Sequencing methods and functional decoding of mRNA modifications

More than 160 types of post-transcriptional RNA modifications have been reported;there is substantial variation in modification type,abundance,site,and function across species,tissues,and RNA type.The recent development of high-throughput detection technology has enabled identification of diverse dynamic and reversible RNA modifications,including N6,2′-O-dimethyladenosine(m6Am),N1-methyladenosine(m1A),5-methylcytosine(m5C),N6-methyladenosine(m6A),pseudouridine(Ψ),and inosine(I).In this review,we focus on eukaryotic mRNA modifications.We summarize their biogenesis,regulatory mechanisms,and biological functions,as well as highthroughput methods for detection of mRNA modifications.We also discuss challenges that must be addressed in mRNA modification research.

不同吸附基团的减水剂对水泥浆体塑性黏度的影响

为揭示主链不同吸附基团对水泥吸附与分散和水泥浆体流变性能的影响,分别以单羧酸吸附基团的丙烯酸(AA)、双羧酸吸附基团的马来酸酐(MA)、磷酸吸附基团的异丙烯膦酸(IPPA)与异戊烯醇聚氧乙烯-聚氧丙烯醚(IPE_(12)PP_(3)G,Mw=762)进行自由基聚合反应制备了酸醚比、主链聚合度、侧链长度和侧链相同,但吸附基团不同的3种减水剂,并对其性能进行凝胶渗透色谱、红外光谱和核磁共振氢谱的分析获得了合成产物的相对分子质量及其分布等结构信息。同时测试了合成减水剂的络合能力、流体力学半径、水泥颗粒表面的吸附量、Zeta电位、吸附层厚度和水泥水化放热速率。结果显示,双羧酸吸附基团的马来酸酐合成的减水剂表现出更强的络合能力(Ca2p结合能与空白样发生化学位移分别为0.17 eV和0.07eV)、更小的流体力学半径(5.46 nm)、更大的吸附量(2.65 mN/m^(2))、Zeta电位(-26.507 mV)和吸附层厚度(1.16 nm),水泥水化放热速也相对最慢。

Gene Network Analysis and Functional Studies of Senescence-associated Genes Reveal Novel Regulators of Arabidopsis Leaf Senescence

Plant leaf senescence has been recognized as the last phase of plant development, a highly ordered process regulated by genes known as senescence associated genes （SAGs）. However, the function of most of SAGs in regulating leaf senescence as well as regulators of those functionally known SAGs are still unclear. We have previously developed a curated database of genes potentially associated with leaf senescence, the Leaf Senescence Database （LSD）. In this study, we built gene networks to identify common regulators of leaf senescence in Arabidopsis thaliana using promoting or delaying senescence genes in LSD. Our results demonstrated that plant hormones cytokinin, auxin, nitric oxide as well as small molecules, such as Ca2＋, delay leaf senescence. By contrast, ethylene, ABA, SA and JA as well as small molecules, such as oxygen, promote leaf senescence, altogether supporting the idea that phytohormones play a critical role in regulating leaf senescence. Functional analysis of candidate SAGs in LSD revealed that a WRKY transcription factor WRKY75 and a Cys2/His2-type transcription factor AZF2 are positive regulators of leaf senescence and loss-of-function of WRKY75 or AZF2 delayed leaf senescence. We also found that silencing of a protein phosphatase, AtMKP2, promoted early senescence. Collectively, LSD can serve as a comprehensive resource for systematic study of the molecular mechanism of leaf senescence as well as offer candidate genes for functional analyses.

