title: The monoclonal antibody 6B9 recognizes CD44 and not cell surface transglutaminase 2.
abstract: The multifunctional enzyme transglutaminase 2 (TG2) can be located intracellularly, in the extracellular matrix (ECM) and on the cell surface. Cell surface TG2 (csTG2) is poorly recognized both by most TG2-specific commercial antibodies and celiac disease-associated anti-TG2 autoantibodies. The recent characterization of a csTG2-specific monoclonal antibody (mAb), which did not recognize ECM-associated TG2, suggested major conformational differences between csTG2 and TG2 found in the ECM. Subsequent findings based on this antibody indicated ubiquitous abundance and novel roles of csTG2 in innate immune responses. We wished to identify the epitope of 6B9 so as to shed light on the disparate antibody binding properties of csTG2- and ECM-associated TG2. Surprisingly, and despite thorough effort, we were unable to isolate TG2 as the antigen of 6B9. We found that 6B9 does not react with recombinant human TG2. In immunoprecipitation experiments, 6B9 pulled down an 85 kDa protein which was identified as CD44 by mass spectrometry. Several flow cytometry experiments including the testing of CD44s transfectants indicated that CD44, and not csTG2, is the antigen of 6B9. We conclude that 6B9 does not recognize csTG2 but rather the cell surface glycoprotein CD44. Thus, recent knowledge of csTG2 gained through the use of 6B9 should be reevaluated.
authors.affiliation: Centre for Immune Regulation, Institute of Immunology, University of Oslo and Rikshospitalet University Hospital, Oslo, Norway. Jorunn.Stamnas@rr-research.no
authors.forename: J
authors.initials: J
authors.lastname: Stamnaes
authors.forename: B
authors.initials: B
authors.lastname: Fleckenstein
authors.forename: F
authors.initials: F
authors.lastname: Lund-Johansen
authors.forename: L M
authors.initials: LM
authors.lastname: Sollid
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015153:Blotting, Western; D001853:Bone Marrow / Q000276:immunology; D000939:Epitopes / Q000276:immunology; D019204:GTP-Binding Proteins / Q000276:immunology*; D006801:Humans; D018960:Hyaluronan Receptors / Q000276:immunology*; D047468:Immunoprecipitation; D019169:Jurkat Cells; D008969:Molecular Sequence Data; D011487:Protein Conformation; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014162:Transfection; D011503:Transglutaminases / Q000276:immunology*; D020298:U937 Cells
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D018960:Hyaluronan Receptors; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; D019204:GTP-Binding Proteins