Genome-Wide Survey and Comparative Analysis of LTR Retrotransposons and Their Captured Genes in Rice and Sorghum

Long terminal repeat (LTR) retrotransposons are the major class I mobile elements in plants. They play crucial roles in gene expansion, diversification and evolution. However, their captured genes are yet to be genome-widely identified and characterized in most of plants although many genomes have been completely sequenced. In this study, we have identified 7,043 and 23,915 full-length LTR retrotransposons in the rice and sorghum genomes, respectively. High percentages of rice full-length LTR retrotransposons were distributed near centromeric region in each of the chromosomes. In contrast, sorghum full-length LTR retrotransposons were not enriched in centromere regions. This dissimilarity could be due to the discrepant retrotransposition during and after divergence from their common ancestor thus might be contributing to species divergence. A total of 672 and 1,343 genes have been captured by these elements in rice and sorghum, respectively. Gene Ontology (GO) and gene set enrichment analysis (GSEA) showed that no over-represented GO term was identified in LTR captured rice genes. For LTR captured sorghum genes, GO terms with functions in DNA/RNA metabolism and chromatin organization were over-represented. Only 36% of LTR captured rice genes were expressed and expression divergence was estimated as 11.9%. Higher percentage of LTR captured rice genes have evolved into pseudogenes under neutral selection. On the contrary, higher percentage of LTR captured sorghum genes were under purifying selection and 72.4% of them were expressed. Thus, higher percentage of LTR captured sorghum genes was functional. Small RNA analysis suggested that some of LTR captured genes in rice and sorghum might have been involved in negative regulation. On the other hand, positive selection has been observed in both rice and sorghum LTR captured genes and some of them were still expressed and functional. The data suggest that some of these LTR captured genes might have evolved into new gene functions.     

家蚕LTR逆转录转座子的鉴定、分类及系统发育分析

转座子是真核生物基因组的重要组成成分。为了研究家蚕Bombyx mori长末端重复序列 (long terminal repeat, LTR)逆转录转座子的分类及进化, 本研究采用de novo预测和同源性搜索相结合的方法, 在家蚕基因组中共鉴定出了38个LTR逆转录转座子家族, 序列长度占整个基因组的0.64%, 远小于先前预测的11.8%, 其中有6个家族为本研究的新发现。38个家族中, 26个家族有表达序列标签 (expression sequence tag, EST)证据, 表明这些家族具有潜在的活性。对有EST证据的6个家族和没有EST证据的5个家族用RT-PCR进行了组织表达谱实验, 结果表明这11个家族在一些组织中有表达, 这进一步证实了这些家族具有转录活性, 基于此我们推测家蚕中大部分的LTR逆转录转座子家族很可能具有潜在活性。对转座子的插入时间进行估计, 结果表明绝大部分元件都是最近1百万年内插入到家蚕基因组中的。我们还比较了黑腹果蝇Drosophila melanogaster、 冈比亚按蚊Anopheles gambiae和家蚕B. mori中Ty3/Gypsy超家族分支的差异, 结果表明不同枝在不同昆虫中有着不同的扩张。家蚕中LTR逆转录转座子的鉴定和系统分析有助于我们理解逆转录转座子在昆虫进化中的作用。     

Young but not relatively old retrotransposons are preferentially located in gene‐rich euchromatic regions in tomato (Solanum lycopersicum) plants

Long terminal repeat (LTR) retrotransposons are the major DNA components of flowering plants. They are generally enriched in pericentromeric heterochromatin regions of their host genomes, which could result from the preferential insertion of LTR retrotransposons and the low effectiveness of purifying selection in these regions. To estimate the relative importance of the actions of these two factors on their distribution pattern, the LTR retrotransposons in Solanum lycopersicum (tomato) plants were characterized at the genome level, and then the distribution of young elements was compared with that of relatively old elements. The current data show that old elements are mainly located in recombination‐suppressed heterochromatin regions, and that young elements are preferentially located in the gene‐rich euchromatic regions. Further analysis showed a negative correlation between the insertion time of LTR retrotransposons and the recombination rate. The data also showed there to be more solo LTRs in genic regions than in intergenic regions or in regions close to genes. These observations indicate that, unlike in many other plant genomes, the current LTR retrotransposons in tomatoes have a tendency to be preferentially located into euchromatic regions, probably caused by their severe suppression of activities in heterochromatic regions. These elements are apt to be maintained in heterochromatin regions, probably as a consequence of the pericentromeric effect in tomatoes. These results also indicate that local recombination rates and intensities of purifying selection in different genomic regions are largely responsible for structural variation and non‐random distribution of LTR retrotransposons in tomato plants.     

