Kaitsuka Taku, Tomizawa Kazuhito, Matsushita Masayuki
Department of Molecular Physiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan.
School of Pharmacy in Fukuoka, International University of Health and Welfare, Okawa, Japan.
Front Mol Biosci. 2021 Jan 14;7:598578. doi: 10.3389/fmolb.2020.598578. eCollection 2020.
Several variant proteins are produced from , including two representative proteins produced via alternative splicing machinery. One protein is the canonical translation eukaryotic elongation factor eEF1Bδ1, and the other is the heat shock-responsive transcription factor eEF1BδL. eEF1Bδ1 is phosphorylated by cyclin-dependent kinase 1 (CDK1), but the machinery controlling eEF1BδL phosphorylation and dephosphorylation has not been clarified. In this study, we found that both proteins were dephosphorylated under heat shock and proteotoxic stress, and this dephosphorylation was inhibited by okadaic acid. Using proteins with mutations at putative phosphorylated residues, we revealed that eEF1Bδ1 and eEF1BδL are phosphorylated at S133 and S499, respectively, and these residues are both CDK1 phosphorylation sites. The eEF1BδL S499A mutant more strongly activated promoter-driven reporter than the wild-type protein and S499D mutant. Furthermore, protein phosphatase 1 (PP1) was co-immunoprecipitated with eEF1Bδ1 and eEF1BδL, and PP1 dephosphorylated both proteins . Thus, this study clarified the role of phosphorylation/dephosphorylation in the functional regulation of eEF1BδL during heat shock.
几种变体蛋白由……产生，包括通过可变剪接机制产生的两种代表性蛋白。一种蛋白是典型的翻译真核延伸因子eEF1Bδ1，另一种是热休克反应转录因子eEF1BδL。eEF1Bδ1被细胞周期蛋白依赖性激酶1（CDK1）磷酸化，但控制eEF1BδL磷酸化和去磷酸化的机制尚未阐明。在本研究中，我们发现这两种蛋白在热休克和蛋白毒性应激下均发生去磷酸化，且这种去磷酸化被冈田酸抑制。利用在假定磷酸化位点具有突变的蛋白，我们发现eEF1Bδ1和eEF1BδL分别在S133和S499处被磷酸化，且这些位点均为CDK1磷酸化位点。eEF1BδL S499A突变体比野生型蛋白和S499D突变体更强烈地激活……启动子驱动的报告基因。此外，蛋白磷酸酶1（PP1）与eEF1Bδ1和eEF1BδL共免疫沉淀，且PP1使这两种蛋白去磷酸化。因此，本研究阐明了热休克期间磷酸化/去磷酸化在eEF1BδL功能调节中的作用。
