Tyrosine phosphorylation is an essential event in the stimulation of B lymphocytes by Staphylococcus aureus Cowan I.
Staphylococcus aureus Cowan I (SAC) is a potent mitogen for purified human B cells. By using Western blotting with antiphosphotyrosine antibodies, we demonstrated that the mitogenic effect of SAC is associated with rapid tyrosine phosphorylation of proteins of 45, 68, 75, 97, and 145 kDa. This tyrosine phosphorylation was detected within 30 s of the addition of SAC; it reached a maximum within 10 min, after which it declined gradually. In contrast to SAC, most soluble anti-IgM antibodies do not induce proliferation of isolated human B cells. As indicated by Western blotting, soluble anti-IgM antibodies induced a similar pattern of tyrosine phosphorylation, with the exception of the 68-kDa protein, which was the most heavily phosphorylated protein in SAC-treated cells. A similar but less intense 68-kDa band was also induced by mitogenic anti-IgM bound to beads. This suggested that tyrosine phosphorylation, especially of p68, may play an important role in B cell mitogenesis. To test this hypothesis, we determined the effect of specific tyrosine kinase inhibitors (tyrphostins) on SAC-induced tyrosine phosphorylation, oncogene expression, and B cell proliferation. The concentration dependencies of inhibition of these processes suggested that they were linked. Nonspecific toxic effects of the tyrphostins were ruled out by the demonstration that the tyrphostins did not alter cell viability and did not inhibit B cell proliferation induced by phorbol esters, which do not induce tyrosine phosphorylation. For maximal inhibition of SAC-induced cell proliferation, the tyrophostins needed to be added before or shortly after addition of SAC. Taken together, these data indicate that tyrosine phosphorylation is an obligatory early signal in B cell proliferation.
Roifman
C M
CM
Division of Immunology and Allergy, Hospital for Sick Children, Toronto, Ontario, Canada.
Chin
K
K
Gazit
A
A
Mills
G B
GB
Gilon
C
C
Levitzki
A
A
D000888:Antibodies, Anti-Idiotypic / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology*; D006801:Humans; D007075:Immunoglobulin M / Q000276:immunology; D066298:In Vitro Techniques; D008213:Lymphocyte Activation / Q000187:drug effects; D008970:Molecular Weight; D010750:Phosphoproteins / Q000378:metabolism*; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D016760:Proto-Oncogene Proteins c-fos; D013211:Staphylococcus aureus / Q000276:immunology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism
D000888:Antibodies, Anti-Idiotypic; D007075:Immunoglobulin M; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D016760:Proto-Oncogene Proteins c-fos; D019000:Phosphotyrosine; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D013755:Tetradecanoylphorbol Acetate