Tyrosine phosphatase activity of lymphoma CD45 (GP180) is regulated by a direct interaction with the cytoskeleton.
GP180 is one of the major transmembrane glycoproteins in mouse T-lymphoma cells. This molecule is an isoform of CD45 and is known to contain an intrinsic protein tyrosine phosphatase (PTPase) activity. Using several complementary biochemical techniques, we have found that fodrin (a spectrin-like protein) is preferentially co-isolated with CD45 (GP180), suggesting that a complex between CD45 (GP180) and the cytoskeleton exists in mouse T-lymphoma cells. Furthermore, we have determined that this CD45 (GP180)-fodrin complex is dissociated by high salt treatment. Using in vitro binding assays, we have shown that CD45 (GP180) binds directly and specifically to fodrin (Kd approximately 1.1 nM) or spectrin (Kd approximately 3.2 nM) in a saturable manner. Additional analyses indicate that a 48-kDa phosphopeptide of CD45 (GP180) contains the fodrin/spectrin-binding domain. Most importantly, the direct binding of fodrin/spectrin to CD45 (GP180) is found to significantly stimulate the PTPase activity of CD45. Enzyme kinetic analysis indicates that fodrin and spectrin increase the Vmax of CD45 (GP180)-mediated dephosphorylation by 7.5 and 3.2-fold, respectively, without significantly changing the Km value. These results strongly suggest that the cytoskeletal proteins, fodrin and spectrin, play an important role in the regulation of the CD45 (GP180) PTPase activity during lymphocyte activation.
Lokeshwar
V B
VB
Department of Cell Biology and Anatomy, School of Medicine, University of Miami, Florida 33101.
Bourguignon
L Y
LY
D000818:Animals; D001345:Autoradiography; D002294:Carcinoma, Squamous Cell; D002352:Carrier Proteins / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D017493:Leukocyte Common Antigens / Q000378:metabolism*; D016399:Lymphoma, T-Cell; D051379:Mice; D008840:Microfilament Proteins / Q000378:metabolism; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D013049:Spectrin / Q000378:metabolism; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; D066246:ErbB Receptors; D017493:Leukocyte Common Antigens; D008840:Microfilament Proteins; D017027:Protein Tyrosine Phosphatases; D013049:Spectrin