InsP7 is a small-molecule regulator of NUDT3-mediated mRNA decapping and processing-body dynamics
InsP7 is a small-molecule regulator of NUDT3-mediated mRNA decapping and processing-body dynamics
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DOI：
10.1073/pnas.1922284117
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发表时间：
2020-08-11
影响因子：
11.1
通讯作者：
Shears, Stephen B.
中科院分区：
文献类型：
--
作者：
Sahu, Soumyadip;Wang, Zhenzhen;Shears, Stephen B.
信使RNA （mRNA）的酶促5'脱帽调控通常会破坏转录物，但它具有动态重塑转录组的能力。例如，免受5'脱帽的保护促进mrna在加工(P)体（无膜的生物分子凝聚体）中的积累。mrna的这种分区化暂时将它们从可翻译库中移除；这些被抑制的转录物被稳定和储存，直到p体溶解允许转录物重新进入细胞质。在这里，我们描述了肌醇焦磷酸信号分子5-InsP（5-二磷酸肌醇五磷酸）对mRNA稳定性和p体动力学的调节。首先，我们在体外证明了5-InsP(7)抑制重组NUDT3 （Nudix[核苷二磷酸连接片段X]型水解酶3）的脱帽。接下来，在完整的HEK293和Hal 16细胞中，我们在CRISPRCas9敲除PPIP5Ks（二磷酸肌醇五磷酸5激酶1型和2型，即PPIP5K KO）后，监测了NUDT3 mRNA底物骨干的稳定性，这将细胞5-InsP(7)水平提高了两到三倍（即在生理流变范围内）。PPIP5K KO细胞表现出NUDT3 mRNA底物水平升高和p体丰度增加。KO背景下5-InsP(7)合成的药理学和遗传学减弱使NUDT3 mRNA底物水平和p -体计数恢复到野生型细胞的水平。此外，脂质体将代谢耐药的5-InsP(7)类似物递送到野生型细胞中，可提高NUDT3 mRNA底物水平，并提高p体丰度。在细胞5-InsP(7)水平通常会随着生物能量环境的变化而波动的背景下，这种肌醇焦磷酸对mRNA结构的调节代表了一个表转录组控制过程。对p体动力学的相关影响与干细胞分化、应激反应的调节有关，并可能改善神经退行性疾病和衰老。
Regulation of enzymatic 5' decapping of messenger RNA (mRNA), which normally commits transcripts to their destruction, has the capacity to dynamically reshape the transcriptome. For example, protection from 5' decapping promotes accumulation of mRNAs into processing (P) bodies-membraneless, biomolecular condensates. Such compartmentalization of mRNAs temporarily removes them from the translatable pool; these repressed transcripts are stabilized and stored until P-body dissolution permits transcript reentry into the cytosol. Here, we describe regulation of mRNA stability and P-body dynamics by the inositol pyrophosphate signaling molecule 5-InsP(7) (5-diphosphoinositol pentakisphosphate). First, we demonstrate 5-InsP(7) inhibits decapping by recombinant NUDT3 (Nudix [nucleoside diphosphate linked moiety X]-type hydrolase 3) in vitro. Next, in intact HEK293 and Hal 16 cells, we monitored the stability of a cadre of NUDT3 mRNA substrates following CRISPRCas9 knockout of PPIP5Ks (diphosphoinositol pentakisphosphate 5-kinases type 1 and 2, i.e., PPIP5K KO), which elevates cellular 5-InsP(7) levels by two- to threefold (i.e., within the physiological rheostatic range). The PPIP5K KO cells exhibited elevated levels of NUDT3 mRNA substrates and increased P-body abundance. Pharmacological and genetic attenuation of 5-InsP(7) synthesis in the KO background reverted both NUDT3 mRNA substrate levels and P-body counts to those of wild-type cells. Furthermore, liposomal delivery of a metabolically resistant 5-InsP(7) analog into wild-type cells elevated levels of NUDT3 mRNA substrates and raised P-body abundance. In the context that cellular 5-InsP(7) levels normally fluctuate in response to changes in the bioenergetic environment, regulation of mRNA structure by this inositol pyrophosphate represents an epitranscriptomic control process. The associated impact on P-body dynamics has relevance to regulation of stem cell differentiation, stress responses, and, potentially, amelioration of neurodegenerative diseases and aging.