Polyclonal B cell activation by a B cell differentiation factor, B151-TRF2. II. Evidence for interaction of B151-TRF2 with glycoprotein on B cell membrane via recognition of terminal N-acetyl-D-glucosamine residue(s).
We investigated the role of carbohydrates in the interaction of a B cell differentiation factor designated as B151-TRF2 derived from B151K12 T cell hybridoma with the corresponding receptor on B cells. Induction of polyclonal differentiation of unprimed B cells into IgM-secreting cells by B151-TRF2 was specifically inhibited by addition of N-acetyl-D-glucosamine (GlcNAc) but not by structurally unrelated monosaccharides such as D-galactose, D-glucose, and N-acetyl-D-galactosamine (GalNAc). Absorption of B151-TRF2 activity with spleen cells was specifically inhibited by the presence of GlcNAc. These results indicate that GlcNAc residues are involved in the interaction of B151-TRF2 with the receptor on B cells. To gain insight into mechanism by which GlcNAc inhibits B151-TRF2-mediated B cell responses, the existence of GlcNAc residues was examined on the B151-TRF2 molecule and the corresponding receptor on the B cell surface. The results revealed that B151-TRF2 molecule was not bound to various lectin-coupled agarose beads so far tested, suggesting absence of carbohydrate moieties on the B151-TRF2 molecule. By contrast, pretreatment of spleen cells with trypsin or glycosidase mixture abolished their ability to absorb B151-TRF2 activity. Moreover, B151-TRF2-absorbing ability of spleen cells disappeared by the pretreatment with beta-N-acetylglucosaminidase, which cleaves terminal GlcNAc. The fact that pnitrophenyl (PNP)-GlcNAc specifically inhibited such enzyme activity on target cells indicates that terminal GlcNAc on the B cell surface plays a crucial role in the interaction with B151-TRF2 molecule. Interestingly, it was also found that B151-TRF2 activity was trapped and eluted from GlcNAc-coupled agarose beads. Taken collectively, these results strongly suggest that B cell membrane receptors for B151-TRF2 comprise glycoproteins with a terminal GlcNAc residue(s), and that binding of B151-TRF2 with terminal GlcNAc on the receptor is important for the subsequent activation of B cells.
D000117:Acetylglucosamine / Q000502:physiology*; D000118:Acetylglucosaminidase / Q000378:metabolism; D000818:Animals; D000917:Antibody Formation; D001402:B-Lymphocytes / Q000166:cytology / Q000276:immunology*; D001665:Binding Sites; D002454:Cell Differentiation; D005944:Glucosamine / Q000031:analogs & derivatives*; D006133:Growth Substances / Q000378:metabolism*; D015847:Interleukin-4; D037102:Lectins / Q000378:metabolism; D008213:Lymphocyte Activation; D008222:Lymphokines / Q000378:metabolism* / Q000502:physiology*; D051379:Mice; D011971:Receptors, Immunologic / Q000502:physiology; D013154:Spleen / Q000166:cytology
D006133:Growth Substances; D037102:Lectins; D008222:Lymphokines; D011971:Receptors, Immunologic; D015847:Interleukin-4; D000118:Acetylglucosaminidase; D005944:Glucosamine; D000117:Acetylglucosamine