Pan-cancer analysis of RNA 5-methylcytosine reader (ALYREF)

The incre asing interest in RNA modifications has signifcantly advanced epigenomic and epitranscriptomic technologies.This study focuses on the immuno oncological impact of ALYREF in human cancer through a pan-cancer analysis,enhancing understanding of this gene's role in cancer.We observed differential ALYREF expression between tumor and normal samples,correl ating strongly with prognosis in various cancers,particularly kidney renal papillary cell carcinoma(KIRP)and liver hepatocellular carcinoma(LIHC).ALYREF showed a negative correlation with most tumor-infitrating cells in lung squamous cell carcinoma(LUSC)and lymphoid neoplasm difuse large B-cell lymphoma(DLBC),while positive correlations were noted in IIHC,kidney chromophobe(KICH),mesothelioma(MESO),KIRP,pheochromocytoma and paraganglioma(PARD),and glioma(GBMLGG).Aditionally,ALYREF expression was closely associated with tumor heterogeneity,stemness indices,and a high mutation rate in TP53 across these cancers.In conclusion,ALYREF may serve as an oncogenic biomarker in numerous cancers,meriting further research attention.

hSOD1-G93A突变的肌萎缩侧索硬化症转基因小鼠脊髓中5-mC水平以及DNMT1、DNMT3A和DNMT3B表达的变化

目的探讨5-甲基胞嘧啶(5-mC)水平以及DNA甲基转移酶1、3A和3B(DNMT1、DNMT3A和DNMT3B)蛋白表达变化与肌萎缩侧索硬化症(amyotrophic lateral sclerosis,ALS)之间的关系。方法选用发病前期(70 d)、发病早期(95 d)、发病中期(108 d)和发病晚期(122 d)的hSOD1-G93A突变的ALS转基因小鼠,以同窝野生型(wild type,WT)小鼠作为对照,应用ELISA和Dot blot技术检测ALS小鼠脊髓中5-mC水平的变化规律,采用Western blot和免疫荧光双标染色技术检测ALS小鼠不同时期的脊髓中DNMT1、DNMT3A、DNMT3B的蛋白水平变化以及定位。结果ELISA和Dot blot结果显示,与WT小鼠相比,ALS转基因小鼠脊髓中5-mC水平升高;Western blot分析显示,与WT小鼠相比,ALS转基因小鼠脊髓中DNMT1蛋白水平在70 d无显著性变化,在95 d、108 d和122 d显著升高;DNMT3A蛋白水平在70 d、95 d和108 d升高,在122 d无显著性变化;DNMT3B蛋白水平在70 d无显著性变化,在95 d、108 d和122 d显著升高。结论ALS转基因小鼠脊髓中5-mC水平及关键DNA甲基转移酶DNMT1、DNMT3A和DNMT3B的蛋白水平异常升高与ALS发病密切相关。

Regulation of RNA methylation and immune infiltration patterns by m5C regulators in head and neck squamous cell carcinoma

5-Methylcytosine(m5C)methylation contributes to the development and progression of various malignant tumors.This study aimed to explore the potential role of m5C methylation regulators(m5CMRs)in head and neck squamous cell carcinoma(HNSCC).Methods:The transcription data of HNSCC samples were obtained from The Cancer Genome Atlas(TCGA)and the Gene Expression Omnibus(GEO)databases.Subsequently,the m5C patterns in HNSCC were evaluated based on 14 m5CMRs.Then,the m5Cscore was developed to quantify m5C patterns by using principal component analysis(PCA)algorithms.Two single-cell RNA sequencing datasets and various methods were employed to assess the prognostic value and sensitivity to immunotherapy.Finally,key prognostic m5CMRs were identified using univariate COX regression analysis,and their clinical significance was validated based on the Human Protein Atlas(HPA)database and by using immunohistochemistry.Results:Two distinct m5C clusters were identified.m5C cluster A is characterized by an immune-activated microenvironment and is associated with a favorable prognosis.Notable differences were observed in prognosis,immune infiltration,and immunotherapy response between the high-and low-m5Cscore groups.Patients in the high-m5Cscore group exhibited high TMB,which is correlated with poor prognosis.The m5Cscore of epithelial cells in HNSCC was higher than that in other cells.Key prognostic m5CMRs,including NSUN2,DNMT3B,ALKBH1,and Y-Box Binding Protein 1(YBX1),were associated with poor prognosis.Conclusion:Our research indicates that in head and neck squamous cell carcinoma,the m5C modification profoundly affects the TME’s diversity and complexity,influencing prognosis and the success of immunotherapy.Targeting m5C regulatory elements may be a new method for enhancing the efficacy of immunotherapy in HNSCC.

