Miniature Inverted-repeat Transposable Elements Research Articles

Genome Analyses of Icelandic Strains ofSulfolobus islandicus, Model Organisms for Genetic and Virus-Host Interaction Studies

The genomes of two Sulfolobus islandicus strains obtained from Icelandic solfataras were sequenced and analyzed. Strain REY15A is a host for a versatile genetic toolbox. It exhibits a genome of minimal size, is stable genetically, and is easy to grow and manipulate. Strain HVE10/4 shows a broad host range for exceptional crenarchaeal viruses and conjugative plasmids and was selected for studying their life cycles and host interactions. The genomes of strains REY15A and HVE10/4 are 2.5 and 2.7 Mb, respectively, and each genome carries a variable region of 0.5 to 0.7 Mb where major differences in gene content and gene order occur. These include gene clusters involved in specific metabolic pathways, multiple copies of VapBC antitoxin-toxin gene pairs, and in strain HVE10/4, a 50-kb region rich in glycosyl transferase genes. The variable region also contains most of the insertion sequence (IS) elements and high proportions of the orphan orfB elements and SMN1 miniature inverted-repeat transposable elements (MITEs), as well as the clustered regular interspaced short palindromic repeat (CRISPR)-based immune systems, which are complex and diverse in both strains, consistent with them having been mobilized both intra- and intercellularly. In contrast, the remainder of the genomes are highly conserved in their protein and RNA gene syntenies, closely resembling those of other S. islandicus and Sulfolobus solfataricus strains, and they exhibit only minor remnants of a few genetic elements, mainly conjugative plasmids, which have integrated at a few tRNA genes lacking introns. This provides a possible rationale for the presence of the introns.

Small RNAs from MITE‐derived stem‐loop precursors regulate abscisic acid signaling and abiotic stress responses in rice

Small silencing RNAs (sRNAs) are non-coding RNA regulators that negatively regulate gene expression by guiding mRNA degradation, translation repression or chromatin modification. Plant sRNAs play crucial roles in various developmental processes, hormone signaling, immune responses and adaptation to a variety of abiotic stresses. miR441 and miR446 were previously annotated as microRNAs (miRNAs) because their precursors can form typical stem-loop structures, but are not considered as real miRNAs because of inconsistency with some ancillary criteria of the recent guidelines for annotation of plant miRNAs. We tentatively rename them small interfering (si)R441 and siR446, respectively, in this study. It has recently been shown that the precursors of siR441 and siR446 might originate from the miniature inverted-repeat transposable element (MITE) Stowaway1. In this report, we show that, in contrast with Dicer-like (DCL)3- and RNA-dependent RNA polymerase (RDR)2-dependent MITE-derived ra-siRNAs, siR441 and siR446 are processed by OsDCL3a but independent of OsRDR2, indicating that siR441 and siR446 are generated from single-stranded stem-loop precursors. We also show that abscisic acid (ABA) and abiotic stresses downregulate the expression of siR441 and siR446 but, surprisingly, increase the accumulation of their precursors in rice plants, implying that processing of siRNA precursors is inhibited. We provide evidence to show that this defective processing is due to increased precursor accumulation per se, possibly by intermolecular self-pairing of the processing intermediate sequences, thus hindering their normal processing. Functional examinations indicate that siR441 and siR446 are positive regulators of rice ABA signaling and tolerance to abiotic stress, possibly by regulating MAIF1 expression.

Recent transposition activity of Xenopus T2 family miniature inverted-repeat transposable elements

To investigate the recent transposition activity of T2 family miniature inverted-repeat transposable elements (MITEs) in Xenopus tropicalis (Western clawed frog), we analyzed the intraspecific polymorphisms associated with MITE insertion in X. tropicalis for three subfamilies of the T2 family (T2-A1, T2-C, and T2-E). A high frequency of MITE-insertion polymorphisms was observed at the T2-A1 (50%) and T2-C insertion loci (60%), but none were noted at the T2-E insertion locus (0%). Analyses of the collected data indicated that members of the T2-A1 and T2-C subfamilies may be currently active in the host species. Identification of these active transpositions will help us in understanding the mechanisms underlying the long-term survival (over several tens of millions of years) of the T2-A1 and T2-C subfamilies.