High rate of chimeric gene origination by retroposition in plant genomes

Retroposition is widely found to play essential roles in origination of new mammalian and other animal genes. However, the scarcity of retrogenes in plants has led to the assumption that plant genomes rarely evolve new gene duplicates by retroposition, despite abundant retrotransposons in plants and a reported long terminal repeat (LTR) retrotransposon-mediated mechanism of retroposing cellular genes in maize (Zea mays). We show extensive retropositions in the rice (Oryza sativa) genome, with 1235 identified primary retrogenes. We identified 27 of these primary retrogenes within LTR retrotransposons, confirming a previously observed role of retroelements in generating plant retrogenes. Substitution analyses revealed that the vast majority are subject to negative selection, suggesting, along with expression data and evidence of age, that they are likely functional retrogenes. In addition, 42% of these retrosequences have recruited new exons from flanking regions, generating a large number of chimerical genes. We also identified young chimerical genes, suggesting that gene origination through retroposition is ongoing, with a rate an order of magnitude higher than the rate in primates. Finally, we observed that retropositions have followed an unexpected spatial pattern in which functional retrogenes avoid centromeric regions, while retropseudogenes are randomly distributed. These observations suggest that retroposition is an important mechanism that governs gene evolution in rice and other grass species.     

Retrotransposon insertion polymorphisms in six rice genes and their evolutionary history

Retrotransposons are abundant in higher plant genomes. Although retrotransposons associated with plant genes have been identified, little is known about their evolutionary conservation at the level of species and subspecies. In the present study, we investigated the phylogenetic distribution of long terminal repeat (LTR) retrotransposon, long interspersed nuclear element (LINE) and short interspersed nuclear element (SINE) insertions in six genes in 95 cultivated and wild rice genotypes. These six genes are likely to be functional based on nonsynonymous (Ka) to synonymous (Ks) substitution ratios which were found to be significantly <1. Different conservation patterns of these retrotransposons in genes were observed in cultivated and wild rice species. Four out of seven retrotransposon insertions appear to predate the ancestral Oryza AA genome. Two of these insertions in genes 4 and 5 occurred early in the evolutionary history of Oryza. Two retrotransposon insertions in gene 1 arose after the divergence of Asian cultivated rice from its wild ancestor. Furthermore, the retrotransposon insertion in gene 3 appears to have occurred in the ancestral lineage leading to temperate japonicas. Conservation of retrotransposon insertions in genes in specific groups, species, and lineages might be related to their specific function.     

Evidence of multiple horizontal transfers of the long terminal repeat retrotransposon RIRE1 within the genus Oryza

Horizontal gene transfer, defined as the transmission of genetic material between reproductively isolated species, has been considered for a long time to be a rare phenomenon. Most well-documented cases of horizontal gene transfer have been described in prokaryotes or in animals and they often involve transposable elements. The most abundant class of transposable elements in plant genomes are the long terminal repeat (LTR) retrotransposons. Because of their propensity to increase their copy number while active, LTR retrotransposons can have a significant impact on genomics changes during evolution. In a previous study, we showed that in the wild rice species Oryza australiensis , 60% of the genome is composed of only three families of LTR retrotransposons named RIRE1 , Wallabi and Kangourou . In the present study, using both in silico and experimental approaches, we show that one of these three families, RIRE1 , has been transferred horizontally between O. australiensis and seven other reproductively isolated Oryza species. This constitutes a new case of horizontal transfer in plants.     