Epitranscriptomics:Toward A Better Understanding of RNA Modifications

Ever since the first RNA nucleoside modification was charac- terized in 1957 [1], over 100 distinct chemical modifications have been identified in RNA to date [2]. Most of these modi- fications were characterized in non-coding RNAs （ncRNAs）, including tRNA, rRNA, and small nuclear RNA （snRNA） [3]. Studies in the past few decades have located various mod- ifications in these ncRNAs and revealed their functional roles [3]. For instance, NLmethyladenosine （mlA）, which is typically found at position 58 in the tRNA T-loop of eukaryotes, func- tions to stabilize tRNA tertiary structure [4] and affect transla- tion by regulating the associations between tRNA and polysome [5]. Pseudouridine （tp） in snRNA can fine-tune branch site interactions and affect mRNA splicing [6].

Epitranscriptomic technologies and analyses

RNA can interact with RNA-binding proteins(RBPs),mRNA,or other non-coding RNAs(ncRNAs)to form complex regulatory networks.High-throughput CLIP-seq,degradome-seq,and RNA-RNA interactome sequencing methods represent powerful approaches to identify biologically relevant ncRNA-target and protein-ncRNA interactions.However,assigning ncRNAs to their regulatory target genes or interacting RNA-binding proteins(RBPs)remains technically challenging.Chemical modifications to mRNA also play important roles in regulating gene expression.Investigation of the functional roles of these modifications relies highly on the detection methods used.RNA structure is also critical at nearly every step of the RNA life cycle.In this review,we summarize recent advances and limitations in CLIP technologies and discuss the computational challenges of and bioinformatics tools used for decoding the functions and regulatory networks of ncRNAs.We also summarize methods used to detect RNA modifications and to probe RNA structure.

Single-base resolution analysis of DNA epigenome via high-throughput sequencing

Epigenetic changes caused by DNA methylation and histone modifications play important roles in the regulation of various cellular processes and development. Recent discoveries of 5-methylcytosine(5m C) oxidation derivatives including 5-hydroxymethylcytosine(5hm C), 5-formylcytsine(5f C) and 5-carboxycytosine(5ca C) in mammalian genome further expand our understanding of the epigenetic regulation. Analysis of DNA modification patterns relies increasingly on sequencing-based profiling methods. A number of different approaches have been established to map the DNA epigenomes with single-base resolution, as represented by the bisulfite-based methods, such as classical bisulfite sequencing(BS-seq), TAB-seq(TET-assisted bisulfite sequencing), ox BS-seq(oxidative bisulfite sequencing) and etc. These methods have been used to generate base-resolution maps of 5m C and its oxidation derivatives in genomic samples. The focus of this review will be to discuss the chemical methodologies that have been developed to detect the cytosine derivatives in the genomic DNA.

Liquid biopsies:DNA methylation analyses in circulating cell-free DNA

Analysis of patient＇s materials like cells or nucleic acids obtained in a minimally invasive or noninvasive manner through the sampling of blood or other body fluids serves as liquid biopsies, which has huge potential for numerous diagnostic applications. Circulating cell-free DNA（cfDNA） is explored as a prognostic or predictive marker of liquid biopsies with the improvements in genomic and molecular methods. DNA methylation is an important epigenetic marker known to affect gene expression. cfDNA methylation detection is a very promising approach as abnormal distribution of DNA methylation is one of the hallmarks of many cancers and methylation changes occur early during carcinogenesis. This re？view summarizes the various investigational applications of cfDNA methylation and its oxidized de？rivatives as biomarkers for cancer diagnosis, prenatal diagnosis and organ transplantation monitoring.The review also provides a brief overview of the technologies for cfDNA methylation analysis based on next generation sequencing.

Metatranscriptomic analysis of host response and vaginal microbiome of patients with severe COVID-19

Dear Editor,The pandemic of coronavirus disease 2019(COVID-19)caused by severe acute respiratory syndrome coronavirus 2(SARS-Co V-2)has become a global public health threat.Here we use a TRACE-seq-based metatranscriptomic analysis to compare host responses and vaginal microbiome of postmenopausal female patients with underlying severe COVID-19 disease with those of healthy females,thereby providing insights into the changes in the microenvironment of women's reproductive system.