Abundant degenerate miniature inverted-repeat transposable elements in genomes of epichloid fungal endophytes of grasses.

Miniature inverted-repeat transposable elements (MITEs) are abundant repeat elements in plant and animal genomes; however, there are few analyses of these elements in fungal genomes. Analysis of the draft genome sequence of the fungal endophyte Epichloë festucae revealed 13 MITE families that make up almost 1% of the E. festucae genome, and relics of putative autonomous parent elements were identified for three families. Sequence and DNA hybridization analyses suggest that at least some of the MITEs identified in the study were active early in the evolution of Epichloë but are not found in closely related genera. Analysis of MITE integration sites showed that these elements have a moderate integration site preference for 5′ genic regions of the E. festucae genome and are particularly enriched near genes for secondary metabolism. Copies of the EFT-3m/Toru element appear to have mediated recombination events that may have abolished synthesis of two fungal alkaloids in different epichloae. This work provides insight into the potential impact of MITEs on epichloae evolution and provides a foundation for analysis in other fungal genomes.

Impact of Small Repeat Sequences on Bacterial Genome Evolution

Intergenic regions of prokaryotic genomes carry multiple copies of terminal inverted repeat (TIR) sequences, the nonautonomous miniature inverted-repeat transposable element (MITE). In addition, there are the repetitive extragenic palindromic (REP) sequences that fold into a small stem loop rich in G–C bonding. And the clustered regularly interspaced short palindromic repeats (CRISPRs) display similar small stem loops but are an integral part of a complex genetic element. Other classes of repeats such as the REP2 element do not have TIRs but show other signatures. With the current availability of a large number of whole-genome sequences, many new repeat elements have been discovered. These sequences display diverse properties. Some show an intimate linkage to integrons, and at least one encodes a small RNA. Many repeats are found fused with chromosomal open reading frames, and some are located within protein coding sequences. Small repeat units appear to work hand in hand with the transcriptional and/or post-transcriptional apparatus of the cell. Functionally, they are multifaceted, and this can range from the control of gene expression, the facilitation of host/pathogen interactions, or stimulation of the mammalian immune system. The CRISPR complex displays dramatic functions such as an acquired immune system that defends against invading viruses and plasmids. Evolutionarily, mobile repeat elements may have influenced a cycle of active versus inactive genes in ancestral organisms, and some repeats are concentrated in regions of the chromosome where there is significant genomic plasticity. Changes in the abundance of genomic repeats during the evolution of an organism may have resulted in a benefit to the cell or posed a disadvantage, and some present day species may reflect a purification process. The diverse structure, eclectic functions, and evolutionary aspects of repeat elements are described.

Genome-wide comparison of cyanobacterial transposable elements, potential genetic diversity indicators

Transposable elements are widely distributed in archaea, bacteria and eukarya domains. Considerable discrepancies of transposable elements in eukaryotes have been reported, however, the studies focusing on the diversity of transposable element systems in prokaryotes were scarce. Understanding the transposable element system in cyanobacteria by the genome-wide analysis will greatly improve the knowledge of cyanobacterial diversity. In this study, the transposable elements of seventeen cyanobacterial genomes were analyzed. The abundance of insertion sequence (IS) elements differs significantly among the cyanobacterial genomes examined. In particular, water bloom forming Microcystis aeruginosa NIES843 was shown to have the highest abundance of IS elements reaching 10.85% of the genome. IS family is a widely acceptable IS classification unit, and IS subfamily, based on probe sequences, was firstly proposed as the basic classification unit for IS element system，therefore both IS family and IS subfamily were suggested as the two hierarchical units for evaluating the IS element system diversity. In total, 1980 predicted IS elements, within 21 IS families and 132 subfamilies, were identified in the examined cyanobacterial genomes. Families IS4, IS5, IS630 and IS200-605 are widely distributed, and therefore supposed to be the ancestral IS families. Analysis on the intactness of IS elements showed that the percentage of the intact IS differs largely among these cyanobacterial strains. Higher percentage of the intact IS detected in the two hot spring cyanobacterial strains implied that the intactness of IS elements may be related to the genomic stabilization of cyanobacteria inhabiting in the extreme environments. The frequencies between IS elements and miniature inverted-repeat transposable elements (MITEs) were shown to have a linear positive correlation. The transposable element system in cyanobacterial genomes is of hypervariabilty. With characterization of easy definition and stability, IS subfamily is considered as a reliable lower classification unit in IS element system. The abundance of intact IS, the composition of IS families and subfamilies, the sequence diversity of IS element nucleotide and transposase amino acid are informative and suitable as the indicators for studies on cyanobacterial diversity. Practically, the transposable system may provide us a new perspective to realize the diversity and evolution of populations of water bloom forming cyanobacterial species.

