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A Small RNA Derived From the 5' End of the IS200 tnpA Transcript Regulates Multiple Virulence Regulons in Salmonella typhimurium.

The insertion sequence IS200 is widely distributed in Eubacteria. Despite its prevalence, IS200 does not appear to be mobile and as such is considered an ancestral component of bacterial genomes. Previous work in Salmonella enterica revealed that the IS200 tnpA transcript is processed to form a small, highly structured RNA (5'tnpA) that participates in the posttranscriptional control of invF expression, encoding a key transcription factor in this enteropathogen's invasion regulon. To further examine the scope of 5'tnpA transcript integration into Salmonella gene expression networks, we performed comparative RNA-seq, revealing the differential expression of over 200 genes in a Salmonella SL1344 5'tnpA disruption strain. This includes the genes for the master regulators of both invasion and flagellar regulons (HilD and FlhDC, respectively), plus genes involved in cysteine biosynthesis and an operon (phsABC) encoding a thiosulfate reductase complex. These expression changes were accompanied by an 80-fold increase in Salmonella invasion of HeLa cells. Follow-up experimentation suggested an additional direct target of 5'tnpA to be the small RNA PinT, which has previously been shown to be a negative regulator of invasion genes through its inhibitory action on key transcription factors governing the Salmonella pathogenicity island 1 regulon. This study provides a powerful new example of bacterial transposon domestication that is based not on the production/use of a regulatory protein or regulatory DNA sequences, but on the function of a transposon-derived small RNA.

Salmonella typhimurium↗

The evolutionary life history of P transposons: from horizontal invaders to domesticated neogenes.

P elements, a family of DNA transposons, are known as aggressive intruders into the hitherto uninfected gene pool of Drosophila melanogaster. Invading through horizontal transmission from an external source they managed to spread rapidly through natural populations within a few decades. Owing to their propensity for rapid propagation within genomes as well as within populations, they are considered as the classic example of selfish DNA, causing havoc in a genomic environment permissive for transpositional activity. Tracing the fate of P transposons on an evolutionary scale we describe different stages in their evolutionary life history. Starting from horizontal transfer events, which now appear to be rather a common phenomenon, the initial transpositional burst in the new host is slowed down by the accumulation of defective copies as well as host-directed epigenetic silencing. This leads to the loss of mobility and, finally, to molecular erosion by random mutations. Possible escape routes from genomic extinction are the reactivation within the original host genome by recombination or suspension of the repressing regime, horizontal emigration to a virgin gene pool, or genomic integration and acquisition of a novel function as a domesticated host gene.

Animals↗

Molecular evolutionary analysis of the widespread piggyBac transposon family and related "domesticated" sequences.

piggyBac is a short inverted-repeat-type DNA transposable element originally isolated from the genome of the moth Trichoplusia ni. It is currently the gene vector of choice for the transformation of various insect species. A few sequences with similarity to piggyBac have previously been identified from organisms such as humans ( Looper), the pufferfish Takifugu rubripes ( Pigibaku), Xenopus ( Tx), Daphnia ( Pokey), and the Oriental fruit fly Bactrocera dorsalis. We have now identified 50 piggyBac-like sequences from publicly available genome sequences and expressed sequence tags (ESTs). This survey allows the first comparative examination of the distinctive piggyBac transposase, suggesting that it might contain a highly divergent DDD domain, comparable to the widespread DDE domain found in many DNA transposases and retroviral integrases which consists of two absolutely conserved aspartic acids separated by about 70 amino acids with a highly conserved glutamic acid about 35 amino acids further away. Many piggyBac-like sequences were found in the genomes of a phylogenetically diverse range of organisms including fungi, plants, insects, crustaceans, urochordates, amphibians, fishes and mammals. Also, several instances of "domestication" of the piggyBac transposase sequence by the host genome for cellular functions were identified. Novel members of the piggyBac family may be useful in genetic engineering of many organisms.

