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Translation machinery: the basis of translational control

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Journal of Genetics and Genomics 2024年第4期51卷 367-378页

作者： Shu Yuan Guilong Zhou Guoyong Xu State Key Laboratory of Hybrid Rice Institute for Advanced Studies(IAS)Wuhan UniversityWuhanHubei 430072China Hubei Hongshan Laboratory WuhanHubei 430070China

Messenger RNA(mRNA)translation consists of initiation,elongation,termination,and ribosome recycling,carried out by the translation machinery,primarily including tRNAs,ribosomes,and translation factors(TrFs).Translational regulators transduce signals of growth and development,as well as biotic and abiotic stresses,to the translation machinery,where global or selective translational control occurs to modulate mRNA translation efficiency(TrE).As the basis of translational control,the translation machinery directly determines the quality and quantity of newly synthesized peptides and,ultimately,the cellular adaption.Thus,regulating the availability of diverse machinery components is reviewed as the central strategy of translational control.We provide classical signaling pathways(e.g.,integrated stress responses)and cellular behaviors(e.g.,liquideliquid phase separation)to exemplify this strategy within different physiological contexts,particularly during hostemicrobe interactions.With new technologies developed,further understanding this strategy will speed up translational medicine and translational agriculture.

关键词： Translational control Ribosome tRNA Translation machinery uORF Phase separation Immune response

Orthogonality of Redesigned tRNA Molecules with Three Stop Codons

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Chinese Journal of Chemistry 2022年第7期40卷 825-831页

作者： Zhao-Yang Zhang Dan-Ni Liao Yu-Xin Ma Bin Jia Ying-Jin Yuan Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering(Ministry of Education) School of Chemical Engineering and TechnologyTianjin UniversityTianjin 300072China Collaborative Innovation Center of Chemical Science and Engineering(Tianjin) Tianjin UniversityTianjin 300072China

The ability to expand genetic code in living cells has emerged as a powerful method with diverse applications.Here,we design re-placement of the anticodons of E.coli tRNAs that recognize codons for 20 natural amino acids,with three anti-stop codons,resulting in a total of 60 engineered tRNA constructs.We test these constructs one by one in E.coli,and found that six tRNAsCUA(tyrV,serX,hisR,trpT,glnV and leuX),two tRNAsUCA(trpT and leuX)and one tRNAUUA(tyrV)allowed efficient expression of Red Fluorescence Protein(RFP)with the presence of a corresponding stop codon in frame.Furthermore,we exploit the mutual orthogonality of tRNASer CUA,tRNATrpUCA and tRNATyrUUA with corresponding stop codons and demonstrated that the tRNASer CUA and the tRNATrp UCA can provide dynamic range and low crosstalk.Finally,we show the TAG and TGA can not only be used as an"AND gate"circuit to control the translation of target gene,but also be used to control the translation of a prodeoxyviolacein(PDV)pathway and a reporter in parallel.Overall,this work provides flexible tools for translational control and holds great potential to promote synthetic biology studies.

关键词： tRNA Orthogonality Synthetic biology Series control Parallel control Biosynthesis

线粒体RNA m^(3)C修饰酶METTL8的修饰机制和底物特异性

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科学通报 2024年第14期69卷 1806-1809页

作者：黄梦涵周小龙国科大杭州高等研究院生命与健康科学学院浙江省系统健康科学重点实验室杭州310024 中国科学院分子细胞科学卓越创新中心/上海生物化学与细胞生物学研究所核糖核酸功能与应用全国重点实验室上海200031

tRNA是遗传信息传递的关键生物大分子之一.在核糖体中,tRNA的反密码子通过与mRNA的三联体密码子互补配对,从而将其携带的氨基酸掺入到新合成的肽链中,对遗传信息的精准传递具有重要作用^([1]).tRNA上存在着大量的转录后核苷酸修饰^([2])... 详细信息

tRNA是遗传信息传递的关键生物大分子之一.在核糖体中,tRNA的反密码子通过与mRNA的三联体密码子互补配对,从而将其携带的氨基酸掺入到新合成的肽链中,对遗传信息的精准传递具有重要作用^([1]).tRNA上存在着大量的转录后核苷酸修饰^([2]).目前已有120多种tRNA修饰被鉴定出来,它们存在于12%~20%的tRNA核苷酸碱基上^([3,4]).tRNA上存在的这些修饰参与并调控一系列生命过程^([5]).众多的修饰能够维持tRNA结构、提高tRNA稳定性、促进反密码子与密码子的精确配对等,在多种生命活动中发挥作用^([6,7]).

