Cellular Basis Of Action Of Gastrointestinal Peptides

最近的研究表明，胃肠道激素与许多生长因子类似，可能通过刺激细胞内酪氨酸磷酸化信号级联反应来刺激细胞生长。然而，目前对这些g蛋白偶联受体激活这些级联反应的能力知之甚少。在这一年中，我们研究了激素/神经递质胆囊收缩素（CCK）激活这些级联反应的能力。CCKA-R激活导致p125FAK、paxillin、p130Cas、PYK2和PKC delta的酪氨酸快速磷酸化。我们最近的研究表明PKC δ激活与[Ca2+]i或PI3K的变化无关，但受PKC- α的激活调节。局灶黏附激酶（p125FAK，PYK）是介导整合素、生长因子、癌基因、生物活性脂质和一些G蛋白偶联受体对生长、黏附、细胞运动和细胞骨架变化的重要细胞内信号。很少有人知道GPCR如CCKA-R改变这些激酶的能力。我们证明CCK可以刺激三个p125FAK位点（Y397,Y577,Y925）和PYK2位点（pY402,Y580,Y881）的酪氨酸磷酸化，然而，它们在动力学，强度，不同受体状态的参与，PKC和胞质钙的变化方面存在差异。这些结果表明，在同一细胞中，这些不同位点的磷酸化受到不同的调控，涉及不同的细胞内机制。这些结果表明，这些不同激酶的磷酸化和调控的差异可能导致它们在激活后对正常和肿瘤疾病的不同细胞作用。

Recent studies show that gastrointestinal hormones, similar to many growth factors, may stimulate cell growth by stimulating intracellular tyrosine phosphorylation signaling cascades. However at present little is known about the ability of these G-protein-coupled receptors to activate these cascades. During the year we have investigated the ability of the hormone/neurotransmitter, cholecystokinin (CCK) to activate these cascades. CCKA-R activation causes rapid tyrosine phosphorylation of p125FAK, paxillin, p130Cas, PYK2 and PKC delta. Our recent studies show PKC delta activation is independent of changes in [Ca2+]i or PI3K but is regulated by activation of PKC-alpha. Focal adhesion kinases (p125FAK,PYK) are important intracellular signals mediating effects of integrins, growth factors, oncogenes, bioactive lipids and some G protein-coupled receptors on growth, adhesion, cellular motility and cytoskeletal changes. Little is known on the ability of GPCR such as the CCKA-R to alter these kinases. We demonstrated CCK can stimulate tyrosine phosphorylation at three p125FAK sites (Y397,Y577,Y925) and PYK2 sites (pY402,Y580,Y881), however, they differ in kinetics, magnitude, participation of the different receptor states, PKC and changes in cytosolic calcium. These results show that phosphorylation of these different sites is differentially regulated and involves different intracellular mechanisms in the same cell. These results suggest that differences in the phosphorylation and regulation of these different kinases likely contribute to their different cellular effects in normal and neoplastic diseases after activation.