosamine; D017493:Leukocyte Common Antigens; C516769:PTPRC protein, human