Induction of the antigen receptor expression on B lymphocytes results in rapid competence for signaling of SLP-65 and Syk.
The binding of antigen to the B cell antigen receptor (BCR) results in the activation of protein tyrosine kinases (PTKs) such as Lyn and Syk, and the phosphorylation of several substrate proteins including HS1 and SLP-65. How these signaling elements are connected to the BCR is not well understood. Using an expression vector for a tamoxifen-regulated Cre recombinase, we have developed a method that allows the inducible expression of the BCR. Disruption of the VH leader reading frame of the immunoglobulin heavy chain by two loxP sites is overcome by Cre-mediated DNA recombination and results in the cell surface expression of the BCR starting 4 h after exposure of transfected B cells to tamoxifen. This method can, in principle, be employed for the inducible expression of any secreted or type I transmembrane protein. By monitoring the activation of signaling elements in pervanadate-stimulated B cells expressing different levels of the BCR, we show here that phosphorylation of SLP-65 and Syk, but not of Lyn, is strictly dependent on the expression of the BCR on the cell surface. These data suggest that the BCR reorganizes its signaling molecules as soon as it appears on the cell surface.
Zhang
Y
Y
Department of Molecular Immunobiology, Biology III, University Freiburg and Max-Planck-Institute for Immunobiology, Stübeweg 51, 79108 Freiburg, Germany.
Wienands
J
J
Zürn
C
C
Reth
M
M
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D001402:B-Lymphocytes / Q000187:drug effects; D001483:Base Sequence; D001692:Biological Transport; D002352:Carrier Proteins; D002460:Cell Line; D004792:Enzyme Precursors / Q000378:metabolism*; D005786:Gene Expression Regulation; D019426:Integrases; D047908:Intracellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011947:Receptors, Antigen, B-Cell / Q000096:biosynthesis*; D011995:Recombination, Genetic; D015398:Signal Transduction; D000072377:Syk Kinase; D013629:Tamoxifen / Q000494:pharmacology; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D014638:Vanadates / Q000494:pharmacology; D014764:Viral Proteins; D019061:src-Family Kinases / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; C113939:B cell linker protein; D002352:Carrier Proteins; D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D010750:Phosphoproteins; D011947:Receptors, Antigen, B-Cell; D014764:Viral Proteins; C076715:pervanadate; D013629:Tamoxifen; D014638:Vanadates; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D000072377:Syk Kinase; D019061:src-Family Kinases; C045073:Cre recombinase; D019426:Integrases