Characterization of AhMITE1 transposition and its association with the mutational and evolutionary origin of botanical types in peanut (Arachis spp.)

AhMITE1 is an active miniature inverted repeat transposable element (MITE) in peanut (Arachis hypogaea L). Its transpositional activity from a particular (FST1-linked) site within the peanut genome was checked using AhMITE1-specifc PCR, which used a forward primer annealing to the 5′-flanking sequence and a reverse primer binding to AhMITE1. It was found that transposition activation was induced by stresses such as ethyl methane sulfonate (EMS), gamma irradiation, environmental conditions, and tissue culture. Excision and insertion of AhMITE1 at this particular site among the mutants led to gross morphological changes resembling alternate subspecies or botanical types. Analysis of South American landraces revealed the presence of AhMITE1 at the site among most of the spp. fastigiata types, whereas the element was predominantly missing from spp. hypogaea types, indicating its strong association. Four accessions of the primitive allotetraploid, A. monticola were devoid of AhMITE1 at the site, indicating only recent activation of the element, possibly because of the “genomic shock” resulting from hybridization followed by allopolyploidization.

A novel class of miniature inverted repeat transposable elements (MITEs) that contain hitchhiking (GTCY)n microsatellites

The movement of miniature inverted repeat transposable elements (MITEs) modifies genome structure and function. We describe the microsatellite-associated interspersed nuclear element 2 (MINE-2), that integrates at consensus WTTTT target sites, creates dinucleotide TT target site duplications (TSDs), and forms predicted MITE-like secondary structures; a 5' subterminal inverted repeat (SIR; AGGGTTCCGTAG) that is partially complementary to a 5' inverted repeat (IR; ACGAAGCCCT) and 3'-SIRs (TTACGGAACCCT). A (GTCY)(n) microsatellite is hitchhiking downstream of conserved 5'MINE-2 secondary structures, causing flanking sequence similarity amongst mobile microsatellite loci. Transfection of insect cell lines indicates that MITE-like secondary structures are sufficient to mediate genome integration, and provides insight into the transposition mechanism used by MINE-2s.

Analysis of SCAR marker nucleotide sequences in maize ( Zea mays L.) somaclones

SCAR (sequence characterized amplified region) markers allow the reliable identification of unique somaclonal variations. Six SCAR markers were developed previously and were thought to be exclusively characteristic of eight maize somaclones. However, we detected two of these markers in maize lines and a cultivar unrelated to the progenitor line of the somaclones. Therefore, we sequenced these markers and performed bioinformatic searches to understand the molecular events that may underlie the variability observed in the somaclones. All changes were found in noncoding sequences and were induced by different molecular events, such as the insertion of long terminal repeat (LTR) transposon(s), precise miniature inverted repeat transposable element (MITE) excision, microdeletion, recombination, and a change in the pool of mitochondrial DNA. For example, the SCAR marker QR is represented by the two variants QR-A and QR-2. The sequences of the two variants were similar, except for a 457-bp fragment found only in QR-A; this region was denoted as Q. Region Q was flanked by the direct 3-bp repeat 5′-TAA-3′ (target site duplication; TSD) and the inverted 14-bp repeat 5′-GGGCCTGTTTGGAA-3′ (terminal inverted repeats; TIRs). These features confer the Q region with similarity to the nonautonomic Tourist-like MITE. In two groups of independently produced somaclones, the same features (morphological, molecular) were variable, which confirms the theory of ‘hot spots’ occurring in the genome. The distribution of one of the SCAR markers was confirmed using Southern blot hybridization. The presence of the same molecular markers in the somaclones and in different non-somaclonal maize variants suggests that in some cases, the same mechanisms determine both in vitro and in vivo variability and that cell culture enhances the rate of heritable genomic changes that naturally occur in living organisms.