Expression of TET and 5-HmC in Trophoblast Villi of Women with Normal Pregnancy and with Early Pregnancy Loss

Increasing evidence suggests that epigenetic dysfunction may influence the stability of normal pregnancy. The ten-eleven translocation （TET） family and 5-hydroxymethylcytosine （5-hmC） were found to be linked with epigenetic reprogramming. The present study aimed to examine the expression of the TET family and 5-hmC in the villi of human embryos and compared their expression between normal pregnancy and early pregnancy loss （EPL）. Embryonic villi were collected from normal pregnant women （control） experiencing medical abortion and from EPL patients at gestation ages of 6, 7 and 8 weeks. The mRNAs of TET family were analysed using quantitative polymerase chain reaction （qPCR）, and TET proteins using Western blotting and immunohistochemical analysis. The MethylFlashTM Kit was used to quantify the absolute amount of 5-methylcytosine （5-mC） and 5-hmC. Our results showed that the expression of the TETs and 5-hmC in the normal villus decreased with increasing gestational age. Immunohistochemistry revealed that the TET proteins were expressed in the cytoplasm of trophoblasts and their expression was the highest in the 6-week tissue samples, which was consistent with the qPCR and Western blot results. The expression of TET1, TET2, and TET3 was lower in the villi in EPL group than in normal pregnancy group （P〈0.05 for all）. It was concluded that the TET family and 5-hmC are critical in epigenetic reprogramming of human embryo. The findings also suggest that a deficiency of TETs in the villus might be associated with human EPL.

Dysregulation of RNA modification systems in clinical populations with neurocognitive disorders

The study of modified RNA known as epitranscriptomics has become increasingly relevant in our understanding of disease-modifying mechanisms.Methylation of N6 adenosine(m^(6)A)and C5 cytosine(m^(5)C)bases occur on mRNAs,tRNA,mt-tRNA,and rRNA species as well as non-coding RNAs.With emerging knowledge of RNA binding proteins that act as writer,reader,and eraser effector proteins,comes a new understanding of physiological processes controlled by these systems.Such processes when spatiotemporally disrupted within cellular nanodomains in highly specialized tissues such as the brain,give rise to different forms of disease.In this review,we discuss accumulating evidence that changes in the m^(6)A and m^(5)C methylation systems contribute to neurocognitive disorders.Early studies first identified mutations within FMR1 to cause intellectual disability Fragile X syndromes several years before FMR1 was identified as an m^(6)A RNA reader protein.Subsequently,familial mutations within the m^(6)A writer gene METTL5,m^(5)C writer genes NSUN2,NSUN3,NSUN5,and NSUN6,as well as THOC2 and THOC6 that form a protein complex with the m^(5)C reader protein ALYREF,were recognized to cause intellectual development disorders.Similarly,differences in expression of the m^(5)C writer and reader effector proteins,NSUN6,NSUN7,and ALYREF in brain tissue are indicated in individuals with Alzheimer's disease,individuals with a high neuropathological load or have suffered traumatic brain injury.Likewise,an abundance of m^(6)A reader and anti-reader proteins are reported to change across brain regions in Lewy bodies diseases,Alzheimer's disease,and individuals with high cognitive reserve.m^(6)A-modified RNAs are also reported significantly more abundant in dementia with Lewy bodies brain tissue but significantly reduced in Parkinson's disease tissue,whilst modified RNAs are misplaced within diseased cells,particularly where synapses are located.In parahippocampal brain tissue,m^(6)A modification is enriched in transcripts associated with psychiatric disorders including conditions with clear cognitive deficits.These findings indicate a diverse set of molecular mechanisms are influenced by RNA methylation systems that can cause neuronal and synaptic dysfunction underlying neurocognitive disorders.Targeting these RNA modification systems brings new prospects for neural regenerative therapies.

