Trypsin stimulates proteinase-activated receptor-2-dependent and -independent activation of mitogen-activated protein kinases

被引:87
作者
Belham, CL
Tate, RJ
Scott, PH
Pemberton, AD
Miller, HRP
Wadsworth, RM
Gould, GW
Plevin, R
机构
[1] UNIV STRATHCLYDE,ROYAL COLL,MOL BIOL LAB,GLASGOW G1 1XW,LANARK,SCOTLAND
[2] UNIV EDINBURGH,ROYAL SCH VET STUDIES,DEPT VET CLIN STUDIES,VET FIELD STN,ROSLIN EH25 9RG,MIDLOTHIAN,SCOTLAND
[3] UNIV GLASGOW,INST BIOMED & LIFE SCI,DIV BIOCHEM & MOL BIOL,GLASGOW G12 8QQ,LANARK,SCOTLAND
关键词
D O I
10.1042/bj3200939
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We have examined protease-mediated activation of the mitogen-activated protein (MAP) kinase cascade in rat aortic smooth-muscle cells and bovine pulmonary arterial fibroblasts. Exposure of smooth-muscle cells to trypsin evoked rapid and transient activation of c-Raf-1, MAP kinase kinase 1 and 2 and MAP kinase that was sensitive to inhibition by soybean trypsin inhibitor. The actions of-trypsin were closely mimicked by the proteinase-activated receptor 2 (PAR-2)-activating peptide sequence SLIGRL but not LSIGRL. Peak MAP kinase activation in response to both trypsin and SLIGRL was also dependent on concentration, with EC(50) values of 12.1+/-3.4 nM and 62.5+/-4.5 mu M respectively. Under conditions where MAP kinase activation by SLIGRL was completely desensitized by prior exposure of smooth-muscle cells to the peptide, trypsin-stimulated MAP kinase activity was markedly attenuated (78.9+/-15.1% desensitization), whereas the response to thrombin was only marginally affected (16.6+/-12.1% desensitization). Trypsin and SLIGRL also weakly stimulated the activation of the MAP kinase homologue p38 in smooth-muscle cells without any detectable activation of c-Jun N-terminal kinase. Strong activation of the MAP kinase cascade and modest activation of p38 by trypsin were also observed in fibroblasts, although in this cell type these effects were not mimicked by SLIGRL nor by the thrombin receptor-activating peptide SFLLRNPNDKYEPF. Reverse transcriptase-PCR analysis confirmed the presence of PAR-2 mRNA in smooth-muscle cells but not fibroblasts. Our results suggest that in vascular smooth-muscle cells, trypsin stimulates the activation of the MAP kinase cascade relatively selectively, in a manner consistent with an interaction with the recently described PAR-2. Activation of MAP kinase by trypsin in vascular fibroblasts, however, seems to be independent of PAR-2 and occurs by an undefined mechanism possibly involving novel receptor species.
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页码:939 / 946
页数:8
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