Division of Radiobiology and Molecular Environmental Research, Eberhard Karls University, Tuebingen, Germany.
Radiother Oncol. 2011 Oct;101(1):140-6. doi: 10.1016/j.radonc.2011.06.004. Epub 2011 Jun 30.
In the present study effect of erbB2 as well as DNA-PKcs on ionizing radiation (IR)- and erbB1 ligand-induced phosphorylation of Akt at S473 in cytoplasmic and nuclear fractions was investigated.
DNA-PKcs proficient and deficient syngeneic colon carcinoma sublines of HCT116 and the glioblastoma cell lines MO59K and MO59J as well as the lung carcinoma cell line A549 were used. Akt-S473 phosphorylation was investigated in cells pre-treated with pharmacological inhibitors or transfected with siRNA by immunoprecipitation, Western blotting and confocal microscopy after different stimuli, i.e., ligands and IR.
IR-induced phosphorylation of Akt in both MO59K and MO59J cell lines but not in HCT116 cells was DNA-PKcs dependent. In A549 cells, IR-induced phosphorylation of nuclear Akt-S473 was dependent on erbB1, erbB2, and DNA-PKcs. EGF induced phosphorylation of nuclear Akt-S473 in a DNA-PKcs and erbB2 independent manner.
Data indicate that the function of DNA-PKcs on IR-induced Akt-S473 phosphorylation is cell line specific. IR-induced, but not EGF-induced phosphorylation of cytoplasmic and/or nuclear Akt-S473 is erbB2 dependent.
本研究旨在探究 erbB2 以及 DNA-PKcs 对细胞质和核部分 Akt 在 S473 位点处因电离辐射（IR）和 erbB1 配体诱导而发生的磷酸化的影响。
使用了 DNA-PKcs 功能正常和缺失的同源结肠癌细胞亚系 HCT116 以及胶质母细胞瘤细胞系 MO59K 和 MO59J ，还有肺腺癌细胞系 A549。通过免疫沉淀、Western blot 和共聚焦显微镜，在细胞经药理学抑制剂预处理或转染 siRNA 后，对不同刺激（如配体和 IR）下 Akt-S473 的磷酸化情况进行研究。
IR 诱导的 MO59K 和 MO59J 细胞系中 Akt 的磷酸化依赖于 DNA-PKcs，但在 HCT116 细胞中则不然。在 A549 细胞中，IR 诱导的核 Akt-S473 的磷酸化依赖于 erbB1、erbB2 和 DNA-PKcs。EGF 以 DNA-PKcs 和 erbB2 非依赖性方式诱导核 Akt-S473 的磷酸化。
数据表明，DNA-PKcs 在 IR 诱导的 Akt-S473 磷酸化中的作用具有细胞系特异性。IR 诱导的、而非 EGF 诱导的细胞质和/或核 Akt-S473 的磷酸化依赖于 erbB2。