当归多糖对崇明白山羊冷冻精液质量与DNA甲基化的影响

为了解决崇明白山羊精液冷冻后总体质量偏低的问题,试验采集8只24~26月龄体型匀称的崇明白山羊精液,检测合格后混匀以消除个体差异,分别用未添加当归多糖(ASP)和添加150,300,600,1200μg/mL ASP的基础稀释液稀释后进行冷冻,解冻后评估精液的质量参数(活率、活力、质膜完整率、顶体完整率、线粒体活性)、相关抗氧化指标[丙二醛(MDA)、活性氧(ROS)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GPH-Px)、一氧化氮(NO)、总抗氧化能力(T-AOC)、过氧化氢酶(CAT)]及DNA甲基化5-MC水平,最终确定ASP的最适宜添加浓度,并分析了精液DNA甲基化5-MC水平与精液质量参数的相关性。结果表明:与未添加ASP相比,添加600μg/mL ASP的精子活率、活力、质膜完整率和线粒体活性都显著升高(P<0.05);添加300μg/mL ASP的精子顶体完整率显著升高(P<0.05);添加600μg/mL ASP的精子SOD、CAT活性及T-AOC水平都显著升高(P<0.05);添加600μg/mL ASP的精子ROS、MDA含量都显著降低(P<0.05);添加300,600μg/mL ASP的精子DNA甲基化5-MC水平显著降低(P<0.05)。精液DNA甲基化5-MC水平与精子活力、活率、质膜完整率、顶体完整率、线粒体活性均呈现极显著负相关(P<0.01),R分别为-0.983,-0.892,-0.992,-0.732,-0.825。说明在本试验条件下,基础稀释液中添加600μg/mL ASP最利于崇明白山羊精液的冷冻保存。

5-Methylcytosine RNA Methylation in Arabidopsis Thaliana

5-Methylcytosine （m^5C） is a well-characterized DNA modification, and is also predominantly reported in abundant non-coding RNAs in both prokaryotes and eukaryotes. However, the distribution and biological functions of m^5C in plant mRNAs remain largely unknown. Here, we report transcriptome-wide profiling of RNA m^5C in Arabidopsis thaliana by applying m^5C RNA immunoprecipitation followed by a deep- sequencing approach （m^5C-RIP-seq）. LC-MS/MS and dot blot analyses reveal a dynamic pattern of m^5C mRNA modification in various tissues and at different developmental stages, m^5C-RIP-seq analysis identified 6045 m^5C peaks in 4465 expressed genes in young seedlings. We found that m^5C is enriched in coding sequences with two peaks located immediately after start codons and before stop codons, and is associated with mRNAs with low translation activity. We further demonstrated that an RNA （cytosine-5）-methyl- transferase, tRNA-specific methyltransferase 4B （TRM4B）, exhibits m^5C RNA methyltransferase activity. Mutations in TRM4B display defects in root development and decreased m^5C peaks. TRM4B affects the transcript levels of the genes involved in root development, which is positively correlated with their mRNA stability and m^5C levels. Our results suggest that m^5C in mRNA is a new epitranscriptome marker inArabidopsis, and that regulation of this modification is an integral part of gene regulatory networks underlying plant development.

Global changes of 5-hydroxymethylcytosine and 5-methylcytosine from normal to tumor tissues are associated with carcinogenesis and prognosis in colorectal cancer

