Schramek H, Schumacher M, Pfaller W
Department of Physiology, University of Innsbruck, Austria.
Am J Physiol. 1996 Aug;271(2 Pt 2):F423-32. doi: 10.1152/ajprenal.1996.271.2.F423.
The duration of extracellular signal-regulated protein kinase (ERK) activation is critical for cell signaling decisions and probably determines whether a stimulus elicits proliferation or differentiation. We studied the intracellular signals regulating sustained ERK-2 activity in glomerular mesangial cells (GMC), utilizing combination of GMC mitogens of different potency. Incubation of GMC with both endothelin-1 (ET-1) and platelet-derived growth factor BB (PDGF-BB) led to a long-lasting, monophasic increase in ERK-2 activity. In contrast, when ET-1 was administered together with epidermal growth factor (EGF), a less pronounced and shorter activation occurred. Long-term stimulation of ERK-2 was accompanied by an increase in p45 MEK activity, which again was more pronounced when ET-1 was administered together with PDGF-BB compared with EGF. In the presence of actinomycin D (Act D), an inhibitor of RNA synthesis, ERK-2 activity induced by ET-1 and PDGF-BB but not by ET-1 and EGF remained elevated more than sixfold throughout the whole incubation period of 6 h. The effect of Act D on ET-1- and PDGF-BB-induced ERK-2 activation was mimicked by the protein phosphatase inhibitor sodium orthovanadate. In addition, vanadate also unmarked an ET-1- and EGF-induced ERK-2 activity after 6 h. The serine/threonine phosphatase inhibitor okadaic acid (OA) did neither alter agonist-induced ERK-2 activity after 6 h (0.5 nM OA) nor after 10 min or 1 h (250 nM). Together these results suggest that, in GMC, long-term activation of the mitogen-activated protein kinase ERK-2 is differentially regulated, depending on the combination of agonists administered. ET-1- and PDGF-BB-induced long-term activation of ERK-2 is regulated by a vanadate-sensitive protein phosphatase(s) and by a transcriptionally regulated protein(s). In contrast, ET-1- and EGF-induced sustained ERK-2 stimulation is regulated by a vanadate-sensitive protein phosphatase(s) but not by a transcriptionally regulated protein. Agonist-specific and time-dependent stimulation of ERK-2-regulating protein phosphatases may be critical for the length of ERK-2 activation in GMC and could thus be of pathophysiological significance in glomerular diseases associated with alterations in cell proliferation or cell differentiation.
细胞外信号调节蛋白激酶（ERK）激活的持续时间对于细胞信号转导决策至关重要，可能决定了一种刺激是引发增殖还是分化。我们利用不同效力的肾小球系膜细胞（GMC）有丝分裂原组合，研究了调节GMC中ERK-2持续活性的细胞内信号。将GMC与内皮素-1（ET-1）和血小板衍生生长因子BB（PDGF-BB）一起孵育，导致ERK-2活性出现持久的单相增加。相比之下，当ET-1与表皮生长因子（EGF）一起给药时，激活作用较弱且持续时间较短。ERK-2的长期刺激伴随着p45 MEK活性的增加，与EGF相比，当ET-1与PDGF-BB一起给药时，这种增加更为明显。在存在RNA合成抑制剂放线菌素D（Act D）的情况下，ET-1和PDGF-BB诱导的ERK-2活性在整个6小时的孵育期内保持升高超过六倍，而ET-1和EGF诱导的ERK-2活性则没有。蛋白磷酸酶抑制剂原钒酸钠模拟了Act D对ET-1和PDGF-BB诱导的ERK-2激活的作用。此外，钒酸盐在6小时后也恢复了ET-1和EGF诱导的ERK-2活性。丝氨酸/苏氨酸磷酸酶抑制剂冈田酸（OA）在6小时（0.5 nM OA）以及10分钟或1小时（250 nM）后均未改变激动剂诱导的ERK-2活性。这些结果共同表明，在GMC中，有丝分裂原激活的蛋白激酶ERK-2的长期激活受到不同的调节，这取决于所给予激动剂的组合。ET-1和PDGF-BB诱导的ERK-2长期激活受钒酸盐敏感的蛋白磷酸酶和转录调节蛋白调控。相比之下，ET-1和EGF诱导的ERK-2持续刺激受钒酸盐敏感的蛋白磷酸酶调控，但不受转录调节蛋白调控。ERK-2调节蛋白磷酸酶的激动剂特异性和时间依赖性刺激可能对GMC中ERK-2激活的持续时间至关重要，因此在与细胞增殖或细胞分化改变相关的肾小球疾病中可能具有病理生理学意义。