MITE-Hunter: a program for discovering miniature inverted-repeat transposable elements from genomic sequences

Miniature inverted-repeat transposable elements (MITEs) are a special type of Class 2 non-autonomous transposable element (TE) that are abundant in the non-coding regions of the genes of many plant and animal species. The accurate identification of MITEs has been a challenge for existing programs because they lack coding sequences and, as such, evolve very rapidly. Because of their importance to gene and genome evolution, we developed MITE-Hunter, a program pipeline that can identify MITEs as well as other small Class 2 non-autonomous TEs from genomic DNA data sets. The output of MITE-Hunter is composed of consensus TE sequences grouped into families that can be used as a library file for homology-based TE detection programs such as RepeatMasker. MITE-Hunter was evaluated by searching the rice genomic database and comparing the output with known rice TEs. It discovered most of the previously reported rice MITEs (97.6%), and found sixteen new elements. MITE-Hunter was also compared with two other MITE discovery programs, FINDMITE and MUST. Unlike MITE-Hunter, neither of these programs can search large genomic data sets including whole genome sequences. More importantly, MITE-Hunter is significantly more accurate than either FINDMITE or MUST as the vast majority of their outputs are false-positives.

Genome-wide comparison of cyanobacterial transposable elements, potential genetic diversity indicators

Abstract Background: Transposable elements are widely distributed in archaea, bacteria and eukarya domains. Considerable discrepancies of transposable elements in eukaryotes have been reported; however, the studies focusing on the diversity of transposable systems in prokaryotes were scarce. Understanding the transposable element system in cyanobacteria by the genome-wide analysis will greatly improve the knowledge of cyanobacterial diversity. Results: In this study, the transposable elements of seventeen cyanobacterial genomes were analyzed. The abundance of insertion sequence (IS) elements differs significantly among the cyanobacterial genomes examined. In particular, water bloom forming Microcystis aeruginosa NIES843 was shown to have the highest abundance of IS elements reaching 10.95% of the genome. IS family is a widely acceptable IS classification unit, and IS subfamily, based on probe sequences, was firstly proposed as the basic classification unit for IS element system. Both of IS family and IS subfamily were set as the two hierarchical units for evaluating the IS element system diversity. Totally, 1982 predicted IS elements, within 21 IS families and 133 subfamilies were identified in the examined cyanobacterial genomes. Families IS4, IS5, IS630 and IS200-605 are widely distributed, and therefore supposed to be the ancestral IS families. Analysis on the intactness of IS elements showed that the percentage of the intact IS differs largely among these cyanobacterial strains. Higher percentage of the intact IS detected in the two hot spring cyanobacterial strains implied that the intactness of IS elements may be related to the genomic stabilization of cyanobacteria inhabiting in the extreme environments. The frequencies between IS elements and miniature inverted-repeat transposable elements (MITEs) were shown to have a linear positive correlation. Conclusions: The transposable element system in cyanobacterial genomes is of hypervariabilty. With characterization of easy definition and stability, IS subfamily is considered as a reliable classification unit in IS element system. The abundance of intact IS, the composition of IS families and subfamilies, the sequence diversity of IS element nucleotide and transposase amino acid are informative and suitable as the indicators for studies on cyanobacterial diversity. Practically, the transposable system may provide us a new perspective to realize the diversity and evolution of populations of water bloom forming cyanobacterial species.