Aberrant DNA methylation has raised widespread attention in tumorigenesis. In this study, we aimed to investigate the changes of global DNA methylation and hydroxymethylation from normal to tumor tissues in colorectal cancer(CRC) and their association with the prognosis. The levels of genomic 5-hydroxymethylcytosine(5hmC) and 5-methylcytosine(5mC) in cancerous tissues were significantly lower than those in corresponding adjacent normal tissues. The genomic levels of 5mC were significantly positively correlated with 5hmC in normal and cancerous tissues(all P<0.05). The ratio of 5mC in cancerous tissues to matched normal tissues(C/N-5mC) was also significantly positively correlated with the ratio of 5hmC in cancerous tissues to matched normal tissues(C/N-5hmC)(P=0.01). The 5mC levels and C/N-5mC ratios decreased with age(all P<0.05). Higher 5mC and 5hmC levels were found in rectal than in colon tissues(all P<0.05). High levels of 5mC in cancerous tissues and high C/N-5hmC ratios were each associated with lymph node metastasis(all P<0.05). Survival analysis indicated that the C/N-5mC ratio(P=0.04) is an independent protective factor for overall survival. The data showed that patients with a combination of high C/N-5hmC and low C/N-5mC ratios tended to have a worse prognosis(P<0.01). Our findings showed that the C/N-5mC ratio may be an independent prognostic factor for CRC outcome. Patients with both a high C/N-5hmC ratio and a low C/N-5mC ratio exhibited the worst survival, suggesting that 5mC and 5hmC can be used as critical markers in tumorigenesis and prognosis estimation.

NSUN2-mediated mRNA m^(5)C Modification Regulates the Progression of Hepatocellular Carcinoma

RNA modifications affect many biological processes and physiological diseases.The 5-methylcytosine(m^(5)C)modification regulates the progression of multiple tumors.However,its characteristics and functions in hepatocellular carcinoma(HCC)remain largely unknown.Here,we found that HCC tissues had a higher m^(5)C methylation level than the adjacent normal tissues.Transcriptome analysis revealed that the hypermethylated genes mainly participated in the phosphokinase signaling pathways,such as the Ras and PI3K-Akt pathways.The m^(5)C methyltransferase NSUN2 was highly expressed in HCC tissues.Interestingly,the expression of many genes was positively correlated with the expression of NSUN2,including GRB2,RNF115,AATF,ADAM15,RTN3,and HDGF.Real-time PCR assays further revealed that the expression of the mRNAs of GRB2,RNF115,and AATF decreased significantly with the down-regulation of NSUN2 expression in HCC cells.Furthermore,NSUN2 could regulate the cellular sensitivity of HCC cells to sorafenib via modulating the Ras signaling pathway.Moreover,knocking down NSUN2 caused cell cycle arrest.Taken together,our study demonstrates the vital role of NSUN2 in the progression of HCC.

Eucaryotic DNA Methylation and Gene Mutation

5-methylcytosine (m5C) as a rare base exists in eucaryotic genomes, it is a normal constituent of many eucaryotic DNA, whose existence is a character of eucaryotic DNA. In the regular physiological conditions, cytosine residue of eucaryotic DNA is methylated to be popular. Up to the present, many people consider that the m5C may be mutation hotspots by the m5C deamination leading to gene mutation. Our theoretical investigations indicated that the spontaneous mutation caused by the transition of G - C-A - T, in eukaryotic DNA, may be a result caused by the tautomer changing base pairs and may also be caused by other factor actions, however it could not be caused by the deamination of m5C.

长爪沙鼠肝脏基因组DNA甲基化水平检测

目的用间接ELISA法检测长爪沙鼠肝脏基因组DNA整体甲基化的水平。方法选取新生仔鼠6只(仔鼠组),3月龄鼠12只[其中青年对照组6只,高脂饮食诱发非酒精性脂肪性肝病(NAFLD)青年模型组6只],8月龄中老龄鼠6只(中老龄),各组均为雄性,按组采集血清(新生鼠除外)用于总胆固醇(TC)、甘油三酯(TG)、高密度脂蛋白(HDL)、低密度脂蛋白(LDL)检测,同时采集肝脏标本二份,一份作常规HE染色的形态学观察,一份提基因组DNA,用ELISA法试剂盒检测肝脏基因组DNA整体甲基化的水平[即5-甲基胞嘧啶(5-mc)含量]。结果青年模型组鼠及中老龄鼠均出现高脂血症,血清TC及TG均不同程度显著上升,青年对照组鼠血脂正常。病理学检查显示,青年模型组与中老龄组鼠肝脏出现较为明显的脂肪变性,而青年对照组肝脏无异常,新生仔鼠肝血窦中可见大量成堆分布有单核细胞和红细胞。肝脏基因组DNA整体甲基化水平,青年模型组沙鼠>新生仔鼠>老龄鼠>5对照组鼠。结论长爪沙鼠NAFLD及年龄增加引起的脂肪性变程度与其基因组DNA甲基化水平相关,间接ELISA法检测肝脏基因组DNA整体甲基化水平简便、迅速,成本低,或可以作为评价其表观遗传学的一种方法。