Animals↗

The implications of DNA transposons in the evolution of P elements in zebrafish (Danio rerio).

We have carried out detailed analyses of Drosophila P-transposon-homologous Pdre elements in zebrafish (Danio rerio) and present a model to explain the evolutionary origin of the complete and terminally truncated copies. Analysis of the most complex terminally truncated element led to the identification of novel DNA transposons, referred to as demio, hexi, and lady elements. We also show that the existence of a P-homologous stable gene can be explained in two ways: it is a P-transposon source gene, and insertion into or recombination with another transposon can lead to mobilization by recruitment of structures necessary for transposition (known as the source gene hypothesis), or it is the result of a domestication event involving a P transposon (domestication hypothesis). Our results indicate that both scenarios are possible and that they reflect different stages of the P-element life cycle.

Animals↗

Homologs of Drosophila P transposons were mobile in zebrafish but have been domesticated in a common ancestor of chicken and human.

A substantial fraction of vertebrate and invertebrate genomes is composed of mobile elements and their derivatives. One of the most intensively studied transposon families, the P elements of Drosophila, was thought to exist exclusively in the genomes of dipteran insects. Based on the data provided by the human genome project, in 2001 our group has identified a P element-homologous sequence in the human genome. This P element-homologous human gene, named Phsa, is 19,533 nucleotides long, comprises six exons and five introns, and encodes a protein of still unknown function with a length of 903 amino acid residues. The N-terminal THAP domain of the putative Phsa protein shows similarities to the site-specific DNA-binding domain of the Drosophila P element transposase. In the present study, FISH analysis and the screening of a human lambda genomic library revealed a single copy of Phsa located on the long arm of chromosome 4, upstream of a gene coding for the hypothetical protein DKFZp686L1814. The same gene arrangement was found for the homologous gene Pgga in the genome of chicken, thus, displaying Pgga at orthologous position on the long arm of chromosome 4. The single-copy gene status and the absence of terminal inverted repeats and target-site duplications indicate that Phsa and Pgga constitute domesticated stationary sequences. In contrast, a considerable number of P-homologous sequences with terminal inverted repeats and intact target-site duplications could be identified in zebrafish, strongly indicating that Pdre elements were mobile within the zebrafish genome. Pdre elements are the first P-like transposons identified in a vertebrate species. With respect to Phsa, gene expression studies showed that Phsa is expressed in a broad range of human tissues, suggesting that the putative Phsa protein plays a not yet understood but essential role in a specific metabolic pathway. We demonstrate that P-homologous DNA sequences occur in the genomes of 21 analyzed vertebrates but only as rudiments in the rodents. Finally, the evolutionary history of P element-homologous vertebrate sequences is discussed in the context of the "molecular domestication" hypothesis versus the "source gene hypothesis."

Amino Acid Sequence↗

Chloramphenicol transposons found in Salmonella naestved and Escherichia coli of domestic animal origin.

Salmonella naestved strain AHI-21, of calf origin, harbors a conjugative R plasmid of group H1, pTE21, which encodes resistance to chloramphenicol (Cm), tetracycline, streptomycin, and sulfadimethoxine. Escherichia coli strain AHI-1, of pig origin, also harbors a conjugative R plasmid of group I alpha, pTE1, which encodes resistance to chloramphenicol and trimethoprim. When either of these R plasmids coexisted with a nonconjugative plasmid, pMK1, which is a composite plasmid of ColE1 and a kanamycin transposon (Tn5), transposition of the Cmr gene into pMK1 occurred independently of the host recA gene function, indicating that both R plasmids contained Cm transposons, Tn3351 and Tn3352. Electron microscopic analysis of self-annealed and heteroduplex molecules showed that they were of approximately 1.7 megadaltons in size and were inserted within the ColE1 loop region of pMK1. However, inverted repeat structures were not seen in these two Cm transposons. Restriction enzyme cleavage analysis showed that both Tn3351 and Tn3352 were indistinguishable in their cleavage patterns, suggesting that they were almost identical in deoxyribonucleic acid sequence despite the difference in their origin. These results suggest that the reciprocal transposition of the Cmr gene might have occurred between Salmonella and E. coli in nature.

