Cellular Basis Of Action Of Gastrointestinal Peptides

最近的研究表明，胃肠激素，类似于许多生长因子，可以刺激细胞内酪氨酸磷酸化信号级联刺激细胞生长。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）激活这些级联的能力。CCKA-R激活导致p125 FAK、桩蛋白、p130 Cas、PYK 2和PKC delta的快速酪氨酸磷酸化。我们最近的研究表明，PKC δ的激活不依赖于[Ca 2 +]i或PI 3 K的变化，但受PKC-α激活的调节。粘着斑激酶（Focal adhesion kinases，p125 FAK，PYK）是介导整合素、生长因子、癌基因、生物活性脂质和某些G蛋白偶联受体对生长、粘附、细胞运动和细胞骨架变化的重要细胞内信号。关于GPCR如CCKA-R改变这些激酶的能力知之甚少。CCK可刺激p125 FAK三个位点（Y397、Y 577、Y 925）和PYK 2位点（pY 402、Y 580、Y881）的酪氨酸磷酸化，但在动力学、强度、不同受体状态的参与、PKC和胞浆钙的变化等方面存在差异。这些结果表明，这些不同位点的磷酸化是差异调节的，并涉及同一细胞中不同的细胞内机制。这些结果表明，这些不同的激酶的磷酸化和调节的差异可能有助于激活后在正常和肿瘤性疾病中的不同细胞效应。

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.