为了应对压力刺激,真核细胞通常抑制蛋白质合成。这导致了多体中mRNA的释放,它们与RNA结合蛋白的缩合,和称为应激颗粒(SGs)的非膜结合细胞质区室的形成。SGs含有40S,但通常缺乏60S核糖体亚基。众所周知,环己酰亚胺,依米汀,和茴香霉素,核糖体抑制剂阻断80S核糖体沿mRNA的进展并稳定多体,防止SG装配。相反,嘌呤霉素,导致过早终止,从多聚体释放mRNA并刺激SGs的形成。同样的效果是由一些翻译起始抑制剂引起的,这导致多聚体分解和以停滞的48S预起始复合物形式的mRNA积累。根据这些和其他数据,据认为,SG形成的触发因素是存在具有延伸的无核糖体片段的mRNA,在细胞内形成凝析油.在这项研究中,我们评估了各种小分子翻译抑制剂在亚砷酸钠诱导的严重氧化应激条件下阻断或刺激SGs组装的能力.与预期相反,我们发现特定类型的核糖体靶向延伸抑制剂,在mRNA编码区的开始处阻止单独的80S核糖体,但不干扰所有随后的核糖体完成翻译和离开转录本(例如三尖杉酯碱,乳酰霉素,或T-2毒素),完全防止亚砷酸盐诱导的SGs的形成。这些观察结果表明,即使mRNA上存在单个80S核糖体,也足以防止其募集到SGs中,并且mRNA的延伸的无核糖体区域的存在不足以形成SG。我们建议mRNA进入SGs可能是由RNA结合蛋白与核糖体相关时仍无法进入的40S亚基上的那些区域之间的特异性接触介导的。
In response to stress stimuli, eukaryotic cells typically suppress protein synthesis. This leads to the release of mRNAs from polysomes, their condensation with RNA-binding proteins, and the formation of non-membrane-bound cytoplasmic compartments called stress granules (SGs). SGs contain 40S but generally lack 60S ribosomal subunits. It is known that cycloheximide, emetine, and anisomycin, the ribosome inhibitors that block the progression of 80S ribosomes along mRNA and stabilize polysomes, prevent SG assembly. Conversely, puromycin, which induces premature termination, releases mRNA from polysomes and stimulates the formation of SGs. The same effect is caused by some translation initiation inhibitors, which lead to polysome disassembly and the accumulation of mRNAs in the form of stalled 48S preinitiation complexes. Based on these and other data, it is believed that the trigger for SG formation is the presence of mRNA with extended ribosome-free segments, which tend to form condensates in the cell. In this study, we evaluated the ability of various small-molecule translation inhibitors to block or stimulate the assembly of SGs under conditions of severe oxidative stress induced by sodium arsenite. Contrary to expectations, we found that ribosome-targeting elongation inhibitors of a specific type, which arrest solitary 80S ribosomes at the beginning of the mRNA coding regions but do not interfere with all subsequent ribosomes in completing translation and leaving the transcripts (such as harringtonine, lactimidomycin, or T-2 toxin), completely prevent the formation of arsenite-induced SGs. These observations suggest that the presence of even a single 80S ribosome on mRNA is sufficient to prevent its recruitment into SGs, and the presence of extended ribosome-free regions of mRNA is not sufficient for SG formation. We propose that mRNA entry into SGs may be mediated by specific contacts between RNA-binding proteins and those regions on 40S subunits that remain inaccessible when ribosomes are associated.