Andreolotti Alberto G, Bragado Maria J, Tapia Jose A, Jensen Robert T, Garcia-Marin Luis J
Departamento de Fisiologia, Universidad de Extremadura, Caceres, Spain.
Eur J Biochem. 2003 Dec;270(23):4706-13. doi: 10.1046/j.1432-1033.2003.03869.x.
Crk belongs to a family of adapter proteins whose structure allows interaction with tyrosine-phosphorylated proteins and is therefore an important modulator of downstream signals, representing a convergence of the actions of numerous stimuli. Recently, it was demonstrated that cholecystokinin (CCK) induced tyrosine phosphorylation of proteins related to fiber stress formation in rat pancreatic acini. Here, we investigated whether CCK receptor activation signals through CrkII and forms complexes with tyrosine-phosphorylated proteins in rat pancreatic acini. We demonstrated that CCK promoted the transient formation of CrkII-paxillin and CrkII-p130Cas complexes with maximal effect at 1 min. Additionally, CCK decreased the electrophoretic mobility of CrkII. This decrease was time- and concentration-dependent and inversely related with its function. Carbachol and bombesin also decreased CrkII electrophoretic mobility, whereas epidermal growth factor, vasoactive intestinal peptide, secretin or pituitary adenylate cyclase-activating polypeptide had no effect. CCK-induced CrkII electrophoretic shift was dependent on the Src family of tyrosine kinases and occurred in the intact animal, suggesting a physiological role of CrkII mediating CCK actions in the exocrine pancreas in vivo.
Crk属于衔接蛋白家族，其结构允许与酪氨酸磷酸化蛋白相互作用，因此是下游信号的重要调节因子，代表了多种刺激作用的汇聚。最近，有研究表明胆囊收缩素（CCK）可诱导大鼠胰腺腺泡中与纤维应激形成相关的蛋白发生酪氨酸磷酸化。在此，我们研究了CCK受体激活是否通过CrkII发出信号并在大鼠胰腺腺泡中与酪氨酸磷酸化蛋白形成复合物。我们证明CCK促进了CrkII-桩蛋白和CrkII-p130Cas复合物的瞬时形成，在1分钟时效果最佳。此外，CCK降低了CrkII的电泳迁移率。这种降低具有时间和浓度依赖性，且与其功能呈负相关。卡巴胆碱和蛙皮素也降低了CrkII的电泳迁移率，而表皮生长因子、血管活性肠肽、促胰液素或垂体腺苷酸环化酶激活多肽则没有作用。CCK诱导的CrkII电泳迁移率变化依赖于酪氨酸激酶的Src家族，且在完整动物体内发生，这表明CrkII在体内外分泌胰腺中介导CCK作用具有生理作用。