Non‐autonomous transposable elements associated with inactivation of microcystin gene clusters in strains of the genus Anabaena isolated from the Baltic Sea

Microcystins are potent peptide toxins produced by a range of distantly related cyanobacteria. They are assembled on a large enzyme complex encoded by the 55 kb microcystin synthetase (mcy) gene cluster. Here we report two strains of the genus Anabaena isolated from the Baltic Sea, which contain the entire mcy gene cluster but do not produce microcystins. Transcription analysis demonstrated that mcy genes were not expressed in these strains. We identified short insertion elements interrupting the mcyA, mcyD and mcyE genes in the two strains isolated in different years from different parts of the Baltic Sea. The 126-207 bp insertion elements encoded both terminal inverted repeats and direct repeats but lacked transposases. However, we found evidence for transposition of these elements despite the absence of genes encoding transposases, suggesting that they are non-autonomous mobile miniature inverted-repeat transposable elements (MITEs) recently described from cyanobacteria. The MITE insertion elements were present in mcyD genes amplified directly from the cyanobacterial community present in the Baltic Sea blooms from 2005. The results demonstrate that mcy gene cluster mutants can make up a stable proportion of the mcy gene pool in the Baltic Sea population.

The Insertion Sequences of Anabaena sp. Strain PCC 7120 and Their Effects on Its Open Reading Frames

Anabaena sp. strain PCC 7120, widely studied, has 145 annotated transposase genes that are part of transposable elements called insertion sequences (ISs). To determine the entirety of the ISs, we aligned transposase genes and their flanking regions; identified the ISs' possible terminal inverted repeats, usually flanked by direct repeats; and compared IS-interrupted sequences with homologous sequences. We thereby determined both ends of 87 ISs bearing 110 transposase genes in eight IS families (http://www-is.biotoul.fr/) and in a cluster of unclassified ISs, and of hitherto unknown miniature inverted-repeat transposable elements. Open reading frames were then identified to which ISs contributed and others--some encoding proteins of predictable function, including protein kinases, and restriction endonucleases--that were interrupted by ISs. Anabaena sp. ISs were often more closely related to exogenous than to other endogenous ISs, suggesting that numerous variant ISs were not degraded within PCC 7120 but transferred from without. This observation leads to the expectation that further sequencing projects will extend this and similar analyses. We also propose an adaptive role for poly(A) sequences in ISs.

Small RNAs, DNA methylation and transposable elements in wheat

BackgroundMore than 80% of the wheat genome is composed of transposable elements (TEs). Since active TEs can move to different locations and potentially impose a significant mutational load, their expression is suppressed in the genome via small non-coding RNAs (sRNAs). sRNAs guide silencing of TEs at the transcriptional (mainly 24-nt sRNAs) and post-transcriptional (mainly 21-nt sRNAs) levels. In this study, we report the distribution of these two types of sRNAs among the different classes of wheat TEs, the regions targeted within the TEs, and their impact on the methylation patterns of the targeted regions.ResultsWe constructed an sRNA library from hexaploid wheat and developed a database that included our library and three other publicly available sRNA libraries from wheat. For five completely-sequenced wheat BAC contigs, most perfectly matching sRNAs represented TE sequences, suggesting that a large fraction of the wheat sRNAs originated from TEs. An analysis of all wheat TEs present in the Triticeae Repeat Sequence database showed that sRNA abundance was correlated with the estimated number of TEs within each class. Most of the sRNAs perfectly matching miniature inverted repeat transposable elements (MITEs) belonged to the 21-nt class and were mainly targeted to the terminal inverted repeats (TIRs). In contrast, most of the sRNAs matching class I and class II TEs belonged to the 24-nt class and were mainly targeted to the long terminal repeats (LTRs) in the class I TEs and to the terminal repeats in CACTA transposons. An analysis of the mutation frequency in potentially methylated sites revealed a three-fold increase in TE mutation frequency relative to intron and untranslated genic regions. This increase is consistent with wheat TEs being preferentially methylated, likely by sRNA targeting.ConclusionsOur study examines the wheat epigenome in relation to known TEs. sRNA-directed transcriptional and post-transcriptional silencing plays important roles in the short-term suppression of TEs in the wheat genome, whereas DNA methylation and increased mutation rates may provide a long-term mechanism to inactivate TEs.