LTR retrotransposons and flowering plant genome size: emergence of the increase/decrease model

Long Terminal Repeat (LTR) retrotransposons are ubiquitous components of plant genomes. Because of their copy-and-paste mode of transposition, these elements tend to increase their copy number while they are active. In addition, it is now well established that the differences in genome size observed in the plant kingdom are accompanied by variations in LTR retrotransposon content, suggesting that LTR retrotransposons might be important players in the evolution of plant genome size, along with polyploidy. The recent availability of large genomic sequences for many crop species has made it possible to examine in detail how LTR retrotransposons actually drive genomic changes in plants. In the present paper, we provide a review of the recent publications that have contributed to the knowledge of plant LTR retrotransposons, as structural components of the genomes, as well as from an evolutionary genomic perspective. These studies have shown that plant genomes undergo genome size increases through bursts of retrotransposition, while there is a counteracting process that tends to eliminate the transposed copies from the genomes. This process involves recombination mechanisms that occur either between the LTRs of the elements, leading to the formation of solo-LTRs, or between direct repeats anywhere in the sequence of the element, leading to internal deletions. All these studies have led to the emergence of a new model for plant genome evolution that takes into account both genome size increases (through retrotransposition) and decreases (through solo-LTR and deletion formation). In the conclusion, we discuss this new model and present the future prospects in the study of plant genome evolution in relation to the activity of transposable elements.     

植物活性长末端重复序列反转录转座子研究进展

长末端重复序列(Long terminal repeat,LTR)反转录转座子是真核生物基因组中普遍存在的一类可移动的DNA序列,它们以RNA为媒介,通过"复制粘贴"机制在基因组中不断自我复制。在高等植物中,许多活性的LTR反转录转座子已被详尽研究并应用于分子标记技术、基因标签、插入型突变及基因功能等分析。本文对植物活性LTR反转录转座子进行全面的调查,并对其结构、拷贝数和分布以及转座特性进行系统的归纳,分析了植物活性LTR反转录转座子的gag(种属特异抗原)和pol(聚合酶)序列特征,以及LTR序列中顺式调控元件的分布。研究发现自主有活性的LTR反转录转座子必须具备LTR区域以及编码Gag、Pr、Int、Rt和Rh蛋白的基因区。其中两端LTR区域具有高度同源性且富含顺式调控元件;Rt蛋白必备RVT结构域;Rh蛋白必备RNase_H1_RT结构域。这些结果为后续植物活性LTR反转录转座子的鉴定和功能分析奠定了重要基础。     

Characterization of genome-wide long terminal repeat retrotransposons provide insights into trait evolution of four grapevine species

Grapevine is one of the most economically important crops in the world. Although long terminal repeat (LTR) retrotransposons are thought to have played an important role in plants, its distribution in grapevine is not clear. Here, we identified genome-wide intact LTR retrotransposons in a total of six high-quality grapevine genomes from Vitis vinifera L., Vitis sylvestris C.C. Gmel., Vitis riparia Michx. and Vitis amurensis Rupr. with an average of 2938 per genome. Among them, the Copia superfamily (particularly for Ale) is a major component of the LTR retrotransposon in grapevine. Insertion time and copy number analysis revealed that the expansion of 70% LTR retrotransposons concentrating on approximately 2.5 Ma was able to drive genome size variation. Phylogenetic tree and syntenic analyses showed that most LTR retrotransposons in these genomes formed and evolved after species divergence. Furthermore, the function and expression of genes inserted by LTR retrotransposons in V. vinifera (Pinot noir) and V. riparia were explored. The length and expression of genes related to starch metabolism and quinone synthesis pathway in Pinot noir and environmental adaptation pathway in V. riparia were significantly affected by LTR retrotransposon insertion. The results improve the understanding of LTR retrotransposons in grapevine genomes and provide insights for its potential contribution to grapevine trait evolution.     