A high methionine,low folate and vitamin B_6/B_(12) containing diet can be associated with memory loss by epigenetic silencing of netrin-1

Memory-epigenetics which is the loss of memory due to epigenetic modifications can be due to the silencing of genes involved in cognitive functions and this is the basis of the current study.We hypothesize that a diet containing high methionine and low vitamins can lead to memory impairment by increasing global DNA methylation and therefore,silencing the netrin-1 gene,which encodes the glycoprotein involved in neurogenesis,axonal guidance and maintenance of the synaptic plasticity.Wild type(C57 BL/6 J) mice were fed with a diet containing excess methionine(1.2%),low-folate(0.08 mg/kg),vitamin B_6(0.01 mg/kg),and B_(12)(10.4 mg/kg) for 6 weeks.Mice were examined weekly for the long-term memory function,using a passive avoidance test,which determined loss of fear-motivated long-term memory starting from the fourth week of diet.Similarly,an increase in brain %5-methyl cytosine was observed starting from the 4 th week of diet in mice.Mice fed with a high methionine,low folate and vitamins containing diet showed a decrease in netrin-1 protein expression and an increase in netrin-1 gene promotor methylation,as determined by methylation-sensitive restriction enzyme-polymerase chain reaction analysis.The increase in methylation of netrin-1 gene was validated by high-resolution melting and sequencing analysis.Furthermore,the association of netrin-1 with memory was established by administering netrin that considerably restored long-term fear motivated memory.Taken together,these results suggest that a diet rich in methionine and lacking in folate and vitamin B_6/B_(12) can induce defects in learning and memory.Furthermore,the data indicates that decrease in netrin-1 expression due to hyper-methylation of its gene can be associated with memory loss.The animal procedures were approved by the Institutional Animal Care and Use Committee,University of Louisville,USA(No.A3586-01) on February 2,2018.

Increased 5-hydroxymethylcytosine and Ten-eleven Translocation Protein Expression in Ultraviolet B-irradiated HaCaT Cells

Background： DNA hydroxymethylation refers to a chemical modification process in which 5-methylcytosine （5mC） is catalyzed to 5-hydroxymethylcytosine （5hmC） by ten-eleven translocation （TET） family proteins. Recent studies have revealed that aberrant TETs expression or 5hmC level may play important roles in the occurrence and development of various pathological and physiological processes including cancer and aging. This study aimed to explore the relation between aberrant DNA hydroxymethylation with skin photoaging and to investigate the levels of TETs, 5mC, and 5hmC expression 24 h after 40 mJ/cm^2 and 80 mJ/cm^2 doses of ultraviolet B （UVB） irradiation to HaCaT cells. Methods： To explore whether aberrant DNA hydroxymethylation is also related to skin photoaging, 40 mJ/cm^2 and 80 mJ/cm^2 doses of UVB were chosen to treat keratinocytes （HaCaT cells）. After 24 h of UVB irradiation, 5mC and 5hmC levels were determined by immunohistochemistry （IHC） and immunofluorescence （IF）, and at the same time, the expression levels of matrix metalloproteinase 1 （MMP-1） and TETs were assessed by reverse transcription-polymerase chain reaction or Western blot analysis. Results： After 40 mJ/cm^2 and 80 mJ/cm^2 doses of UVB exposure, both IHC and IF results showed that 5hmC levels increased significantly, while the 5mC levels did not exhibit significant changes in HaCaT cells, compared with HaCat cells without UVB exposure. Moreover, compared with HaCat cells without UVB exposure, the levels ofTET1, TET2, and TET3 mRNA and protein expression were significantly upregulated （mRNA： P = 0.0022 and 0.0043 for TET1; all P 〈 0.0001 for TET2; all P = 0.0006 for TET3; protein： P = 0.0012 and 0.0006 tbr TET 1 ; all P = 0.0022 for TET2; and all P = 0.0002 for TET3）, and the levels of MMP- 1 mRNA expression increased dose dependently in 40 mJ/cm^2 and 80 mJ/cm^2 UVB-irradiated groups. Conclusion： UVB radiation could cause increased 5hmC and TET expression, which might become a novel biomarker in UVB-related skin aging.

