Selective activation of p42 mitogen-activated protein (MAP) kinase in murine B lymphoma cell lines by membrane immunoglobulin cross-linking. Evidence for protein kinase C-independent and -dependent mechanisms of activation.
Cross-linking of membrane immunoglobulin (mIg), the B lymphocyte antigen receptor, with anti-receptor antibodies stimulates tyrosine phosphorylation of a number of proteins, including one of 42 kDa. Proteins with a similar molecular mass are tyrosine-phosphorylated in response to receptor stimulation in other cell types and have been identified as serine/threonine kinases, termed mitogen-activated protein (MAP) kinases or extracellular signal-regulated kinases (ERKs). The MAP kinases constitute a family of related kinases, at least three of which have molecular masses of 40-45 kDa. In this paper we show that mIg cross-linking stimulated the myelin basic protein phosphotransferase activity characteristic of MAP kinase in both mature and immature murine B cell lines. This enzyme activity co-purified on three different columns with a 42 kDa protein that was tyrosine-phosphorylated (pp42) in response to mIg cross-linking and which reacted with a panel of anti-(MAP kinase) antibodies. Although immunoblotting with the anti-(MAP kinase) antibodies showed that these B cell lines expressed both 42 kDa and 44 kDa forms of MAP kinase, only the 42 kDa form was activated and tyrosine-phosphorylated to a significant extent. Activation of protein kinase C (PKC) with phorbol esters also resulted in selective tyrosine phosphorylation and activation of the 42 kDa MAP kinase. This suggested that mIg-induced MAP kinase activation could be due to stimulation of PKC by mIg. However, mIg-stimulated MAP kinase activation and pp42 tyrosine phosphorylation was only partially blocked by a PKC inhibitor, the staurosporine analogue Compound 3. In contrast, Compound 3 completely blocked the ability of phorbol esters to stimulate MAP kinase activity and induce tyrosine phosphorylation of pp42. Thus mIg may activate MAP kinase by both PKC-dependent and -independent mechanisms.
Gold
M R
MR
Biomedical Research Centre, University of British Columbia, Vancouver, Canada.
Sanghera
J S
JS
Stewart
J
J
Pelech
S L
SL
D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000201:enzymology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D004789:Enzyme Activation; D066298:In Vitro Techniques; D016393:Lymphoma, B-Cell; D051379:Mice; D008969:Molecular Sequence Data; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D019000:Phosphotyrosine; D011493:Protein Kinase C / Q000502:physiology*; D011506:Proteins / Q000378:metabolism*; D011940:Receptor Aggregation; D011947:Receptors, Antigen, B-Cell / Q000378:metabolism* / Q000502:physiology; D015398:Signal Transduction; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
D011506:Proteins; D011947:Receptors, Antigen, B-Cell; D019000:Phosphotyrosine; D015240:Phorbol 12,13-Dibutyrate; D014443:Tyrosine; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases