Pig H3K4me3,H3K27ac,and gene expression profiles reveal reproductive tissue-specific activity of transposable elements

Regulatory sequences and transposable elements(TEs)account for a large proportion of the genomic sequences of species;however,their roles in gene transcription,especially tissue-specific expression,remain largely unknown.Pigs serve as an excellent animal model for studying genomic sequence biology due to the extensive diversity among their wild and domesticated populations.Here,we conducted an integrated analysis using H3K27ac ChIP-seq,H3K4me3 ChIP-seq,and RNA-seq data from 10 different tissues of seven fetuses and eight closely related adult pigs.We aimed to annotate the regulatory elements and TEs to elucidate their associations with histone modifications and mRNA expression across different tissues and developmental stages.Based on correlation analysis between mRNA expression and H3K27ac and H3K4me3 peak activity,results indicated that H3K27ac exhibited stronger associations with gene expression than H3K4me3.Furthermore,1.45%of TEs overlapped with either the H3K27ac or H3K4me3 peaks,with the majority displaying tissue-specific activity.Notably,a TE subfamily(LTR4C_SS),containing binding motifs for SIX1 and SIX4,showed specific enrichment in the H3K27ac peaks of the adult and fetal ovaries.RNA-seq analysis also revealed widespread expression of TEs in the exons or promoters of genes,including 4688 TE-containing transcripts with distinct development stage-specific and tissue-specific expression.Of note,1967 TE-containing transcripts were enriched in the testes.We identified a long terminal repeat(LTR),MLT1F1,acting as a testis-specific alternative promoter in SRPK2(a cell cycle-related protein kinase)in our pig dataset.This element was also conserved in humans and mice,suggesting either an ancient integration of TEs in genes specifically expressed in the testes or parallel evolutionary patterns.Collectively,our findings demonstrate that TEs are deeply embedded in the genome and exhibit important tissue-specific biological functions,particularly in the reproductive organs.

Transposable elements in <i>Escherichia coli</i>antimicrobial resistance

Transposable elements are capable of switching their positions on the genome thereby causing gene arrangements and contributing to genome evolution. The aim of this review is to specifically discuss the role of transposable elements in transferring antimicrobial resistance genes in E. coli, thus contributing to increase in virulence and conferring the possibility of multidrug resistance. Different types of transposable elements such as transposons and integrons and their profound influence on E. coli antimicrobial resistance are the focus of this review.

High genetic variation and recombination events in the vicinity of non-autonomous transposable elements from ‘Candidatus Liberibacter asiaticus'

Two miniature inverted-repeat transposable elements（MITEs）, MCLas-A and MCLas-B, were recently identified from ‘Candidatus Liberibacter asiaticus＇ known to be associated with citrus Huanglongbing（HLB, yellow shoot disease）. MCLas-A was suggested as an active MITE because of its mobility. The immediate upstream gene of the two MITEs was predicted to be a putative transposase. The goal of this study is to analyze the sequence variation in the upstream putative transposase of MITEs and explore the possible correlation between sequence variation of transposase gene and MITE activity. PCR and sequence analysis showed that 12 sequence types were found in six major amplicon types from 43 representative ‘Ca. L. asiaticus＇ isolates from China, the United States and Brazil. Out of the 12 sequence types, three（T4, T5-2, T6） were reported for the first time. Recombination events were found in the two unique sequence types（T5-2 and T6） which were detected in all Brazilian isolates. Notably, no sequence variation or recombination events were detected in the upstream putative transposase gene of MCLas-A, suggesting the conservation of the transposase gene might be closely related with the MITE activity. Phylogenetic analysis demonstrated two well supported clades including five subclades were identified, clearly reflecting the geographical origins of isolates, especially that of Ruili isolates, S？o Paulo isolates and a few Florida isolates.