Replication of nonautonomous retroelements in soybean appears to be both recent and common

Retrotransposons and their remnants often constitute more than 50% of higher plant genomes. Although extensively studied in monocot crops such as maize (Zea mays) and rice (Oryza sativa), the impact of retrotransposons on dicot crop genomes is not well documented. Here, we present an analysis of retrotransposons in soybean (Glycine max). Analysis of approximately 3.7 megabases (Mb) of genomic sequence, including 0.87 Mb of pericentromeric sequence, uncovered 45 intact long terminal repeat (LTR)-retrotransposons. The ratio of intact elements to solo LTRs was 8:1, one of the highest reported to date in plants, suggesting that removal of retrotransposons by homologous recombination between LTRs is occurring more slowly in soybean than in previously characterized plant species. Analysis of paired LTR sequences uncovered a low frequency of deletions relative to base substitutions, indicating that removal of retrotransposon sequences by illegitimate recombination is also operating more slowly. Significantly, we identified three subfamilies of nonautonomous elements that have replicated in the recent past, suggesting that retrotransposition can be catalyzed in trans by autonomous elements elsewhere in the genome. Analysis of 1.6 Mb of sequence from Glycine tomentella, a wild perennial relative of soybean, uncovered 23 intact retroelements, two of which had accumulated no mutations in their LTRs, indicating very recent insertion. A similar pattern was found in 0.94 Mb of sequence from Phaseolus vulgaris (common bean). Thus, autonomous and nonautonomous retrotransposons appear to be both abundant and active in Glycine and Phaseolus. The impact of nonautonomous retrotransposon replication on genome size appears to be much greater than previously appreciated.     

TEnest: automated chronological annotation and visualization of nested plant transposable elements

Organisms with a high density of transposable elements (TEs) exhibit nesting, with subsequent repeats found inside previously inserted elements. Nesting splits the sequence structure of TEs and makes annotation of repetitive areas challenging. We present TEnest, a repeat identification and display tool made specifically for highly repetitive genomes. TEnest identifies repetitive sequences and reconstructs separated sections to provide full-length repeats and, for long-terminal repeat (LTR) retrotransposons, calculates age since insertion based on LTR divergence. TEnest provides a chronological insertion display to give an accurate visual representation of TE integration history showing timeline, location, and families of each TE identified, thus creating a framework from which evolutionary comparisons can be made among various regions of the genome. A database of repeats has been developed for maize (Zea mays), rice (Oryza sativa), wheat (Triticum aestivum), and barley (Hordeum vulgare) to illustrate the potential of TEnest software. All currently finished maize bacterial artificial chromosomes totaling 29.3 Mb were analyzed with TEnest to provide a characterization of the repeat insertions. Sixty-seven percent of the maize genome was found to be made up of TEs; of these, 95% are LTR retrotransposons. The rate of solo LTR formation is shown to be dissimilar across retrotransposon families. Phylogenetic analysis of TE families reveals specific events of extreme TE proliferation, which may explain the high quantities of certain TE families found throughout the maize genome. The TEnest software package is available for use on PlantGDB under the tools section (http://www.plantgdb.org/prj/TE_nest/TE_nest.html); the source code is available from (http://wiselab.org).     

Young,intact and nested retrotransposons are abundant in the onion and asparagus genomes

Background and Aims

Although monocotyledonous plants comprise one of the two major groups of angiosperms and include >65 000 species, comprehensive genome analysis has been focused mainly on the Poaceae (grass) family. Due to this bias, most of the conclusions that have been drawn for monocot genome evolution are based on grasses. It is not known whether these conclusions apply to many other monocots.

Methods

To extend our understanding of genome evolution in the monocots, Asparagales genomic sequence data were acquired and the structural properties of asparagus and onion genomes were analysed. Specifically, several available onion and asparagus bacterial artificial chromosomes (BACs) with contig sizes >35 kb were annotated and analysed, with a particular focus on the characterization of long terminal repeat (LTR) retrotransposons.

Key Results

The results reveal that LTR retrotransposons are the major components of the onion and garden asparagus genomes. These elements are mostly intact (i.e. with two LTRs), have mainly inserted within the past 6 million years and are piled up into nested structures. Analysis of shotgun genomic sequence data and the observation of two copies for some transposable elements (TEs) in annotated BACs indicates that some families have become particularly abundant, as high as 4–5 % (asparagus) or 3–4 % (onion) of the genome for the most abundant families, as also seen in large grass genomes such as wheat and maize.

Conclusions

Although previous annotations of contiguous genomic sequences have suggested that LTR retrotransposons were highly fragmented in these two Asparagales genomes, the results presented here show that this was largely due to the methodology used. In contrast, this current work indicates an ensemble of genomic features similar to those observed in the Poaceae.     

