Nature Structural & Molecular Biology ( IF 12.5 ) Pub Date : 2024-02-19 , DOI: 10.1038/s41594-023-01187-1
Young-Suk Lee 1, 2 , Yevgen Levdansky 3 , Yoonseok Jung 1 , V Narry Kim 1, 4 , Eugene Valkov 3
|
Shortening of messenger RNA poly(A) tails, or deadenylation, is a rate-limiting step in mRNA decay and is highly regulated during gene expression. The incorporation of non-adenosines in poly(A) tails, or ‘mixed tailing’, has been observed in vertebrates and viruses. Here, to quantitate the effect of mixed tails, we mathematically modeled deadenylation reactions at single-nucleotide resolution using an in vitro deadenylation system reconstituted with the complete human CCR4–NOT complex. Applying this model, we assessed the disrupting impact of single guanosine, uridine or cytosine to be equivalent to approximately 6, 8 or 11 adenosines, respectively. CCR4–NOT stalls at the 0, −1 and −2 positions relative to the non-adenosine residue. CAF1 and CCR4 enzyme subunits commonly prefer adenosine but exhibit distinct sequence selectivities and stalling positions. Our study provides an analytical framework to monitor deadenylation and reveals the molecular basis of tail sequence-dependent regulation of mRNA stability.
中文翻译:

单核苷酸分辨率下混合 Poly(A) 尾的脱腺苷酸化动力学
信使 RNA Poly(A) 尾部的缩短或脱腺苷化是 mRNA 衰减的限速步骤,并且在基因表达过程中受到高度调控。已在脊椎动物和病毒中观察到非腺苷掺入聚腺苷酸尾部或“混合尾部”。在这里,为了定量混合尾的影响,我们使用用完整的人 CCR4-NOT 复合物重建的体外去腺苷化系统,以单核苷酸分辨率对去腺苷化反应进行了数学建模。应用该模型,我们评估了单个鸟苷、尿苷或胞嘧啶的破坏影响分别相当于大约 6、8 或 11 个腺苷。 CCR4–NOT 相对于非腺苷残基停在 0、-1 和 -2 位置。 CAF1 和 CCR4 酶亚基通常更喜欢腺苷,但表现出不同的序列选择性和停滞位置。我们的研究提供了一个监测去腺苷化的分析框架,并揭示了尾部序列依赖性调节 mRNA 稳定性的分子基础。