Schäfer C, Ross S E, Bragado M J, Groblewski G E, Ernst S A, Williams J A
Department of Physiology and Internal Medicine, University of Michigan, Ann Arbor, Michigan 48109-0622, USA.
J Biol Chem. 1998 Sep 11;273(37):24173-80. doi: 10.1074/jbc.273.37.24173.
Cholecystokinin (CCK) and other pancreatic secretagogues have recently been shown to activate signaling kinase cascades in pancreatic acinar cells, leading to the activation of extracellular signal-regulated kinases and Jun N-terminal kinases. We now show the presence of a third kinase cascade activating p38 mitogen-activated protein (MAP) kinase in isolated rat pancreatic acini. CCK and osmotic stress induced by sorbitol activated p38 MAP kinase within minutes; their effects were dose-dependent, with maximal activation of 2.8- and 4.4-fold, respectively. The effects of carbachol and bombesin on p38 MAP kinase activity were similar to those of CCK, whereas phorbol ester, epidermal growth factor, and vasoactive intestinal polypeptide stimulated p38 MAP kinase by 2-fold or less. Both CCK and sorbitol also increased the tyrosyl phosphorylation of p38 MAP kinase. Using the specific inhibitor of p38 MAP kinase, SB 203580, we found that p38 MAP kinase activity was required for MAP kinase-activated protein kinase-2 activation in pancreatic acini. SB 203580 reduced the level of basal phosphorylation and blocked the increased phosphorylation of Hsp 27 after stimulation with either CCK or sorbitol. CCK treatment induced an initial rapid decrease in total F-actin content of acini, followed by an increase after 40 min. Preincubation with SB 203580 significantly inhibited these changes in F-actin content. Staining of the actin cytoskeleton with rhodamine-conjugated phalloidin and analysis by confocal fluorescence microscopy showed disruption of the actin cytoskeleton after 10 and 40 min of CCK stimulation. Pretreatment with SB 203580 reduced these changes. These findings demonstrate that the activation of p38 MAP kinase is involved not only in response to stress, but also in physiological signaling by gastrointestinal hormones such as CCK, where activation of Gq-coupled receptors stimulates a cascade in which p38 MAP kinase activates MAP kinase-activated protein kinase-2, resulting in Hsp 27 phosphorylation. Activation of p38 MAP kinase, most likely through phosphorylation of Hsp 27, plays a role in the organization of the actin cytoskeleton in pancreatic acini.
胆囊收缩素（CCK）及其他促胰液素最近已被证明可激活胰腺腺泡细胞中的信号激酶级联反应，从而导致细胞外信号调节激酶和Jun N端激酶的激活。我们现在发现，在分离的大鼠胰腺腺泡中存在第三种激活p38丝裂原活化蛋白（MAP）激酶的激酶级联反应。CCK和山梨醇诱导的渗透应激在数分钟内即可激活p38 MAP激酶；它们的作用呈剂量依赖性，最大激活倍数分别为2.8倍和4.4倍。卡巴胆碱和蛙皮素对p38 MAP激酶活性的影响与CCK相似，而佛波酯、表皮生长因子和血管活性肠肽对p38 MAP激酶的刺激作用则在2倍或以下。CCK和山梨醇均还增加了p38 MAP激酶的酪氨酰磷酸化。使用p38 MAP激酶的特异性抑制剂SB 203580，我们发现p38 MAP激酶活性是胰腺腺泡中MAP激酶激活的蛋白激酶-2激活所必需的。SB 203580降低了基础磷酸化水平，并阻断了CCK或山梨醇刺激后Hsp 27磷酸化的增加。CCK处理导致腺泡总F-肌动蛋白含量最初迅速下降，随后在40分钟后增加。用SB 203580预孵育可显著抑制F-肌动蛋白含量的这些变化。用罗丹明偶联的鬼笔环肽对肌动蛋白细胞骨架进行染色并通过共聚焦荧光显微镜分析显示，CCK刺激10分钟和40分钟后肌动蛋白细胞骨架遭到破坏。用SB 203580预处理可减少这些变化。这些发现表明，p38 MAP激酶的激活不仅参与应激反应，还参与诸如CCK等胃肠激素的生理信号传导，其中Gq偶联受体的激活会刺激一个级联反应，在该反应中p38 MAP激酶激活MAP激酶激活的蛋白激酶-2，导致Hsp 27磷酸化。p38 MAP激酶的激活，很可能是通过Hsp 27的磷酸化，在胰腺腺泡肌动蛋白细胞骨架的组织中发挥作用。