Dynamic evolution of transposable elements,demographic history,and gene content of paleognathous birds

Palaeognathae includes ratite and tinamou species that are important for understanding early avian evolution.Here,we analyzed the whole-genome sequences of 15 paleognathous species to infer their demographic histories,which are presently unknown.We found that most species showed a reduction of population size since the beginning of the last glacial period,except for those species distributed in Australasia and in the far south of South America.Different degrees of contraction and expansion of transposable elements(TE)have shaped the paleognathous genome architecture,with a higher transposon removal rate in tinamous than in ratites.One repeat family,AviRTE,likely underwent horizontal transfer from tropical parasites to the ancestor of little and undulated tinamous about 30 million years ago.Our analysis of gene families identified rapid turnover of immune and reproductionrelated genes but found no evidence of gene family changes underlying the convergent evolution of flightlessness among ratites.We also found that mitochondrial genes have experienced a faster evolutionary rate in tinamous than in ratites,with the former also showing more degenerated W chromosomes.This result can be explained by the Hill-Robertson interference affecting genetically linked W chromosomes and mitochondria.Overall,we reconstructed the evolutionary history of the Palaeognathae populations,genes,and TEs.Our findings of co-evolution between mitochondria and W chromosomes highlight the key difference in genome evolution between species with ZW sex chromosomes and those with XY sex chromosomes.

Histone H3K27me3 methylation regulates the expression of secreted proteins distributed at fast-evolving regions through transcriptional repression of transposable elements

The fine-tuned expression dynamics of the effector genes are pivotal for the transition from vegetative growth to host colonization of pathogenic filamentous fungi.However,mechanisms underlying the dynamic regulation of these genes remain largely unknown.Here,through comparative transcriptome and chromatin immunoprecipitation sequencing(ChIP-seq)analyses of the methyltransferase PoKmt6 in rice blast fungus Pyricularia oryzae(syn.Magnaporthe oryzae),we found that PoKmt6-mediated H3K27me3 deposition was enriched mainly at fast-evolving regions and contributed to the silencing of a subset of secreted proteins(SP)and transposable element(TE)families during the vegetative growth of P.oryzae.Intriguingly,we observed that a group of SP genes,which were depleted of H3K27me3 modification,could also be silenced via the H3K27me3-mediated repression of the nearby TEs.In conclusion,our results indicate that H3K27me3 modification mediated by PoKmt6 regulates the expression of some SP genes in fast-evolving regions through the suppression of nearby TEs.

Molecular Characterization of Type II Transposable Elements in Cowpea [<i>Vigna unguiculata</i>(L.) Walp]

Previous genetic studies in cowpea [Vigna unguiculata (L.) Walp] have shown that an active bipartite transposable element (TE) is responsible for a range of mutant phenotypes of its leaf, stem and flower. Since type II TEs have not been characterized at the molecular level in cowpea, this study was initiated to survey the presence of type II TEs in the cowpea genome. Type II TEs: Enhancer/Suppressor-mutator (En/Spm) and Miniature Inverted-repeat Transposable Elements (MITEs) were isolated and characterized. The sequence identity between the EnSpm TE clones was 46% at the nucleotide level (NL) and 30% at the amino acid level (AL) while that of MITEs was 71% at NL and 63% at AL. These cowpea En/Spm TEs were 80% homologous with En/Spm elements of other crops at NL and 46% at AL. The MITEs were 96% similar at NL and 18% homologous at AL. DNA gel blot analysis confirmed the presence of the En/Spm TEs in cowpea. RT-PCR (reverse transcriptase polymerase chain reaction) analysis showed that the VuEnSpm-3 and the MITE clone, VuPIF-1 were actively transcribed in wild type and mutant cowpea tissues. Overall, our data show that multiple, divergent lineages of En/Spm and MITEs are present in the cowpea genome, some of which are actively transcribed. Our findings also offer new molecular resource to further investigate the genetic determinants underlying previously described mutant cowpea phenotypes.

Isolating the Mutator Transposable Element Insertional Mutant Gene mio16 of Maize Using Double SelectedAmplification of Insertion Flanking Fragments (DSAIFF)

Mutator transposable element （Mu） has been used as an effective tool to clone maize （Zea mays L.） genes. One opaque endosperm mutant （miol6） was identified in a pool of Mu inserted mutants. A modified method, termed the double selected amplification of insertion flanking fragments （DSAIFF）, was employed to isolate the Mu flanking fragments （MFFs） of miol6. The target site duplications （TSDs） isolated from the Msp I and Mse I digested MFFs had a same 9-bp sequence and were confirmed to be the flanking sequence of one identically inserted gene. Co-segregation analysis suggested that the MFFs were associated with the mutant opaque endosperm, and miol6 was mapped in silico onto the physical position ranged from 229 965 021 to 229 965 409 bp of the maize chromosome 4.09 bin. The full-length cDNA of the wild-type gene was obtained by an RT-PCR primer-scanning technique, and Mio16 was found to putatively encode a homolog of the Arabidopsis MAP3K delta-1 protein kinase. RT-PCR result the mRNA expression of miol6 region anchored by primers Mu20 and af276 was not interrupted by Mu insertion. Further researches will be done to elucidate how the expression of miol6 is alternated by Mu insertion.