Characterization of a Group of MITEs with Unusual Features from Two Coral Genomes

BackgroundMiniature inverted-repeat transposable elements (MITEs), which are common in eukaryotic genomes, are small non-coding elements that transpose by utilizing transposases encoded by autonomous transposons. Recent genome-wide analyses and cross-mobilization assays have greatly improved our knowledge on MITE proliferation, however, specific mechanisms for the origin and evolution of MITEs are still unclear.Principal FindingsA group of coral MITEs called CMITE were identified from two corals, Acropora millepora and Acropora palmata. CMITEs conform to many common characteristics of MITEs, but also present several unusual features. The most unusual feature of CMITEs is conservation of the internal region, which is more conserved between MITE families than the TIRs. The origin of this internal region remains unknown, although we found one CMITE family that seems to be derived from a piggyBac-like transposon in A. millepora. CMITEs can form tandem arrays, suggesting an unconventional way for MITEs to increase copy numbers. We also describe a case in which a novel transposable element was created by a CMITE insertion event.ConclusionsTo our knowledge, this is the first report of identification of MITEs from coral genomes. Proliferation of CMITEs seems to be related to the transposition machinery of piggyBac-like autonomous transposons. The highly conserved internal region of CMITEs suggests a potential role for this region in their successful transposition. However, the origin of these unusual features in CMITEs remains unclear, and thus represents an intriguing topic for future investigations.

Transcript expression and regulatory characteristics of a rice glycosyltransferase OsGT61-1 gene

Glycosyltransferases (GTs) catalyze the reaction of glycosylation of various cellular macromolecules. Rice genome contains 609 potential GT genes which have been grouped into 40 gene families according to CAZy (Carbohydrate Active Enzyme) database. Phylogenetic tree showed that the members of GT61 family are related across genera. Close relationship among various rice GT61 proteins was also reflected from MEME analysis. Full-length cDNA corresponding to MSU rice locus Os02g22380 and encoding for xylosyltransferase was obtained (referred to as OsGT61-1). OsGT61-1 transcript was noted to be ABA- and NaCl-responsive and its expression was more prominent in root than in shoot tissues. Experiments involving fusion of OsGT61-1 promoter (1410 bp upstream to ATG) with GUS reporter gene showed that GUS expression was in consonance with the transcript expression. 1410 bp OsGT61-1 promoter contains two miniature inverted repeat transposable element sequences namely Gaijin (144 bp) and Wanderer (211 bp). Using yeast as trans-host, we noted that these sequences negatively influence the downstream transcript expression.

A helitron-like transposon superfamily from lepidoptera disrupts (GAAA)(n) microsatellites and is responsible for flanking sequence similarity within a microsatellite family.

Transposable elements (TEs) are mobile DNA regions that alter host genome structure and gene expression. A novel 588 bp non-autonomous high copy number TE in the Ostrinia nubilalis genome has features in common with miniature inverted-repeat transposable elements (MITEs): high A + T content (62.3%), lack of internal protein coding sequence, and secondary structure consisting of subterminal inverted repeats (SIRs). The O. nubilalis TE has inserted at (GAAA)(n) microsatellite loci, and was named the microsatellite-associated interspersed nuclear element (MINE-1). Non-autonomous MINE-1 superfamily members also were identified downstream of (GAAA)(n) microsatellites within Bombyx mori and Pectinophora gossypiella genomes. Of 316 (GAAA)(n) microsatellites from the B. mori whole genome sequence, 201 (63.6%) have associated autonomous or non-autonomous MINE-1 elements. Autonomous B. mori MINE-1s a encode a helicase and endonuclease domain RepHel-like protein (BMHELp1) indicating their classification as Helitron-like transposons and were renamed Helitron1_BM. Transposition of MINE-1 members in Lepidoptera has resulted in the disruption of (GAAA)(n) microsatellite loci, has impacted the application of microsatellite-based genetic markers, and suggests genome sequence that flanks TT/AA dinucleotides may be required for target site recognition by RepHel endonuclease domains.