Chloramphenicol↗

MUSTANG is a novel family of domesticated transposase genes found in diverse angiosperms.

While transposons have traditionally been viewed as genomic parasites or "junk DNA," the discovery of transposon-derived host genes has fueled an ongoing debate over the evolutionary role of transposons. In particular, while mobility-related open reading frames have been known to acquire host functions, the contribution of these types of events to the evolution of genes is not well understood. Here we report that genome-wide searches for Mutator transposase-derived host genes in Arabidopsis thaliana (Columbia-0) and Oryza sativa ssp. japonica (cv. Nipponbare) (domesticated rice) identified 121 sequences, including the taxonomically conserved MUSTANG1. Syntenic MUSTANG1 orthologs in such varied plant species as rice, poplar, Arabidopsis, and Medicago truncatula appear to be under purifying selection. However, despite the evidence of this pathway of gene evolution, MUSTANG1 belongs to one of only two Mutator-like gene families with members in both monocotyledonous and dicotyledonous plants, suggesting that Mutator-like elements seldom evolve into taxonomically widespread host genes.

Arabidopsis↗

A deep-branching clade of retrovirus-like retrotransposons in bdelloid rotifers.

Rotifers of class Bdelloidea, a group of aquatic invertebrates in which males and meiosis have never been documented, are also unusual in their lack of multicopy LINE-like and gypsy-like retrotransposons, groups inhabiting the genomes of nearly all other metazoans. Bdelloids do contain numerous DNA transposons, both intact and decayed, and domesticated Penelope-like retroelements Athena, concentrated at telomeric regions. Here we describe two LTR retrotransposons, each found at low copy number in a different bdelloid species, which define a clade different from previously known clades of LTR retrotransposons. Like bdelloid DNA transposons and Athena, these elements are found preferentially in telomeric regions. Unlike bdelloid DNA transposons, many of which are decayed, the newly described elements, named Vesta and Juno, inhabiting the genomes of Philodina roseola and Adineta vaga, respectively, appear to be intact and represent recent insertions, possibly from an exogenous source. We describe the retrovirus-like structure of the new elements, containing gag, pol, and env-like open reading frames, and discuss their possible origins, transmission, and behavior in bdelloid genomes.

Amino Acid Sequence↗

Transposable Element Dynamics Drive the Genomic Evolution and Phenotypic Diversification of Allotetraploid Common Carp.

An important question in evolutionary biology is how polyploidization generates raw material for phenotypic diversification. Transposable elements (TEs) represent an underestimated source of genetic variation in eukaryotic genomes. By integrating 516 whole-genome resequencing datasets and 236 transcriptomes from common carp (Cyprinus carpio), a representative allotetraploid fish, we constructed the first population-scale landscape of TE insertions in teleosts. TE insertions are widespread in the carp genome and preferentially associated with stress-responsive genes, with DNA transposons as major contributors. Relaxed purifying selection and TE burst events coexist, generating abundant variation for subsequent subspecies differentiation. Compared with a closely related diploid species, carp exhibits more exonic TE insertions and shorter TE-gene distances, and multiple TE superfamilies expanded during tetraploidization. Genome-wide association analyses uncovered intragenic TE variants underlying domesticated traits missed by SNPs, including DNA transposon deletions associated with scale reduction and altered body shape. Notably, lighter-colored individuals harbor homozygous deletions of LTR and DNA transposons within mdfic2, whose knockout in zebrafish reduces pigmentation. Most trait-associated variants reflect lineage-specific loss of ancient TE insertions rather than recent transposition. Overall, these findings highlight the distinct role of TEs in polyploid genome evolution and phenotypic diversification, providing new insights into TE dynamics in vertebrates.

allotetraploidization↗

Evidence for dual pathways of Tc1/mariner domestication in Drosophila.