Diversity and association analysis of important agricultural trait based on miniature inverted-repeat transposable element specific marker in Brassica napus L.

Miniature inverted-repeat transposable elements(MITEs)are a group of DNA transposable element(TE)which preferentially distributed with gene associated regions.Tens of MITEs families have been revealed in Brassica napus genome,they scatter across the genome with tens of thousands copies and produce polymorphisms both intra-and inter-species.Our previous studies revealed a Tourist-like MITE,Monkey King,associated with vernalization requirement of B.napus,however there are still few studies reveal MITE association with agricultural traits in B.napus.In the present study,80 polymorphic markers were developed from 55 MITEs,and used to evaluate genetic diversity in a panel of B.napus accessions consisting of 101 natural and 25 synthetic genotypes.Five agricultural traits,oil content,glucosinolate content,erucic acid content,weight of thousand seeds(WTS)and plant height,were investigated across 3-years field experiments,in addition,two traits,hypocotyl length and root length,were evaluated at the 4-leaf stage in the laboratory.Correlations between the MITE-based markers and seven traits were analyzed,finally,10 polymorphic markers produced by 6 pairs of MITE specific primers were revealed relatively high correlation with 5 traits.Two polymorphic markers were anchored with two candidate genes,BnaA02g13530D and BnaA08g20010D,respectively,which may contribute to glucosinolate content and WTS.This research may contribute to genetic improvement through utilization of MITE-induced polymorphisms in Brassica species.

Construction of a transposase accessible chromatin landscape reveals chromatin state of repeat elements and potential causal variant for complex traits in pigs

Background:A comprehensive landscape of chromatin states for multiple mammalian tissues is essential for elucidating the molecular mechanism underlying regulatory variants on complex traits.However,the genome-wide chromatin accessibility has been only reported in limited tissue types in pigs.Results:Here we report a genome-wide landscape of chromatin accessibility of 20 tissues in two female pigs at ages of 6 months using ATAC-seq,and identified 557,273 merged peaks,which greatly expanded the pig regulatory ele-ment repository.We revealed tissue-specific regulatory elements which were associated with tissue-relevant biologi-cal functions.We identified both positive and negative significant correlations between the regulatory elements and gene transcripts,which showed distinct distributions in terms of their strength and distances from corresponding genes.We investigated the presence of transposable elements(TEs)in open chromatin regions across all tissues,these included identifications of porcine endogenous retroviruses(PERVs)exhibiting high accessibility in liver and homology of porcine specific virus sequences to universally accessible transposable elements.Furthermore,we prior-itized a potential causal variant for polyunsaturated fatty acid in the muscle.Conclusions:Our data provides a novel multi-tissues accessible chromatin landscape that serve as an important resource for interpreting regulatory sequences in tissue-specific and conserved biological functions,as well as regula-tory variants of loci associated with complex traits in pigs.

Study on the Homologous Sequences of copia-like Retrotransposons in a Twin ovary Mutant of Rice

A twin ovary mutant derived from the doubled haploid (DH) progeny of a cross,02428/Gui 630, was presumably related to the transposition of some transposable elements. Up to date, all reported the active transposable elements in rice (Oryza sativa L.) are copia like retrotransposons. In the present study, the reverse transcriptase domains of copia like retrotransposons were amplified from the total DNA isolated from the mutant plants with the degenerated oligonucleotide primers for the domain. Three cloned insert DNAs, R33 1, R33 4 and R33 8, representing putative different copia like retrotransposons were screened out. Two of them displayed high polymorphism between “Zhaiyeqing 8” and “Jingxi 17”. Nine of the polymorphic bands were mapped on seven rice chromosomes. Sequencing analysis revealed that stop codons frequently occur in the sequence of R33 8, while both R33 1 and R33 4 contain a continuous coding region for 81 putative amino acid residues. No significant variation in hybridization patterns was found between indica and japonica rice or among 26 varieties of indica rice when R33 1 was used as a probe. Nevertheless, when R33 4 was used as a probe, high polymorphisms were detected both between indica and japonica rice and among 26 indica varieties, implying that this element might still be active in rice genomes.

