Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD, UK.
Department of Chemical Engineering, Centre for Process Systems Engineering, Institute for Systems and Synthetic Biology, Imperial College London, South Kensington Campus, London SW7 2AZ, UK.
Cells. 2019 Jul 31;8(8):800. doi: 10.3390/cells8080800.
In eukaryotes, the binding of poly(A) binding protein (PAB) to the poly(A) tail is central to maintaining mRNA stability. PABP interacts with the translation termination apparatus, and with eIF4G to maintain 3'-5' mRNA interactions as part of an mRNA closed loop. It is however unclear how ribosome recycling on a closed loop mRNA is influenced by the proximity of the stop codon to the poly(A) tail, and how post-termination ribosome recycling affects mRNA stability. We show that in a yeast disabled for nonsense mediated mRNA decay (NMD), a mRNA with an early stop codon at codon 22 of the reading frame is still highly unstable, and that this instability cannot be significantly countered even when 50% stop codon readthrough is triggered. In an NMD-deficient mutant yeast, stable reporter alleles with more 3' proximal stop codons could not be rendered unstable through Rli1-depletion, inferring defective Rli1 ribosome recycling is insufficient in itself to trigger mRNA instability. Mathematical modelling of a translation system including the effect of ribosome recycling and poly(A) tail shortening supports the hypothesis that impaired ribosome recycling from 5' proximal stop codons may compromise initiation processes and thus destabilize the mRNA. A model is proposed wherein ribosomes undergo a maturation process during early elongation steps, and acquire competency to re-initiate on the same mRNA as translation elongation progresses beyond the very 5' proximal regions of the mRNA.
在真核生物中，多聚腺苷酸结合蛋白 (PAB) 与多聚腺苷酸尾巴的结合对于维持 mRNA 的稳定性至关重要。PABP 与翻译终止装置相互作用，并与 eIF4G 结合，以维持 3'-5' mRNA 相互作用，作为 mRNA 闭环的一部分。然而，尚不清楚核糖体在闭环 mRNA 上的回收如何受到终止密码子与多聚腺苷酸尾巴的接近程度的影响，以及翻译终止后核糖体的回收如何影响 mRNA 的稳定性。我们表明，在酵母中，无义介导的 mRNA 降解 (NMD) 失活时，阅读框中密码子 22 处有早期终止密码子的 mRNA 仍然高度不稳定，即使触发 50%的终止密码子通读，这种不稳定性也不能得到显著抑制。在 NMD 缺陷型酵母突变体中，即使耗尽 Rli1 也不能使带有更靠近 3' 端终止密码子的稳定报告基因等位基因不稳定，这表明 Rli1 核糖体回收缺陷本身不足以触发 mRNA 不稳定。包括核糖体回收和多聚腺苷酸尾巴缩短效应的翻译系统的数学模型支持这样一种假设，即从 5' 近端终止密码子的核糖体回收受损可能会损害起始过程，从而使 mRNA 不稳定。提出了一种模型，其中核糖体在早期延伸步骤中经历成熟过程，并且在翻译延伸超过 mRNA 的非常 5' 近端区域时，获得在同一 mRNA 上重新起始的能力。
