Initiation of antigen receptor endocytosis and B lymphocyte activation lie on independent biochemical pathways.
Peroxidase-conjugated anti-surface immunoglobulin (sIg) was used quantitatively to monitor endocytosis of crosslinked sIg on murine B lymphocytes. The role of biochemical second messengers in the initiation of endocytosis was assessed by employing several inhibitors. A novel peroxidase detection system was used and temperature-dependent decreases in sIg density on immunoperoxidase-labelled murine lymphocytes were monitored. Metabolic inhibitors as well as colchicine and cytochalasin D were utilized to confirm that the internalization of sIg could be blocked by classical inhibitors of the endocytosis process. The role of tyrosine kinase activity was established by the fact that endocytosis was significantly reduced with 100 micrograms/mL genistein. Experiments using EGTA or 1,2-bis(beta-aminophenoxy)ethane-N-N,N'-tetraacetic acid (BAPTA) to chelate Ca2+ indicated that Ca2+ plays little role in endocytosis. Likewise, protein kinase C (PKC) was not found to be involved in endocytosis, as activation of PKC with 50 ng/mL phorbol 12-myristate 13-acetate, or inhibition of the enzyme with 1 nmol/L or 5 nmol/L staurosporin, did not modulate endocytosis. Taken together, results suggested that ligand-induced endocytosis of antigen receptors is mediated primarily through localized membrane events and is not dependent upon the classical B lymphocyte activation signals, such as the biochemical events in the inositol phosphate cascade.
Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, PA 19107.
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Shuler
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Owen
D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D004705:Endocytosis / Q000276:immunology*; D005260:Female; D008213:Lymphocyte Activation / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009195:Peroxidase; D011493:Protein Kinase C / Q000276:immunology / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000276:immunology / Q000378:metabolism; D011946:Receptors, Antigen / Q000276:immunology* / Q000378:metabolism; D011947:Receptors, Antigen, B-Cell / Q000276:immunology; D015398:Signal Transduction / Q000276:immunology; D013154:Spleen / Q000276:immunology; D013696:Temperature
D011946:Receptors, Antigen; D011947:Receptors, Antigen, B-Cell; D009195:Peroxidase; D011505:Protein-Tyrosine Kinases; D011493:Protein Kinase C; D002118:Calcium