Review Article: ISTR, a Retrotransposons-Based Marker to Assess Plant Genome Variability with Special Emphasis in the Genera <i>Zea</i>and <i>Agave</i>

A review is presented for give information and highlights the advantage of ISTR as molecular marker which with the genome changes after manipulation or breeding intervention can be detected.

Vectors for gene therapy:A place for DNA transposon

Gene therapy offers important perspectives in current and future medicine but suffers from imperfect vectors for the delivery of the therapeutic gene. Most preclinical and clinical trials have been based on the use of viral vectors, which have evident advantages but also some serious disadvantages. In the past decade the use of DNA transposon-based systems for gene delivery has emerged as a non-viral alternative. DNA transposon vector engineering remains largely in a preclinical phase but some interesting results have been obtained. This mini-review aims to provide the current state of the art on DNA transposon vectors used in a gene therapy perspective.

玉米非自主性转座子rDt的体细胞转座序列特征

Dotted/rDt是玉米遗传学中最早发现的双元转座子系统之一。为了揭示玉米中非自主性转座子rDt在其自主性转座子Dt调控下的转座遗传特性,选取了a1-rDt;Dt和a1-m1::rDt;Dt2个rDt转座子插入突变等位基因,检测玉米籽粒紫色斑点表型差异的遗传基础,利用巢式PCR与特异性酶切相结合的方法检测并鉴定了这2种材料叶片组织中rDt体细胞转座的印迹序列类型。通过构建遗传杂交群体,统计分析各群体的后代籽粒表型以及A1野生型基因的回复突变频率。结果显示,在籽粒糊粉层紫色斑点大小均一但数目极低的a1-rDt;Dt材料中,仅检测到2种体细胞转座的印迹序列类型,其中1种是没有转座子插入前A1野生型。而在籽粒糊粉层紫色斑点大小不一、排列密集的a1-m1::rDt;Dt材料中可以检测到5种体细胞转座的印迹序列类型,其中3种类型均保持A1'回复突变基因的开放阅读框;同时,a1-m1::rDt;Dt材料中A1'的回复突变频率是a1-rDt;Dt材料的大约2.6倍。研究表明,rDt在a1插入位点体细胞转座后的修复产物序列组成相对简单,与Ac/Ds等hAT超家族转座子相似,且rDt体细胞转座产生的印迹序列类型及丰富度是两个a1基因插入突变体中A1'回复突变频率高低及玉米籽粒糊粉层表型差异的遗传基础。

干旱条件下玉米转座子插入关联的表观调控分析

【目的】干旱是全球范围影响玉米生产的最主要胁迫因素之一。解析抗旱性的遗传基础与分子机制为玉米的抗旱改良提供依据。【方法】利用代表性玉米自交系,以叶片相对含水量和散粉-吐丝间隔为指标开展田间抗旱性精准鉴定。筛选2个抗旱性极端差异的自交系,开展基因组重测序分析和转座子插入鉴定;利用全基因组重亚硫酸盐测序(WGBS)方法分析不同水分处理下叶片和根系组织的DNA甲基化水平;同时利用转录组测序方法对相同样品的基因表达进行分析;通过比较分析获得2个材料间的转座子插入缺失变异、差异甲基化区域和差异表达基因,并综合分析这三者间的相关关系。针对前期克隆的玉米抗旱基因ZCN7,分析该基因区域转座子插入缺失变异介导的DNA甲基化和基因表达变化情况。【结果】在田间干旱处理下,自交系H082183的叶片相对含水量和散粉-吐丝间隔均与正常处理没有显著差异,而旅28在所有试验材料中表现最低的叶片相对含水量和最大的散粉-吐丝间隔。利用H082183和旅28这两个抗旱性极端差异的玉米自交系开展基因组重测序和转座子插入分析,分别检测到333754和333296个转座子插入,其中,有89954个转座子插入在2个自交系间具有多态性。基因组DNA甲基化分析表明,转座子、内含子和启动子区域较外显子和非编码区呈现较高的CG和CHG甲基化水平,经差异甲基化分析,在2个自交系间共检测到41352个差异甲基化区域,其中60%的差异甲基化区域位于转座子插入缺失变异的上下游5 kb范围内。基因表达水平与基因的CG和CHG甲基化水平负相关,在2个自交系干旱下的叶片和根系中分别鉴定到4196和3500个差异表达基因,其中19.5%和19.7%与差异甲基化区域关联。通过对抗旱相关基因ZCN7的研究,发现该基因34 kb区间内的3个LTR类转座子插入,造成自交系旅28在干旱和正常处理下的CG和CHG甲基化显著高于抗旱自交系H082183,并且H082183中ZCN7表达量也显著高于旅28。【结论】揭示了转座子介导的表观遗传调控在玉米响应干旱胁迫中的重要作用,进一步扩展了转座子变异和DNA甲基化调控抗旱基因ZCN7表达的分子机制。

