德国海德堡大学Johanna Schott开发出新方法，可用于测量核糖体装载时间并揭示对核糖体密度的动力学影响。相关论文于2021年9月3日在线发表于国际学术期刊《自然—方法学》。

为了测量新生mRNA上的核糖体密度（RD），研究人员将Ribo-Seq与渐进式4-硫代尿苷标记相结合，开发了新生Ribo-Seq。在小鼠巨噬细胞中，研究人员通过实验确定了新生mRNA的出现和它与核糖体结合之间的滞后期，经计算，大量mRNA的滞后期为20-22分钟。在小鼠胚胎干细胞中，nRibo-Seq显示核糖体装载的滞后性更强，达到35-38分钟。

在用脂多糖刺激巨噬细胞后，细胞质和翻译的mRNA之间的滞后导致输入和核糖体保护的片段之间的解耦，这在mRNA数量远离稳态表达的条件下引起扭曲的RD测量。因此，研究人员证明转录变化以一种被动的方式影响RD。

据介绍，一般来说，mRNA被认为在进入细胞质后会立即被核糖体装载。

附：英文原文

Title: Nascent Ribo-Seq measures ribosomal loading time and reveals kinetic impact on ribosome density

Author: Schott, Johanna, Reitter, Sonja, Lindner, Doris, Grosser, Jan, Bruer, Marius, Shenoy, Anjana, Geiger, Tamar, Mathes, Arthur, Dobreva, Gergana, Stoecklin, Georg

Issue&Volume: 2021-09-03

Abstract: In general, mRNAs are assumed to be loaded with ribosomes instantly upon entry into the cytoplasm. To measure ribosome density (RD) on nascent mRNA, we developed nascent Ribo-Seq by combining Ribo-Seq with progressive 4-thiouridine labeling. In mouse macrophages, we determined experimentally the lag between the appearance of nascent mRNA and its association with ribosomes, which was calculated to be 20–22min for bulk mRNA. In mouse embryonic stem cells, nRibo-Seq revealed an even stronger lag of 35–38min in ribosome loading. After stimulation of macrophages with lipopolysaccharide, the lag between cytoplasmic and translated mRNA leads to uncoupling between input and ribosome-protected fragments, which gives rise to distorted RD measurements under conditions where mRNA amounts are far from steady-state expression. As a result, we demonstrate that transcriptional changes affect RD in a passive way.

DOI: 10.1038/s41592-021-01250-z

Source: https://www.nature.com/articles/s41592-021-01250-z