TY - JOUR
T1 - Proteinase-activated receptor-2-mediated activation of stress-activated protein kinases and inhibitory kappa b kinases in NCTC 2544 keratinocytes
AU - Kanke, T.
AU - MacFarlane, S.R.
AU - Seatter, M.J.
AU - Davenport, E.L.
AU - Paul, A.
AU - McKenzie, R.C.
AU - Plevin, R.J.
PY - 2001
Y1 - 2001
N2 - In this study we examined the regulation of the stress-activated protein (SAP) kinases and inhibitory κB kinases (IKKs) through stimulation of the novel G-protein-coupled receptor proteinase-activated receptor-2 in the human keratinocyte cell line NCTC2544. Trypsin and the peptide SLIGKV stimulated a time-dependent increase in both c-Jun N-terminal kinase and p38 mitogen-activated protein kinase activity. Trypsin also stimulated NFκB-DNA binding and the activation of the upstream kinases IKKα and -β. Phorbol 12-myristate 13-acetate also strongly activated both SAP kinases and IKK isoforms, suggesting the potential for a protein kinase C-mediated regulatory mechanism underlying the effects of trypsin. Pre-incubation with selective protein kinase C (PKC) inhibitors GF109203X and Gö6983, or transfection of dominant negative (DN)-PKCα, abolished phorbol 12-myristate 13-acetate-mediated c-Jun N-terminal kinase activity, although it only partially inhibited the response to trypsin. In contrast, Gö6983 reduced trypsin-stimulated p38 mitogen-activated protein kinase activity to a greater extent than GF109203X, although DN-PKCα or PKCζ had no substantial effect. Additionally, inhibitors of PKC partially reduced trypsin-stimulated IKKα activity but abolished that of IKKβ, whereas DN-PKCα but not DN-PKCζ substantially reduced trypsin-stimulated Flag-IKKβ activity. This study shows for the first time proteinase-activated receptor-2-mediated stimulation of both SAP kinase and IKK signaling and differing roles for PKC isoforms in the regulation of each pathway.
AB - In this study we examined the regulation of the stress-activated protein (SAP) kinases and inhibitory κB kinases (IKKs) through stimulation of the novel G-protein-coupled receptor proteinase-activated receptor-2 in the human keratinocyte cell line NCTC2544. Trypsin and the peptide SLIGKV stimulated a time-dependent increase in both c-Jun N-terminal kinase and p38 mitogen-activated protein kinase activity. Trypsin also stimulated NFκB-DNA binding and the activation of the upstream kinases IKKα and -β. Phorbol 12-myristate 13-acetate also strongly activated both SAP kinases and IKK isoforms, suggesting the potential for a protein kinase C-mediated regulatory mechanism underlying the effects of trypsin. Pre-incubation with selective protein kinase C (PKC) inhibitors GF109203X and Gö6983, or transfection of dominant negative (DN)-PKCα, abolished phorbol 12-myristate 13-acetate-mediated c-Jun N-terminal kinase activity, although it only partially inhibited the response to trypsin. In contrast, Gö6983 reduced trypsin-stimulated p38 mitogen-activated protein kinase activity to a greater extent than GF109203X, although DN-PKCα or PKCζ had no substantial effect. Additionally, inhibitors of PKC partially reduced trypsin-stimulated IKKα activity but abolished that of IKKβ, whereas DN-PKCα but not DN-PKCζ substantially reduced trypsin-stimulated Flag-IKKβ activity. This study shows for the first time proteinase-activated receptor-2-mediated stimulation of both SAP kinase and IKK signaling and differing roles for PKC isoforms in the regulation of each pathway.
KW - stress-activated protein
KW - protein kinases
KW - kappa b kinases
KW - keratinocytes
U2 - 10.1074/jbc.M100377200
DO - 10.1074/jbc.M100377200
M3 - Article
VL - 276
SP - 31657
EP - 31666
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 34
ER -