Codon Usage Biases of Transposable Elements and Host Nuclear Genes in Arabidopsis thaliana and Oryza sativa

Transposable elements （TEs） are mobile genetic entities ubiquitously distributed in nearly all genomes. High frequency of codons ending in A/T in TEs has been previously observed in some species. In this study, the biases in nucleotide composition and codon usage of TE transposases and host nuclear genes were investigated in the AT-rich genome of Arabidopsis thaliana and the GC-rich genome of Oryza sativa. Codons ending in A/T are more frequently used by TEs compared with their host nuclear genes. A remarkable positive correlation between highly expressed nuclear genes and C/G-ending codons were detected in O. sativa （r=0.944 and 0.839, respectively, P〈0.0001） but not in A. thaliana, indicating a close association between the GC content and gene expression level in monocot species. In both species, TE codon usage biases are similar to that of weakly expressed genes. The expression and activity of TEs may be strictly controlled in plant genomes. Mutation bias and selection pressure have simultaneously acted on the TE evolution in A. thaliana and O. sativa. The consistently observed biases of nucleotide composition and codon usage of TEs may also provide a useful clue to accurately detect TE sequences in different species.

Gapless indica rice genome reveals synergistic contributions of active transposable elements and segmental duplications to rice genome evolution

The ultimate goal of genome assembly is a high-accuracy gapless genome.Here,we report a new assembly pipeline that is used to produce a gapless genome for the indica rice cultivar Minghui 63.The resulting 397.71-Mb final assembly is composed of 12 contigs with a contig N50 size of 31.93 Mb.Each chromosome is represented by a single contig and the genomic sequences of all chromosomes are gapless.Quality evaluation of this gapless genome assembly showed that gene regions in our assembly have the highest completeness compared with the other 15 reported high-quality rice genomes.Further comparison with the japonica rice genome revealed that the gapless indica genome assembly contains more transposable elements(TEs)and segmental duplications(SDs),the latter of which produce many duplicated genes that can affect agronomic traits through dose effect or sub-/neo-functionalization.The insertion of TEs can also affect the expression of duplicated genes,which may drive the evolution of these genes.Furthermore,we found the expansion of nucleotide-binding site with leucine-rich repeat disease-resistance genes and cis-zeatin-O-glucosyltransferase growth-related genes in SDs in the gapless indica genome assembly,suggesting that SDs contribute to the adaptive evolution of rice disease resistance and developmental processes.Collectively,our findings suggest that active TEs and SDs synergistically contribute to rice genome evolution.

非生物胁迫对毛竹转座子衍生TUCP转录活性的影响

【目的】转座子(TE)是真核细胞基因组的重要组成部分,在毛竹Phyllostachys edulis基因组超过63%时,易受胁迫诱导激活。分析非生物胁迫下,来源于转座子的不确定编码潜力转录本(TUCP)的表达模式,为转座子参与毛竹抗逆分子机制提供参考。【方法】采用生物信息学技术和手段,在低温、高温、高盐、紫外照射等4种胁迫处理下,研究毛竹TE-TUCPs及转座子邻近基因的转录特性和转录模式。通过实时荧光定量PCR(RT-qPCR)验证转录组来源的TETUCPs差异表达数据的可靠性。【结果】在毛竹4个胁迫处理转录本中,共鉴定出57627个TE-TUCPs。TE-TUCPs应对不同非生物胁迫表现出特异性表达模式。高温、高盐、紫外照射处理可以促进具有转录活性的TE-TUCPs附近基因差异表达,但是低温会抑制具有转录活性的TE-TUCPs附近基因差异表达。【结论】TE-TUCPs主要来源于Ty1/Copia和Ty3/Gypsy超家族。基因的表达潜能与近距离的TE-TUCPs表达潜能互相抑制。TE-TUCPs转录情况会受到非生物胁迫作用来调控附近基因的表达以适应胁迫影响。

