Recombinant IL-6 activates p42 and p44 mitogen-activated protein kinases in the IL-6 responsive B cell line, AF-10.
IL-6 is a multi-functional cytokine that utilizes 80-kDa ligand-binding and 130-kDa signal-transducing subunits to stimulate diverse cellular responses. Although IL-6R ligation has been associated with tyrosine protein phosphorylation and activation of an unidentified serine/threonine kinase, very little is known about the intermediary signaling events between the cell membrane and the nucleus. rIL-6 treatment of the human B cell line, AF-10, induced MAP kinase (mitogen-activated protein kinase) activity as determined by in vitro phosphorylation of microtubule-associated protein-2 (MAP-2) and the synthetic peptide APRTPGGRR, corresponding to amino acids 95-98 of bovine myelin basic protein. The kinetics of the response was rapid and dependent on the dose of rIL-6. The response was cytokine specific, did not require the presence of extracellular Ca2+, and was minimally affected by the presence of staurosporine. MAP kinase activation in AF-10 cells occurred in parallel with appearance of 42- and 44-kDa tyrosine phosphoproteins (p42 and p44). Moreover, MAP kinase activation was diminished when AF-10 cells were stimulated with rIL-6 in the presence of tyrosine protein kinase inhibitors, genistein and geldanomycin. p42 and p44 co-electrophoresed on SDS-PAGE with extracellular signal-related kinase (ERK)-2, and ERK-1, respectively; both are members of the ERK family. In addition to p42MAPK and p44MAPK, rIL-6 also activated a MAP-2 kinase that eluted at a lower salt concentration (20 to 60 mM NaCl, peak I) from Mono-Q resin than p42MAPK (120 to 180 mM NaCl, peak II). The identify of this kinase is unknown but it is not an MPB kinase or a protein that exhibits immunoreactivity with anti-ERK antisera. In another IL-6-responsive B cell line, SKW6.4, rIL-6-activated peak I MAP-2 kinase but failed to activate ERK-2. The protein kinase C agonist, PMA, did, however, activate ERK-2 in SKW6.4 cells. These results show that the pleiotrophic cytokine, IL-6, activates p42MAPK/ERK-2 and at least one other serine/threonine kinase in B cell lines.
Department of Medicine, UCLA School of Medicine, Los Angeles, CA 90024.
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Daeipour
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Kumar
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Amaral
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D000595:Amino Acid Sequence; D001402:B-Lymphocytes / Q000187:drug effects / Q000201:enzymology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002460:Cell Line; D002852:Chromatography, Ion Exchange; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D015850:Interleukin-6 / Q000494:pharmacology*; D008213:Lymphocyte Activation / Q000187:drug effects*; D019950:Mitogen-Activated Protein Kinase 1; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000096:biosynthesis; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000187:drug effects* / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000187:drug effects* / Q000378:metabolism; D011971:Receptors, Immunologic / Q000502:physiology; D019947:Receptors, Interleukin-6; D011994:Recombinant Proteins / Q000494:pharmacology
D015850:Interleukin-6; D010750:Phosphoproteins; D011971:Receptors, Immunologic; D019947:Receptors, Interleukin-6; D011994:Recombinant Proteins; D011494:Protein Kinases; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1