Site-specific quantification of 5-carboxylcytosine in DNA by chemical conversion coupled with ligation-based PCR

5-Methylcytosine(5mC)is the most important epigenetic modification in mammals.The active DNA demethylation could be achieved through the ten-eleven translocation(TET)protein-mediated oxidization of 5mC with the generation of 5-hydroxymethylcytosine(5hmC),5-formylcytosine(5fC)and 5-carboxylcytosine(5caC).It has been known that 5mC,5hmC and 5fC play critical roles in modulating gene expression.However,unlike the 5mC,5hmC,and 5fC,the functions of 5caC are still underexplored.Investigation of the functions of 5caC relies on the accurate quantification and localization analysis of 5caC in DNA.In the current study,we developed a method by chemical conversion in conjugation with ligation-based real-time quantitative PCR(qPCR)for the site-specific quantification of 5caC in DNA.This method depends on the selective conversion of 5caC to form dihydrouracil(DHU)by pyridine borane treatment.DHU behaves like thymine and pairs with adenine(DHU-A).Thus,the chemical conversion by pyridine borane leads to the transformation of base paring from 5caC-G to DHU-A,which is utilized to achieve the site-specific detection and quantification of 5caC in DNA.As a proof-of-concept,the developed method was successfully applied in the site-specific quantification of 5caC in synthesized DNA spiked in complex biological samples.The method is rapid,straightforward and cost-effective,and shows promising in promoting the investigation of the functional roles of 5caC in future study.

Research of total levels on DNA methylation in plant based on HPLC analysis

HPLC analysis is important for determination of total level on DNA methylation in plants. It can be used to help characterise epigenetic changes during growth, development and stress. HPLC methods have been optimised for mammalian and microbial DNA, but not for plants. This article examines several important factors in the HPLC analysis of plant DNA methylation including extraction and purification of DNA and HPLC conditions choice by using leaves of rice seedling. The experimental results showed that RNA of nucleic acid was removed by using RNase A. This study also identified critical components of HPLC analysis. With the optimized method of HPLC conditions, the better result was achieved in the chromatogram of cytosine and 5-methylcytosine in genomic DNA acid hydrolysis. The study would offer a comprehensive guide for the stringent analysis of DNA methylation in plants.

Recent Advances in Deciphering the Mechanisms and Biological Functions of DNA Demethylation

5-Methylcytosine (5mC) is a dynamic and reversible epigenetic modification in genomic DNA of higher eukaryotes.It has been well-established that the demethylation of 5mC occurs through the ten-eleven translocation (TET)-mediated oxidation of 5mC followed by thymine DNA glycosylase (TDG)-initiated base excision repair (BER).Recent findings also have identified an alternative pathway of DNA demethylation.In this pathway,TET enzymes directly oxidize 5mC to form 5-formylcytosine (5fC) or 5-carboxylcytosine (5caC).These modified bases can undergo direct deformylation or decarboxylation,respectively.Additionally,DNA demethylation can also occur through the deamination of 5mC and 5hmC,resulting in the production of thymine and 5-hydroxymethyluracil (5hmU),respectively.Various DNA demethylation pathways possess critical functional implications and roles in biological processes.This Recent Advances article will focus on the studies of mechanisms and biological functions of DNA demethylation,shedding light on the reversible nature of the epigenetic modification of 5mC.

