| Summary: | 博士 === 國立成功大學 === 臨床醫學研究所 === 104 === AKT regulates activated glycogen synthase kinase (GSK)-3β to facilitate IFN-γ signaling through the inhibition of Src homology-2 domain-containing phosphatase (SHP) 2. Mutated phosphoinositide 3-kinase (PI3K) and phosphatase and tensin homolog (PTEN) cause AKT activation and GSK-3β inactivation to induce SHP2-activated cellular hyporesponsiveness to IFN-γ in human gastric cancer AGS cells. Here, we want to discover whether some molecules for promoting AKT activation, GSK-3β inactivation, and SHP2 activation to cause IFN-γ resistance. First, the potential role of galectin-3 acts upstream of AKT will be investigated. Increasing or decreasing galectin-3 could change IFN-γ signaling. Following cisplatin-induced galectin-3 up-regulation, surviving cells showed cellular hyporesponsiveness to IFN-γ. Galectin-3 induced IFN-γ resistance independent of its extracellular β-galactoside-binding activity. Galectin-3 expression was not regulated by PI3K activation or a decrease in PTEN. Increased galectin-3 might cause GSK-3β inactivation and SHP2 activation by promoting PDK1-induced AKT phosphorylation at a threonine residue. Overexpression of AKT, inactive GSK-3βR96A, SHP2, or active SHP2D61A caused cellular hyporesponsiveness to IFN-γ in IFN-γ-sensitive MKN45 cells. IFN-γ-induced growth inhibition and apoptosis in AGS cells were observed until galectin-3 expression was down-regulated. In the first part, these results demonstrate that an increase in galectin-3 facilitates AKT/GSK-3β/SHP2 signaling, causing cellular hyporesponsiveness to IFN-γ. Integrin-linked kinase (ILK), a serine/ threonine kinase, regulates cell adhesion, migration, and proliferation, also particularly by promoting AKT signaling. However, aberrant increased ILK did not affect AKT/GSK-3β and IFN-γ-activated IRF1, indicating ILK does not contribute to IFN-γ resistance. The role of ILK regulation is not clearly in AGS cells. Previous study shows that ILK increased in gastric cancer is correlated with tumor grade and metastasis. The regulation for ILK overexpression and mechanisms for ILK-regulated gastric tumorigenesis has not documented. We next want to identify the molecular basis for ILK regulation and its alternative role in the ERK1/2/NF-κB-mediated stimulation of proliferation, migration, and survival. Genetically or pharmacologically inhibiting ILK abolished NF-κB-regulated cell proliferation. Unpredictably, ILK stimulated Ras activity by facilitating the activation of c-Raf/MEK1/2/ERK1/2/ribosomal S6 kinase/inhibitor of κBα/NF-κB signaling by a complex of IQ motif containing GTPase activating protein (IQGAP) 1 and Ras. Enforced enzymatic ILK expression promoted cell proliferation by facilitating ERK1/2/NF-κB signaling. PI3K activation or the decreased expression of PTEN prolonged ERK1/2 activation through the protection of ILK from proteasome-mediated degradation. The C-terminus of heat shock cognate 70 interacting protein, a HSP90-associated E3 ubiquitin ligase, mediated ILK ubiquitination to control PI3K- and HSP90-regulated ILK stabilization and signaling. In addition to proliferation, this pathway also determined cell migration and restricted the sensitivity of anticancer agents to 5-fluorouracil and cisplatin. Additionally, validating EGF/EGFR signaling confirmed the ERK-regulatory roles of ILK and IQGAP1. These results demonstrate that an increase in ILK facilitates IQGAP1 and Ras interaction and non-canonically promotes ERK1/2/NF-κB activation to benefit cell growth. Taken together, we investigate IFN-γ susceptibility and oncogenic regulation in gastric cancer cells.
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