J Biol Chem. 2014 Jul 25;289(30):20706-16. doi: 10.1074/jbc.M114.547828.
Initial steps in protein synthesis are highly regulated processes as they define the reading frame of the translation machinery. Eukaryotic translation initiation is a process facilitated by numerous factors (eIFs), aimed to form a "scanning" mechanism toward the initiation codon. Translation initiation of the main open reading frame (ORF) in an mRNA transcript has been reported to be regulated by upstream open reading frames (uORFs) in a manner of re-initiation. This mode of regulation is governed by the phosphorylation status of eIF2α and controlled by cellular stresses. Another mode of translational initiation regulation is leaky scanning, and this regulatory process has not been extensively studied. We have identified arsenite- inducible regulatory particle-associated protein (AIRAP) transcript to be translationally induced during arsenite stress conditions. AIRAP transcript contains a single uORF in a poor-kozak context. AIRAP translation induction is governed by means of leaky scanning and not re-initiation. This induction of AIRAP is solely dependent on eIF1 and the uORF kozak context. We show that eIF1 is phosphorylated under specific conditions that induce protein misfolding and have biochemically characterized this site of phosphorylation. Our data indicate that leaky scanning like re-initiation is responsive to stress conditions and that leaky scanning can induce ORF translation by bypassing poor kozak context of a single uORF transcript.
蛋白质合成的初始步骤是高度受调控的过程，因为它们定义了翻译机器的阅读框架。真核翻译起始是一个由众多因子（eIFs）促进的过程，旨在形成一个“扫描”机制，朝向起始密码子。已经报道了 mRNA 转录物中的主要开放阅读框（ORF）的翻译起始受上游开放阅读框（uORF）以重新起始的方式调控。这种调节模式受 eIF2α 的磷酸化状态控制，并受细胞应激调节。另一种翻译起始调控模式是漏扫，并且这种调控过程尚未得到广泛研究。我们已经确定在亚砷酸盐应激条件下，砷诱导调节颗粒相关蛋白（AIRAP）转录本被翻译诱导。AIRAP 转录本在较差的 Kozak 背景下含有单个 uORF。AIRAP 翻译诱导是通过漏扫而不是重新起始来控制的。AIRAP 的这种诱导仅依赖于 eIF1 和 uORF Kozak 上下文。我们表明，在诱导蛋白质错误折叠的特定条件下，eIF1 被磷酸化，并对该磷酸化位点进行了生化表征。我们的数据表明，与重新起始一样，漏扫对应激条件有反应，并且漏扫可以通过绕过单个 uORF 转录本较差的 Kozak 上下文来诱导 ORF 翻译。
