Cells adjust to hypoxic stress within the tumor microenvironment by downregulating energy-consuming processes including translation. To delineate mechanisms of cellular adaptation to hypoxia, we performed RNA-Seq of normoxic and hypoxic head and neck cancer cells. These data revealed a significant down regulation of genes known to regulate RNA processing and splicing. Exon-level analyses classified > 1,000 mRNAs as alternatively spliced under hypoxia and uncovered a unique retained intron (RI) in the master regulator of translation initiation, EIF2B5. Notably, this intron was expressed in solid tumors in a stage-dependent manner. We investigated the biological consequence of this RI and demonstrate that its inclusion creates a premature termination codon (PTC), that leads to a 65kDa truncated protein isoform that opposes full-length eIF2Bε to inhibit global translation. Furthermore, expression of 65kDa eIF2Bε led to increased survival of head and neck cancer cells under hypoxia, providing evidence that this isoform enables cells to adapt to conditions of low oxygen. Additional work to uncover -cis and -trans regulators of EIF2B5 splicing identified several factors that influence intron retention in EIF2B5: a weak splicing potential at the RI, hypoxia-induced expression and binding of the splicing factor SRSF3, and increased binding of total and phospho-Ser2 RNA polymerase II specifically at the intron retained under hypoxia. Altogether, these data reveal differential splicing as a previously uncharacterized mode of translational control under hypoxia and are supported by a model in which hypoxia-induced changes to cotranscriptional processing lead to selective retention of a PTC-containing intron in EIF2B5.

细胞通过下调包括翻译在内的耗能过程来适应肿瘤微环境中的低氧应激。为了描述细胞适应缺氧的机制，我们进行了常氧和低氧的头颈癌细胞的RNA-Seq。这些数据揭示了已知调节RNA加工和剪接的基因的显着下调。外显子水平分析将> 1,000个mRNA划分为在缺氧条件下进行剪接，并在翻译起始的主调节子EIF2B5中发现了一个独特的内含子（RI）。值得注意的是，该内含子在实体瘤中以阶段依赖性的方式表达。我们调查了此RI的生物学结果，并证明了其包含会产生一个过早的终止密码子（PTC），从而导致65kDa的截短蛋白同工型与全长eIF2Bε相对，以抑制全局翻译。此外，在缺氧条件下65kDaeIF2Bε的表达导致头颈癌细胞的存活率提高，这证明该同种型能够使细胞适应低氧条件。额外的工作，揪出顺和反式的监管EIF2B5拼接确定了几个因素内含子保留的影响EIF2B5：在RI上的剪接能力弱，缺氧诱导的剪接因子SRSF3的表达和结合，总和磷酸Ser2 RNA聚合酶II的结合增加，特别是在缺氧条件下保留的内含子上。总而言之，这些数据揭示了差异剪接作为缺氧条件下翻译控制的先前未表征模式，并且得到了模型的支持，在该模型中，缺氧诱导的共转录加工变化导致EIF2B5中含PTC的内含子的选择性保留。