Transposable elements play an important role during cotton genome evolution and fiber cell development

Transposable elements(TEs)usually occupy largest fractions of plant genome and are also the most variable part of the structure.Although traditionally it is hallmarked as"junk and selfish DNA",today more and more evidence points out TE’s participation in gene regulations including gene mutation,duplication,movement and novel gene creation via genetic and epigenetic mechanisms.The recently sequenced genomes of diploid cottons Gossypium arboreum(AA)and Gossypium raimondii(DD)together with their allotetraploid progeny Gossypium hirsutum(At At Dt Dt)provides a unique opportunity to compare genome variations in the Gossypium genus and to analyze the functions of TEs during its evolution.TEs accounted for 57%,68.5%and67.2%,respectively in DD,AA and At At Dt Dt genomes.The 1,694 Mb A-genome was found to harbor more LTR(long terminal repeat)-type retrotransposons that made cardinal contributions to the twofold increase in its genome size after evolution from the 775.2 Mb D-genome.Although the 2,173 Mb At At Dt Dt genome showed similar TE content to the A-genome,the total numbers of LTR-gypsy and LTR-copia type TEs varied significantly between these two genomes.Considering their roles on rewiring gene regulatory networks,we believe that TEs may somehow be involved in cotton fiber cell development.Indeed,the insertion or deletion of different TEs in the upstream region of two important transcription factor genes in At or Dt subgenomes resulted in qualitative differences in target gene expression.We suggest that our findings may open a window for improving cotton agronomic traits by editing TE activities.

Differentiation of a Miniature Inverted Transposable Element （MITE） System in Asian Rice Cultivars and Its Inference for a Diphyletic Origin of Two Subspecies of Asian Cultivated Rice

In the present study, we report a survey on a Miniature Inverted Transposable Element （MITE） system known as mPing in 102 varieties of Asian cultivated rice （Oryza sativa L.）. We found that mPing populations could be generalized Into two families, mPing-1 and mPing-2, according to their sequence structures. Further analysis showed that these two families of mPing had significant bias in their distribution pattern in two subspecies of rice, namely O. sativa ssp. japonica and indica. 0. sativa japonica has a higher proportion of mPing-1 as a general trait, whereas 0. sativa indica has a higher proportion of roPing-2. We also examined the mPing system In a doubled haploid （DH） cross-breeding population of jingxi 17 （japonica） and zhaiyeqing 8 （indica） varieties and observed that the mPing system was not tightly linked to major subspecies-determining genes. Furthermore, we checked the mPing system in 28 accessions of Asian common wild rice O. rufipogon and found the roPing system in 0. rufipogon. The distribution pattern of the roPing system in O. rufipogon indicated a diphyletlc origin of the Asian cultivated rice O. sativa species. We did not find the mPing system in another 20 Oryza species. These results substantiated a previous hypothesis that O. ruflpogon and O. nivara species were the closest relatives of O. sativa and that the two extant subspecies of O. sativa were evolved independently from corresponding ecotypes of O. ruflpogon.

PacBio Sequencing Reveals Transposable Elements as a Key Contributor to Genomic Plasticity and Virulence Variation in Magnaporthe oryzae

Dear Editor ：The sustainable cultivation of rice, which serves as staple food crop for more than half of the world＇s population, is under serious threat due to the huge yield losses inflicted by rice blast disease caused by the globally destructive fungus Magnaporthe oryzae （Pyricularia oryzae） （Dean et al., 2012; Nalley et al., 2016; Deng et al., 2017）. This filamentous ascomycete fungus is also capable of causing blast infection on other economically important cereal crops, including wheat, millet, and barley, making it the world＇s most important plant pathogenic fungus （Zhong et al., 2016）.