关键词：反密码子三联体密码生物大分子修饰酶 tRNA 遗传信息传递底物特异性 ).

线粒体RNAm^(3)C修饰酶METTL8的活力依赖于亚型特异的N-末端延伸结构且METTL8能修饰多种非天然底物tRNA

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科学通报:英文版 2023年第18期68卷 2094-2105,M0004页

作者：黄梦涵王金涛张剑辉毛雪玲彭桂鑫林秀颖吕岱竹袁晨林桓王恩多周小龙 Key Laboratory of RNA Science and Engineering State Key Laboratory of Molecular BiologyCAS Center for Excellence in Molecular Cell ScienceShanghai Institute of Biochemistry and Cell BiologyChinese Academy of SciencesUniversity of Chinese Academy of SciencesShanghai 200031China Key Laboratory of Systems Health Science of Zhejiang Province School of Life ScienceHangzhou Institute for Advanced StudyUniversity of Chinese Academy of SciencesHangzhou 310024China School of Life Science and Technology ShanghaiTech UniversityShanghai 201210China State Key Laboratory of Marine Resource Utilization in South China Sea Hainan UniversityHaikou 570228China School of Life Sciences Hainan UniversityHaikou 570228China Analysis and Testing Center Chinese Academy of Tropical Agricultural SciencesHaikou 571101China School of Pharmacy China Pharmaceutical UniversityNanjing 211198China

定位于线粒体的METTL8-Iso1和分布在核仁的METTL8-Iso4均为甲基转移酶基因METTL8通过mRNA选择性剪接产生的亚型,它们的区别仅为前者有N-末端延伸结构.METTL8-Iso1的功能是催化产生线粒体tRNATh和tRNASe(UCN)第32位3-甲基胞苷(mC32)修饰.... 详细信息

定位于线粒体的METTL8-Iso1和分布在核仁的METTL8-Iso4均为甲基转移酶基因METTL8通过mRNA选择性剪接产生的亚型,它们的区别仅为前者有N-末端延伸结构.METTL8-Iso1的功能是催化产生线粒体tRNATh和tRNASe(UCN)第32位3-甲基胞苷(mC32)修饰.METTL8-Iso4是否也具有tRNAmC32修饰活性以及N-末端延伸在线粒体tRNAmC32修饰中的作用尚不清楚。我们发现,N-末端延伸上有几个保守的关键氨基酸残基,而METTL8-Iso4由于缺少N-未端延伸而不具备tRNAmC32修饰活力.体外实验和体内实验的结果表明,甲基转移酶METTL2A和Trm140上对应的这些关键位点也是各自对底物tRNAmC32修饰活性所必需的。在跨细胞区室与跨物种的酶对tRNA体外催化的交又实验中,我们意外地发现METTL8-Iso1也能催化几种人细胞质tRNA甚至大肠杆菌tRNA的mC32修饰。此外,mC32修饰并不影响tRNA的N-苏氨酰氨基甲酰腺苷(t'A)修饰和氨基酰化活力.除了METTL8与人线粒体丝氨酰-tRNA合成酶(SARS2)的相互作用外,我们还进一步发现了人线粒体苏氨酰-tRNA合成酶(TARS2)与METTL8-IsoI之间的相互作用。体外实验中,METTL8-Iso1显著促进了SARS2和TARS2的氨酰化活力,表明线粒体IRNA的修饰和氨基酰化之间存在功能联系:总之,该结果加深了对线粒体mC32修饰生物发生机制的理解,并提供了一种制备仅含mC32修饰的人细胞质或细菌tRNA的方法,有助于未来研究mC32修饰对tRNA结构和功能的影响。

关键词： tRNA 3-methylcytidine Methyltransferase Aminoacylation

The occurrence order and cross-talk of different tRNA modifications

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Science China(Life Sciences) 2021年第9期64卷 1423-1436页

作者： Jing Li Wen-Yu Zhu Wen-Qing Yang Cai-Tao Li Ru-Juan Liu School of Life Science and Technology ShanghaiTech UniversityShanghai 201210China