Transposition of the Tourist-MITE mPing in yeast: an assay that retains key features of catalysis by the class 2 PIF/Harbinger superfamily

BackgroundPIF/Harbinger is the most recently discovered DNA transposon superfamily and is now known to populate genomes from fungi to plants to animals. Mobilization of superfamily members requires two separate element-encoded proteins (ORF1 and TPase). Members of this superfamily also mobilize Tourist-like miniature inverted repeat transposable elements (MITEs), which are the most abundant transposable elements associated with the genes of plants, especially the cereal grasses. The phylogenetic analysis of many plant genomes indicates that MITEs can amplify rapidly from one or a few elements to hundreds or thousands.The most active DNA transposon identified to date in plants or animals is mPing, a rice Tourist-like MITE that is a deletion derivative of the autonomous Ping element. Ping and the closely related Pong are the only known naturally active PIF/Harbinger elements. Some rice strains accumulate ~40 new mPing insertions per plant per generation. In this study we report the development of a yeast transposition assay as a first step in deciphering the mechanism underlying the amplification of Tourist-MITEs.ResultsThe ORF1 and TPase proteins encoded by Ping and Pong have been shown to mobilize mPing in rice and in transgenic Arabidopsis. Initial tests of the native proteins in a yeast assay resulted in very low transposition. Significantly higher activities were obtained by mutation of a putative nuclear export signal (NES) in the TPase that increased the amount of TPase in the nucleus. When introduced into Arabidopsis, the NES mutant protein also catalyzed higher frequencies of mPing excision from the gfp reporter gene. Our yeast assay retains key features of excision and insertion of mPing including precise excision, extended insertion sequence preference, and a requirement for two proteins that can come from either Ping or Pong or both elements.ConclusionsThe yeast transposition assay provides a robust platform for analysis of the mechanism underlying transposition catalyzed by the two proteins of PIF/Harbinger elements. It recapitulates all of the features of excision and reinsertion of mPing as seen in plant systems. Furthermore, a mutation of a putative NES in the TPase increased transposition both in yeast and plants.

A systematic search and classification of T2 family miniature inverted-repeat transposable elements (MITEs) in Xenopus tropicalis suggests the existence of recently active MITE subfamilies

To reveal the genome-wide aspects of Xenopus T2 family miniature inverted-repeat transposable elements (MITEs), we performed a systematic search and classification of MITEs by a newly developed procedure. A terminal sequence motif (T2-motif: TTAAAGGRR) was retrieved from the Xenopus tropicalis genome database. We then selected 51- to 1,000-bp MITE candidates framed by an inverted pair of 2 T2-motifs. The 34,398 candidates were classified into possible clusters by a novel terminal sequence (TS)-clustering method on the basis of differences in their short terminal sequences. Finally, 19,242 MITEs were classified into 16 major MITE subfamilies (TS subfamilies), 10 of which showed apparent homologies to known T2 MITE subfamilies, and the rest were novel TS subfamilies. Intra- and inter-subfamily similarities or differences were investigated by analyses of diversity in GC content, total length, and sequence alignments. Furthermore, genome-wide conservation of the inverted pair structure of subfamily-specific TS stretches and their target site sequence (TTAA) were analyzed. The results suggested that some TS subfamilies might include active or at least recently active MITEs for transposition and/or amplification, but some others might have lost such activities a long time ago. The present methodology was efficient in identifying and classifying MITEs, thereby providing information on the evolutionary dynamics of MITEs.

Identification of NbME MITE families: Potential molecular markers in the microsporidia Nosema bombycis

Six novel families of miniature inverted-repeat transposable elements (MITEs) were characterized in the microsporidia Nosema bombycis and were named NbMEs. The structural characteristics and the distribution of NbME copies in the N. bombycis genome were investigated, and it was found that portions of NbMEs are associated with gene sections. Potential molecular markers for various N. bombycis strains were identified in this study through utilization of the MITE-AFLP technique. Three distinct pathogenic isolates collected from different areas were distinguished, and polymorphisms were detected using the NbME5 marker, thereby establishing this NbME as a potential marker for studying isolate variation in N. bombycis.