The cloned thrombin receptor is necessary and sufficient for activation of mitogen-activated protein kinase and mitogenesis in mouse lung fibroblasts - Loss of responses in fibroblasts from receptor knockout mice

被引:115
作者
Trejo, J
Connolly, AJ
Coughlin, SR
机构
[1] UNIV CALIF SAN FRANCISCO,DAIICHI RES CTR,INST CARDIOVASC RES,SAN FRANCISCO,CA 94143
[2] UNIV CALIF SAN FRANCISCO,DEPT MED,SAN FRANCISCO,CA 94143
[3] UNIV CALIF SAN FRANCISCO,DEPT PATHOL,SAN FRANCISCO,CA 94143
关键词
D O I
10.1074/jbc.271.35.21536
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
The mitogenic activity of thrombin on fibroblasts and smooth muscle cells may contribute to embryonic development and normal wound healing, and it may also play a role in pathological responses to vascular injury. To examine the importance of thrombin signaling in vivo and to define the cloned thrombin receptor's role, we disrupted the thrombin receptor gene (tr) in mice. Platelets from tr(-/-) mice responded normally to thrombin, but tr(-/-) fibroblasts showed no thrombin-induced calcium mobilization or phosphoinositide hydrolysis. Thus distinct thrombin receptors act in different tissues. This study focuses on the role of the thrombin receptor in thrombin-induced mitogenesis and mitogen-activated protein (MAP) kinase activation in mesenchymal cells. Thrombin and thrombin receptor agonist peptide both stimulated DNA synthesis and MAP kinase activation in fibroblasts derived from wild-type mice. These responses were selectively lost in fibroblasts from tr(-/-) mice. Activation of the cloned thrombin receptor is therefore necessary and sufficient for thrombin-induced mitogenesis and MAP kinase activation in mouse lung fibroblasts. The tr(-/-) mouse thus provides a valuable model for defining the role of thrombin-induced proliferative events in vivo. Because thrombin-induced MAP kinase activation was attributable to a single receptor expressed at natural levels, mouse lung fibroblasts presented an opportunity to define the pathways that normally mediate activation of MAP kinase by the thrombin receptor. Elimination of phorbol-sensitive protein kinase C by prolonged exposure to phorbol ester only partially inhibited MAP kinase activation by thrombin but completely blocked c-Raf kinase activation. Pertussis toxin partially inhibited MAP kinase activation by thrombin but had no significant effect on c-Raf kinase activation. Thus in mouse lung fibroblasts, one thrombin receptor utilizes two pathways for MAP kinase activation: one is protein kinase C- and c-Raf dependent, and a second is G(i)-dependent and c-Raf-independent.
引用
收藏
页码:21536 / 21541
页数:6
相关论文
共 35 条
[1]   DEVELOPMENT OF 3T3-LIKE LINES FROM BALB/C MOUSE EMBRYO CULTURES - TRANSFORMATION SUSCEPTIBILITY TO SV40 [J].
AARONSON, SA ;
TODARO, GJ .
JOURNAL OF CELLULAR PHYSIOLOGY, 1968, 72 (2P1) :141-&
[2]  
ALBLAS J, 1993, J BIOL CHEM, V268, P22235
[3]  
BAFFY G, 1994, J BIOL CHEM, V269, P8483
[4]   DIVERSITY IN FUNCTION AND REGULATION OF MAP KINASE PATHWAYS [J].
BLUMER, KJ ;
JOHNSON, GL .
TRENDS IN BIOCHEMICAL SCIENCES, 1994, 19 (06) :236-240
[5]   HOW MAP KINASES ARE REGULATED [J].
COBB, MH ;
GOLDSMITH, EJ .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1995, 270 (25) :14843-14846
[6]   THROMBIN RECEPTOR ACTIVATING PEPTIDES - IMPORTANCE OF THE N-TERMINAL SERINE AND ITS IONIZATION STATE AS JUDGED BY PH-DEPENDENCE, NUCLEAR-MAGNETIC-RESONANCE SPECTROSCOPY, AND CLEAVAGE BY AMINOPEPTIDASE-M [J].
COLLER, BS ;
WARD, P ;
CERUSO, M ;
SCUDDER, LE ;
SPRINGER, K ;
KUTOK, J ;
PRESTWICH, GD .
BIOCHEMISTRY, 1992, 31 (47) :11713-11720
[7]   Role of the thrombin receptor In development and evidence for a second receptor [J].
Connolly, AJ ;
Ishihara, H ;
Kahn, ML ;
Farese, RV ;
Coughlin, SR .
NATURE, 1996, 381 (6582) :516-519
[8]  
CRESPO P, 1994, J BIOL CHEM, V269, P21103
[9]   RAS-DEPENDENT ACTIVATION OF MAP KINASE PATHWAY MEDIATED BY G-PROTEIN BETA-GAMMA-SUBUNITS [J].
CRESPO, P ;
XU, NZ ;
SIMONDS, WF ;
GUTKIND, JS .
NATURE, 1994, 369 (6479) :418-420
[10]  
DAVIS RJ, 1993, J BIOL CHEM, V268, P14553