Chemical modifications expand the composition of RNA molecules from four standard nucleosides to over 160 modified nucleosides,which greatly increase the complexity and utility of RNAs.Transfer RNAs(tRNAs)are the most heavily modified cellular RNA molecules and contain the largest variety of modifications.Modification of tRNAs is pivotal for protein synthesis and also precisely regulates the noncanonical functions of tRNAs.Defects in tRNA modifications lead to numerous human diseases.Up to now,more than 100 types of modifications have been found in tRNAs.Intriguingly,some modifications occur widely on all tRNAs,while others only occur on a subgroup of tRNAs or even only a specific tRNA.The modification frequency of each tRNA is approximately 7% to 25%,with 5-20 modification sites present on each tRNA.The occurrence and modulation of tRNA modifications are specifically noticeable as plenty of interplays among different sites and modifications have been discovered.In particular,tRNA modifications are responsive to environmental changes,indicating their dynamic and highly organized nature.In this review,we summarized the known occurrence order,cross-talk,and cooperativity of tRNA modifications.

关键词： tRNA RNA modification RNA methylation occurrence order cross-talk

Extensive profiling of the expressions of tRNAs and tRNA-derived fragments(tRFs)reveals the complexities of tRNA and tRF populations in plants

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Science China(Life Sciences) 2021年第4期64卷 495-511页

作者： Xuan Ma Chunyan Liu Xiaocong Kong Jing Liu Siju Zhang Shanshan Liang Weijiang Luan Xiaofeng Cao Tianjin Key Laboratory of Animal and Plant Resistance College of Life SciencesTianjin Normal UniversinTianjin 300387China State Key Laboratory of Plant Genomics and National Center for Plant Gene Research Institute of Genetics and Developmental BiologyChinese Academy of Sciences.Beijing 100101China

Evidence is emerging that t RNA-derived fragments(t RFs)are regulatory molecules.Studies of t RFs in plants have been based on conventional small RNA sequencing,and focused on profiling of t RF-5 and t RF-3 species.A more comprehensive and quantitative analysis of the entire t RF population is highly necessary.Here,we employ t RNA-seq and YAMAT-seq,and develop a bioinformatics tool to comprehensively profile the expressions of t RNAs and t RFs in plants.We show that in Arabidopsis,approximately half of t RNA genes are extremely weakly expressed,accounting for only 1%of total t RNA abundance,while~12%of t RNA genes contribute to~80%of t RNA abundance.Our t RNA sequencings in various plants reveal that t RNA expression profiles exhibit a cross-species conserved pattern.By characterizing the composition of a highly heterogeneous t RF population,we show that t RNA halves and previously unnoticed 10–16-nt tiny t RFs represent substantial portions.The highly accumulated 13-nt and 16-nt tiny t RFs in Arabidopsis indicate that tiny t RFs are not random t RNA degradation products.Finally,we provide a user-friendly database for displaying the dynamic spatiotemporal expressions of t RNAs and t RFs in the model plants Arabidopsis and rice.

关键词： tRNA tRFs tRNA halves tiny tRFs tRNA-seq

tRNA-sgRNA/Cas9系统介导多年生黑麦草原生质体的基因编辑

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草业学报 2023年第4期32卷 129-141页

作者：姚佳明郝欢欢张敬徐彬南京农业大学草业学院江苏南京210095

tRNA可以将多个sgRNAs连接起来合并成一个多顺反子基因,再与CRISPR/Cas9表达载体结合形成多顺反子tRNA-sgRNA/Cas9(PTG/Cas9)系统来对多靶点进行基因编辑。该系统已经在水稻中验证其可促进sgRNAs的转录和提高多靶点的编辑效率。因此,为... 详细信息

tRNA可以将多个sgRNAs连接起来合并成一个多顺反子基因,再与CRISPR/Cas9表达载体结合形成多顺反子tRNA-sgRNA/Cas9(PTG/Cas9)系统来对多靶点进行基因编辑。该系统已经在水稻中验证其可促进sgRNAs的转录和提高多靶点的编辑效率。因此,为了在多年生黑麦草中实现高效的基因编辑,本研究中,构建了2个带有tRNA的CRISPR中间载体,并提供了一种快速、灵活的PTG/Cas9载体构建方法。为了快速验证PTG/Cas9系统是否可以在多年生黑麦草基因组中发挥功能,用聚乙二醇4000(PEG 4000)介导PTG/Cas9质粒转化多年生黑麦草原生质体,后提取原生质体DNA扩增目的序列检测是否有目标基因被编辑的细胞。试验结果显示,PTG/Cas9系统可用于多年生黑麦草基因编辑,且基因编辑效率约为6.7%。试验结果为多年生黑麦草遗传研究和育种提供了重要的基础。