Structure and evolution of full-length LTR retrotransposons in rice genome

The long terminal repeat (LTR) retrotransposons are the most abundant class of transposable elements in plant genomes and play important roles in genome divergence and evolution. Their accumulation is the main factor influencing genome size increase in plants. Rice (Oryza sativa L.) is a model monocot and is the focus of much biological research due to its economic importance. We conducted a comprehensive survey of full-length LTR retrotransposons based on the completed genome of japonica rice variety Nipponbare (TIGR Release 5), with the newly published tool LTR-FINDER. The elements could be categorized into 29 structural domain categories (SDCs), and their total copy number identified was estimated at >6,000. Most of them were relatively young: more than 90% were less than 10 My. There existed a high level of activity among them as a whole at 0–1 Mya, but different categories possessed distinct amplification patterns. Most recently inserted elements were specific to the rice genome, while a few were conserved across species. This study provides new insights into the structure and evolutionary history of the full-length retroelements in the rice genome.     

The heterochromatic copies of the LTR retrotransposons as a record of the genomic events that have shaped the Drosophila melanogaster genome

Transposable elements, which are major components of most genomes, are known to accumulate in heterochromatic regions in which they have progressively diverged in sequence by mutations and internal deletions and insertions (indels) during the course of evolution. They therefore provide a record of the genomic events that have shaped the genomes, some of which could correspond to speciation events. Using the sequence divergence between the long terminal repeats (LTRs), we estimated the date of the insertion events of the LTR retrotransposon copies embedded within the heterochromatin regions of the Drosophila melanogaster genome. We did not detect traces of any specific waves of mobilization of retrotransposons within heterochromatin, apart from a very recent wave, which corresponds to the numerous LTR retrotransposon copies found in euchromatin.     

Development of retrotransposon-based molecular markers and their application in genetic mapping in chokecherry (<Emphasis Type="Italic">Prunus virginiana</Emphasis> L.)

Retrotransposons are the largest group of transposable elements (TEs) that are ubiquitous and well dispersed in plant genomes. Transposition/insertion of TEs on chromosomes often generates unique repeat junctions (RJs) between TEs and their flanking sequences. Long terminal repeats (LTR) are well conserved and abundant in plant genomes, making LTR retrotransposons valuable for development of TE junction-based markers. In this study, LTR retrotransposons and their RJs were detected from chokecherry genome sequences generated by Roche 454 sequencing. A total of 1246 LTR retrotransposons were identified, and 338 polymerase chain reaction primer pairs were designed. Of those, 336 were used to amplify DNA from chokecherry and other rosaceous species. An average of 283 of 336 (84.2 %) LTR primer pairs effectively amplified DNA from chokecherries. One hundred and seventeen chokecherry LTR primers also produced amplification in other Prunus (99) or rosaceous species (19). A total of 59 of 78 polymorphic LTR markers were qualified for linkage map construction according to the segregation distortion Chi-square (χ ²) test. Forty-eight LTR markers were successfully located on a previously constructed chokecherry map. The majority of the LTR markers were mapped on LG XI of the chokecherry map. Our results suggest that LTR marker development using random genome sequences is rapid and cost-efficient. Confirmed applicability of LTR markers in map construction and genetic mapping will facilitate genetic research in chokecherry and other rosaceous species.     

Swine liver methylomes of Berkshire,Duroc and Landrace breeds by MeDIPS

Using a methyl‐DNA immunoprecipitation technique in combination with next‐generation deep sequencing, we conducted comprehensive DNA methylation profiling of liver genomes from three pig breeds: Berkshire, Duroc and Landrace. The profiles revealed that the distribution patterns of methylation signals along the genome are conserved among the three pig breeds. Specifically, many signals in coding genes were found in introns, and most signals in the repetitive elements were identified in non‐long terminal repeat (LTR) retrotransposons such as long and short interspersed repetitive elements, implying a significant association with alternative splicing and expression of retrotransposable elements respectively. Differentially methylated regions among the three pig breeds were identified in the non‐LTR retrotransposons, suggesting that they may lead to differential retrotransposable element activity. Altogether, this study provides advanced swine methylome data and valuable resources for understanding the function of DNA methylation in the evolutionary divergence of different pig breeds.