Identifying RNA Modifications by Direct RNA Sequencing Reveals Complexity of Epitranscriptomic Dynamics in Rice

Transcriptome analysis based on high-throughput sequencing of a cDNA library has been widely applied to functional genomic studies.However,the cDNA dependence of most RNA sequencing techniques constrains their ability to detect base modifications on RNA,which is an important element for the post-transcriptional regulation of gene expression.To comprehensively profile the N^(6)-methyladenosine(m^(6)A)and N^(5)-methylcytosine(m5 C)modifications on RNA,direct RNA sequencing(DRS)using the latest Oxford Nanopore Technology was applied to analyze the transcriptome of six tissues in rice.Approximately 94 million reads were generated,with an average length ranging from 619 nt to 1013 nt,and a total of 45,707 transcripts across 34,763 genes were detected.Expression profiles of transcripts at the isoform level were quantified among tissues.Transcriptome-wide mapping of m^(6)A and m5 C demonstrated that both modifications exhibited tissue-specific characteristics.The transcripts with m^(6)A modifications tended to be modified by m5 C,and the transcripts with modifications presented higher expression levels along with shorter poly(A)tails than transcripts without modifications,suggesting the complexity of gene expression regulation.Gene Ontology analysis demonstrated that m^(6)A-and m5 C-modified transcripts were involved in central metabolic pathways related to the life cycle,with modifications on the target genes selected in a tissue-specific manner.Furthermore,most modified sites were located within quantitative trait loci that control important agronomic traits,highlighting the value of cloning functional loci.The results provide new insights into the expression regulation complexity and data resource of the transcriptome and epitranscriptome,improving our understanding of the rice genome.

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.

Global methylation status of sperm DNA in carriers of chromosome structural aberrations

Male infertility might be clearly associated with aberrant DNA methylation patterns in human spermatozoa. An association between oxidative stress and the global methylation status of the sperm genome has also been suggested. The aim of the present study was to determine whether the global sperm DNA methylation status was affected in the spermatozoa of carriers of chromosome structural aberrations. The relationships between the 5-methylcytosine （msC） levels in spermatozoa and chromatin integrity status were evaluated. The study patients comprised male carriers of chromosome structural aberrations with reproductive failure （n = 24）, and the controls comprised normozoospermic sperm volunteers （n = 23）. The global msC level was measured using thin-layer chromatography （TLC） and immunofluorescence （IF） techniques. The sperm chromatin integrity was assessed using aniline blue （AB） staining and TUNEL assay. The mean msC levels were similar between the investigated chromosome structural aberrations carriers （P） and controls （K）. However, sperm chromatin integrity tests revealed significantly higher values in chromosomal rearrangement carriers than in controls （P 〈 0.05）. Although the potential relationship between sperm chromatin integrity status and sperm DNA fragmentation and the msC level juxtaposed in both analyzed groups （P vs K） was represented in a clearly opposite manner, the low chromatin integrity might be associated with the high hypomethylation status of the sperm DNA observed in carriers of chromosome structural aberrations.

Multiple Functions of Ten-eleven Translocation 1 during Tumorigenesis

Objective： Aberrant expression of ten-eleven translocation 1 （TET1） plays a critical role in tumor development and progression. We systematically summarized the latest research progress on the role and mechanisms of TET1 in cancer biology. Data Sources： Relevant articles published in English from 1980 to April 2016 were selected from the PubMed database. Tile terms ＂＇ten-eleven translocation 1,＂ ＂＇SmC,＂ ＇5hmC,＂ ＇microRNA,＇＂ ＂hypoxia,＂ and ＂＇embryonic stem cell＇＂ were used tbr the search. Study Selection： Articles focusing on the role and mechanism ofTETI in tumor were reviewed, including clinical and basic research articles. Results： TET proteins, the key enzymes converting 5-methylcytosine to 5-hydroxymethylcytosine, play vital roles in DNA demethylation regulation. Recent studies have shown that loss of TETI is associated with tumorigenesis and can be used as a potential biomarker for cancer therapy, which indicates that TETI serves as tumor suppressor gene. Moreover, besides its dioxygenase activity, TET1 could induce epithelial-mesenchymal transition and act as a coactivator to regulate gene transcription, such as developmental regulator in embryonic stem cells （ESCs） and hypoxia-responsive gene in cancer. The regulation of TETl is also correlated with microRNA in a posttranscriptional modification process. Hence, it is complex but critical to coaprehend the mechanisms of TETI in the biology of ESCs and cancer. Conclusions： TET I not only serves as a demethylation enzyme but also plays multiple roles during tumorigenesis and progression. More studies should be carried out to elucidate the exact mechanisms of TETI and its associations with cancer betbre considering it as a therapeutic tool.