Table of Content

10 October 2022, Volume 43 Issue Album-4

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Article

Effects of Exogenous N⁶-methyladenosine (m⁶A) Incorporation on the Expression of Cellular mRNA Transcripts

ZHANG Qingyi, CAO Jie, SHU Xiao, LIU Jianzhao

2022, 43(Album-4):  20220173.  doi:10.7503/cjcu20220173

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N6-methyladenosine (m6A) is the most prevalent and reversible internal mRNA modification in eukaryotic cells, and plays an essential role in RNA fate determination. In this work, we studied the exogenous incorporation of m6A into cellular mRNA through salvage pathway and evaluated its resultant biological effects. First, we found that feeding of HeLa cells with m6A nucleoside significantly altered cell morphology and viability. We then synthesized isotope-labelled d3-m6A (N6-methyl-d3-adenosine) and examined the d3-m6A incorporation rate in cellular mRNA along with incubation time using mass spectrometry. Next, RNA sequencing (RNA-seq) was used to study the biological effects of exogenous m6A incorporation. Finally, thousands of genes were found to be differentially expressed, and were dramatically enriched in pathways such as ribosome biogenesis, mRNA metabolic process and cell morphogenesis differentiation. All these results suggested that exogenous m6A could be incorporated into mRNAs through a metabolic pathway to influence cellular gene expression.



Review

Progress on the Stereocontrolled Synthesis of Phosphorothioate Oligonucleotides

CAO Shujie, LI Hongjun, GUAN Wenli, REN Mengtian, ZHOU Chuanzheng

2022, 43(Album-4):  20220304.  doi:10.7503/cjcu20220304

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Phosphorothioate (PS) oligonucleotides have found a wide range of applications in biochemical mechanism studies, biomedicine, materials and related fields. The stereo configuration of PS has a remarkable effect on the biochemical properties of PS nucleic acids, which has aroused wide interest in developing methods for efficient and stereoselective synthesis of PS oligonucleotides in the past 30 years. This review summarizes the methods for stereocontrolled synthesis of PS oligonucleotides, with a focus on the research progress in the past decade. The advantages and disadvantages of different methods are comparatively analyzed. Finally, the prospect of stereocontrolled synthesis of PS oligonucleotides is briefly discussed.



Application of Nanopore Sequencing Technology in the Detection of Nucleic Acid Modifications

CHEN Jialu, HUANG Shuo

2022, 43(Album-4):  20220333.  doi:10.7503/cjcu20220333

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A wide variety of chemical modifications have been found in DNA and RNA. These nucleic acid modifications play critical roles in the regulation of gene expression, affect biological processes such as growth and development, and cause diseases including cancer. Accurate identification and localization of nucleic acid modifications can help to understand their functional mechanism, and contributes to diagnosis and treatment of relevant diseases. Nanopore sequencing is an emerging single-molecule sequencing technology and can directly detect nucleic acid modifications. It can simultaneously detect multiple modifications based on the difference in pore blockage signals between modified and canonical bases. This review briefly summarizes the principle and recent development of nanopore sequencing, computer algorithms for nucleic acid modification identification and applications of nanopore sequencing. By the end of this review, prospects of future development of nanopore sequencing is also proposed.



Research Progress on Chemical Intervention of N⁶-methyladenosine Modification

HUAN Xinyu, LAI Ganqiang, HUANG Yue, YANG Caiguang

2022, 43(Album-4):  20220340.  doi:10.7503/cjcu20220340

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There are multiple modifications on mRNA, which includes N6-methyladenosine (m6A), N1-methyladenosine (m1A), 5-methylcytosine (m5C), etc. As the most common modification on mRNA, m6A has an impact on the processing of the 5’ends and 3’ ends of mRNA, as well as the procedure of localization, degradation, and translation. It also regulates the expression of genes at the post-transcriptional level and participates in manifold physiological activities. This paper makes a review of the mechanisms on the molecular level of the m6A modification and the relationship between the modification and various diseases. Then it outlines the development course of the m6A identification technology, emphasizes on the latest research progress on m6A chemical intervention in order to enable the readers to form a comprehensive understanding of m6A modification and provide references for the development of small-molecular drugs targeting on m6A modification.



Non-viral delivery of CRISPR/Cas9 Genome Editing

Jinhan Sheng, Qizhen Zheng, Ming Wang

2022, 43(Album-4):  20220344.  doi:10.7503/cjcu20220344

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Genome editing based on the clustered regularly interspaced short palindromic repeats (CRISPR/Cas9) provides an efficient and rapid tool for precise control and regulation of mammalian cell genome. Its chemical biology and biomedical application is however challenged by the delivery of the CRISPR/Cas9 gene editing tool that composed of Cas9 protein and gRNA into mammalian cells. In recent years, a large variety of non-viral delivery vectors have been designed for CRISPR/Cas9 gene editing delivery in the form of DNA, messenger RNA (mRNA) and ribonucleoprotein complex (RNP). In this article, we summarize the most recent progress of non-viral CRISPR/Cas9 genome editing tool, and the application prospect of CRISPR/Cas9 genome editing technology in chemical biology.



Application of Gene Editing in Mitochondrial Diseases

CHANG Liying, LING Xinyu, CHEN Heqi, WANG Xue, LIU Tao

2022, 43(Album-4):  20220363.  doi:10.7503/cjcu20220363

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Mitochondria, as energy factories of cells, play an important role in maintaining cell energy metabolism and human life activities. The mutation of mitochondrial genome has led to a series of mitochondrial diseases, which seriously threaten human life and health. The development of gene editing methods targeting mitochondria is of great significance for the treatment of mitochondrial diseases. In recent years, a series of gene editing methods including restriction nuclease, zinc finger nuclease, transcriptional activator like effector nuclease, CRISPR and mitochondrial base editor have been developed. This article reviews the recent progress, deficiencies and development direction of the application of gene editing tools for mitochondria gene editing. We hope it could provide reference value for the development of mitochondrial disease treatment technology.