Transformation of elongation factor 1Bdelta into heat shock response transcription factor by alternative splicing. Homo sapiens

Protein translation factors play crucial roles in a variety of stress responses. Here, we show that the eukaryotic elongation factor 1Bdelta (eEF1Bdelta) changes its structure and function from a translation factor into a heat shock response transcription factor by alternative splicing. While eEF1Bdelta is specifically localized in the cytoplasm, the long isoform of eEF1Bdelta (eEF1BdeltaL) is localized in the nucleus and induces heat shock element (HSE)-containing genes in cooperation with heat-shock transcription factor 1 (HSF1). Moreover, the N-terminal domain of eEF1BdeltaL binds with NF-E2-related factor 2 (Nrf2) and induces stress response heme oxygenase 1 (HO-1). Specific inhibition of eEF1BdeltaL with siRNA completely inhibits Nrf2-dependent HO-1 induction. In addition, eEF1BdeltaL directly binds to HSE oligo DNA in vitro and associates with HSE containing the HO-1-enhancer region in vivo. Thus, the transcriptional role of eEF1BdeltaL could provide new insights into the molecular mechanism of stress responses. We performed microarray analysis to compare the gene expression induced by eEF1Bdelta1 or eEF1BdeltaL overexpression. Overall design: HEK293 cells transfected with expression plasmids encoding flag-tagged-eEF1Bdelta1 or eEF1BdeltaL protein

蛋白质翻译因子在多种应激反应中发挥关键作用。本研究发现，真核延伸因子1Bδ（eEF1Bdelta）可通过可变剪接，从翻译因子转变为热激反应转录因子，同时改变自身结构与功能。常规状态下，eEF1Bdelta特异性定位于细胞质，而其长亚型eEF1BdeltaL则定位于细胞核，并可与热激转录因子1（HSF1）协同诱导携带热激元件（HSE）的基因表达。此外，eEF1BdeltaL的N端结构域可与NF-E2相关因子2（Nrf2）结合，进而诱导应激反应相关基因血红素加氧酶1（HO-1）的转录。通过小干扰RNA（siRNA）特异性抑制eEF1BdeltaL，可完全阻断Nrf2介导的HO-1诱导过程。进一步实验证实，eEF1BdeltaL在体外可直接结合携带HSE的寡聚DNA，在体内则可与HO-1增强子区域的HSE位点相结合。综上，eEF1BdeltaL的转录调控功能可为应激反应的分子机制研究提供全新视角。
本研究通过基因芯片分析，比较了eEF1Bdelta1与eEF1BdeltaL过表达所诱导的基因表达差异。实验整体设计：将转染了编码FLAG标签标记的eEF1Bdelta1或eEF1BdeltaL蛋白的表达质粒的HEK293细胞作为实验样本。