关键词： CRISPR/Cas9 tRNA 多基因编辑载体质粒原生质体多年生黑麦草

tRNA cleavage: a new insight

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Neural Regeneration Research 2020年第1期15卷 47-52页

作者： Sherif Rashad Kuniyasu Niizuma Teiji Tominaga Department of Neurosurgery Tohoku University Graduate School of MedicineSendaiJapan Department of Neurosurgical Engineering and Translational Neuroscience Tohoku University Graduate School of MedicineSendaiJapan Department of Neurosurgical Engineering and Translational Neuroscience Graduate School of Biomedical EngineeringTohoku UniversitySendaiJapan

Over the past decades,tRNA was found to be a rich hub of RNA modifications such as 1-methyladenosine and 5-methycytosine modifications and others,holding more than half of all modifications occurring in RNA molecules.Moreover,tRNA was discovered to be a source of various small noncoding RNA species,such as the stress induced angiogenin cleaved tRNA halves(tiRNA)or the miRNA like tRNA derived fragments.tRNA cleavage under stress was fist discovered in bacteria and later was found to be conserved across different species,including mammals.Under cellular stress conditions,tRNA undergoes conformational changes and angiogenin cleaves it into 3′and 5′halves.5′tiRNA halves were shown to repress protein translations.tRNA cleavage is thought of to be a cytoprotective mechanism by which cells evade apoptosis,however some data hints to the opposite;that tiRNA are cytotoxic or at least related to apoptosis initiation.tRNA cleavage also was shown to be affected by tRNA modifications via different enzymes in the cytosol and mitochondria.In this review,we will highlight the biology of tRNA cleavage,show the evidence of it being cytoprotective or a marker of cell death and shed a light on its role in disease models and human diseases as well as possible future directions in this field of RNA research.

关键词： angiogenin apoptosis cell stress RNA modification stress granules stroke tiRNA translation repression tRNA tRNA cleavage

Hearing impairment-associated KARS mutations lead to defects in aminoacylation of both cytoplasmic and mitochondrial tRNA^(Lys)

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Science China(Life Sciences) 2020年第8期63卷 1227-1239页

作者： Yong Wang Jing-Bo Zhou Qi-Yu Zeng Siqi Wu Mei-Qin Xue Pengfei Fang En-Duo Wang Xiao-Long Zhou State Key Laboratory of Molecular Biology CAS Center for Excellence in Molecular Cell ScienceShanghai Institute of Biochemistry and Cell BiologyChinese Academy of SciencesUniversity of Chinese Academy of SciencesShanghai 200031China School of Life Science and Technology ShanghaiTech UniversityShanghai 201210China State Key Laboratory of Bioorganic and Natural Products Chemistry Center for Excellence in Molecular SynthesisShanghai Institute of Organic ChemistryChinese Academy of SciencesShanghai 200032China

Aminoacyl-tRNA synthetases(aaRSs)are ubiquitously expressed,essential enzymes,synthesizing aminoacyl-tRNAs for protein synthesis.Functional defects of aaRSs frequently cause various human disorders.Human KARS encodes both cytosolic and mitochondrial lysyl-tRNA synthetases(LysRSs).Previously,two mutations(c.1129 G>A and c.517 T>C)were identified that led to hearing impairment;however,the underlying biochemical mechanism is unclear.In the present study,we found that the two mutations have no impact on the incorporation of LysRS into the multiple-synthetase complex in the cytosol,but affect the cytosolic LysRS level,its tertiary structure,and cytosolic tRNA aminoacylation in vitro.As for mitochondrial translation,the two mutations have little effect on the steady-state level,mitochondrial targeting,and tRNA binding affinity of mitochondrial LysRS.However,they exhibit striking differences in charging mitochondrial tRNALys,with the c.517T>C mutant being completely deficient in vitro and in vivo.We constructed two yeast genetic models,which are powerful tools to test the in vivo aminoacylation activity of KARS mutations at both the cytosolic and mitochondrial levels.Overall,our data provided biochemical insights into the potentially molecular pathological mechanism of KARS c.1129G>A and c.517T>C mutations and provided yeast genetic bases to investigate other KARS mutations in the future.

关键词： aminoacyl-tRNA synthetase tRNA aminoacylation protein synthesis