Wennström S, Landgren E, Blume-Jensen P, Claesson-Welsh L
Ludwig Institute for Cancer Research, Uppsala Branch, Sweden.
J Biol Chem. 1992 Jul 5;267(19):13749-56.
Signal transduction by tyrosine kinase growth factor receptors involves ligand-induced phosphorylation of substrates for the kinase, resulting in mediation of common or receptor-specific biological signals. We have compared signal transduction pathways for the fibroblast growth factor receptor-1 (FGFR-1), the platelet-derived growth factor beta-receptor (PDGFR-beta), and a chimeric FGFR-1 molecule, FGFRchim, in which the FGFR-1 kinase insert was replaced with that of the PDGFR-beta. The different receptors were characterized and found to be functional as ligand-stimulatable kinases, after expression of the respective human cDNAs in porcine aortic endothelial cells. Substrates for the receptors were analyzed by ligand stimulation of [32P]orthophosphate-labeled cells and immunoprecipitation with phosphotyrosine antiserum. A number of phosphoproteins were induced in all the different types of cells, but components specifically induced after stimulation of FGFR-1 and PDGFR-beta expressing cells could also be detected. Examination of receptor-associated substrates by in vitro kinase assays revealed phosphoproteins of 65 and 85 kDa, which were associated with PDGFR-beta and FGFRchim, but not with FGFR-1. The 85-kDa phosphoprotein could correspond to the regulatory subunit of phosphatidylinositol 3' kinase (PI3-K), since phosphatidylinositol 3' kinase activity was detected after ligand stimulation of FGFRchim- and PDGFR-beta- but not FGFR-1-expressing cells. In addition, ligand stimulation of FGFRchim- and PDGFR-beta-expressing cells, but not FGFR-1-expressing cells, led to induction of actin reorganization in the form of circular membrane ruffling. Thus, replacement of a discrete segment of the intracellular domain of the FGFR-1 with the corresponding stretch from the PDGFR-beta resulted in transfer of PDGFR-beta-specific signaling properties to the chimeric molecule.
酪氨酸激酶生长因子受体介导的信号转导涉及激酶底物的配体诱导磷酸化，从而介导常见的或受体特异性的生物学信号。我们比较了成纤维细胞生长因子受体-1（FGFR-1）、血小板衍生生长因子β受体（PDGFR-β）以及一种嵌合型FGFR-1分子FGFRchim的信号转导途径，在FGFRchim中，FGFR-1的激酶插入区被PDGFR-β的激酶插入区所取代。在猪主动脉内皮细胞中分别表达各自的人cDNA后，对不同的受体进行了表征，发现它们作为可被配体刺激的激酶发挥功能。通过对[32P]正磷酸盐标记的细胞进行配体刺激并用抗磷酸酪氨酸抗血清进行免疫沉淀，分析了受体的底物。在所有不同类型的细胞中都诱导产生了一些磷酸化蛋白，但也能检测到在刺激表达FGFR-1和PDGFR-β的细胞后特异性诱导产生的成分。通过体外激酶测定对受体相关底物进行检测，发现了65 kDa和85 kDa的磷酸化蛋白，它们与PDGFR-β和FGFRchim相关，但与FGFR-1无关。85 kDa的磷酸化蛋白可能对应于磷脂酰肌醇3'激酶（PI3-K）的调节亚基，因为在配体刺激表达FGFRchim和PDGFR-β的细胞后检测到了磷脂酰肌醇3'激酶活性，而在刺激表达FGFR-1的细胞后未检测到。此外，配体刺激表达FGFRchim和PDGFR-β的细胞，而不是表达FGFR-1的细胞，导致以圆形膜皱襞形式的肌动蛋白重组。因此，用PDGFR-β的相应片段替换FGFR-1细胞内结构域的一个离散片段，导致PDGFR-β特异性信号特性转移到嵌合分子上。
