Center for RNA Biology, Columbus, OH 43210, USA.
Department of Biological Chemistry and Pharmacology, Columbus, OH 43210, USA.
Nucleic Acids Res. 2020 Apr 17;48(7):3806-3815. doi: 10.1093/nar/gkaa046.
Cap homeostasis is the cyclical process of decapping and recapping that maintains the translation and stability of a subset of the transcriptome. Previous work showed levels of some recapping targets decline following transient expression of an inactive form of RNMT (ΔN-RNMT), likely due to degradation of mRNAs with improperly methylated caps. The current study examined transcriptome-wide changes following inhibition of cytoplasmic cap methylation. This identified mRNAs with 5'-terminal oligopyrimidine (TOP) sequences as the largest single class of recapping targets. Cap end mapping of several TOP mRNAs identified recapping events at native 5' ends and downstream of the TOP sequence of EIF3K and EIF3D. This provides the first direct evidence for downstream recapping. Inhibition of cytoplasmic cap methylation was also associated with mRNA abundance increases for a number of transcription, splicing, and 3' processing factors. Previous work suggested a role for alternative polyadenylation in target selection, but this proved not to be the case. However, inhibition of cytoplasmic cap methylation resulted in a shift of upstream polyadenylation sites to annotated 3' ends. Together, these results solidify cap homeostasis as a fundamental process of gene expression control and show cytoplasmic recapping can impact regulatory elements present at the ends of mRNA molecules.
帽状结构的动态平衡是帽结构的去帽和再帽的循环过程，它维持了转录组的一部分的翻译和稳定性。以前的工作表明，在非活性形式的 RNMT（ΔN-RNMT）短暂表达后，一些再帽靶标水平下降，这可能是由于带有不正确甲基化帽的 mRNAs 的降解。本研究在抑制细胞质帽甲基化后，对全转录组范围的变化进行了研究。这确定了具有 5'-末端寡嘧啶（TOP）序列的 mRNAs 作为最大的再帽靶标单一类别。对几种 TOP mRNAs 的帽端映射确定了在 EIF3K 和 EIF3D 的天然 5'端和 TOP 序列下游的再帽事件。这首次提供了下游再帽的直接证据。细胞质帽甲基化的抑制也与许多转录、剪接和 3'加工因子的 mRNA 丰度增加有关。以前的工作表明，可变多聚腺苷酸化在靶标选择中起作用，但事实并非如此。然而，细胞质帽甲基化的抑制导致上游多聚腺苷酸化位点向注释的 3'末端转移。总之，这些结果证实了帽状结构的动态平衡是基因表达调控的一个基本过程，并表明细胞质再帽可以影响 mRNA 分子末端的调节元件。