BACKGROUND: The domestication of transposable elements is a key source of evolutionary innovation, yet the pathways by which their functional modules are repurposed by the host remain poorly understood. The Tc1/mariner superfamily is a widespread group of DNA transposons, but the prevalence and patterns of their domestication are underexplored. RESULTS: We performed a systematic genomic screen across 43 drosophilid species using stringent criteria for molecular domestication. This analysis identified five high-confidence, evolutionarily conserved genes derived from Tc1/mariner transposases. Phylogenetic and structural analyses suggest domestication via two distinct molecular pathways: co-option of the DNA-binding module and co-option of the catalytic domain. The DNA-binding module pathway includes CG4570, the previously known genes cag and toy (the latter fused with a homeodomain), and a lineage-restricted gene in the Drosophila obscura group that exhibits signatures of recent domestication. In contrast, the catalytic domain pathway is represented solely by CG14478. Structural modeling reveals that CG14478 protein preserves a canonical DDE endonuclease fold. Co-expression network analysis suggests potential cellular roles of these genes: CG14478 is linked to RNA/chromatin-related processes, CG4570 to cell cycle/chromosome functions, cag to ciliary and nuclear functions, and toy to neuronal development. CONCLUSIONS: This study establishes a stringent framework for identifying domesticated TEs, demonstrating that Tc1/mariner elements are co-opted via two distinct pathways: retention of either catalytic or DNA-binding modules. Our findings suggest that domestication is a dynamic continuum, ranging from recent, lineage-specific events to ancient, conserved genes, and underscore how genomic conflict with TEs can drive eukaryotic evolution and regulatory complexity.

Animals↗

Diverse DNA transposons in rotifers of the class Bdelloidea.

We surveyed the diversity, structural organization, and patterns of evolution of DNA transposons in rotifers of the class Bdelloidea, a group of basal triploblast animals that appears to have evolved for millions of years without sexual reproduction. Representatives of five superfamilies were identified: ITm (IS630/Tc/mariner), hAT, piggyBac, helitron, and foldback. Except for mariners, no fully intact copies were found. Mariners, both intact and decayed, are present in high copy number, and those described here may be grouped in several closely related lineages. Comparisons across lineages show strong evidence of purifying selection, whereas there is little or no evidence of such selection within lineages. This pattern could have resulted from repeated horizontal transfers from an exogenous source, followed by limited intragenomic proliferation, or, less plausibly, from within-host formation of new lineages under host- or element-based selection for function, in either case followed by eventual inactivation and decay. Unexpectedly, the flanking sequences surrounding the majority of mariners are very similar, indicating either insertion specificity or proliferation as part of larger DNA segments. Members of all superfamilies are present near chromosome ends, associated with the apparently domesticated retroelement Athena, in large clusters composed of diverse DNA transposons, often inserted into each other, whereas the examined gene-rich regions are nearly transposon-free.

Acari↗

Flower color variation: a model for the experimental study of evolution.

We review the study of flower color polymorphisms in the morning glory as a model for the analysis of adaptation. The pathway involved in the determination of flower color phenotype is traced from the molecular and genetic levels to the phenotypic level. Many of the genes that determine the enzymatic components of flavonoid biosynthesis are redundant, but, despite this complexity, it is possible to associate discrete floral phenotypes with individual genes. An important finding is that almost all of the mutations that determine phenotypic differences are the result of transposon insertions. Thus, the flower color diversity seized on by early human domesticators of this plant is a consequence of the rich variety of mobile elements that reside in the morning glory genome. We then consider a long history of research aimed at uncovering the ecological fate of these various flower phenotypes in the southeastern U.S. A large body of work has shown that insect pollinators discriminate against white phenotypes when white flowers are rare in populations. Because the plant is self-compatible, pollinator bias causes an increase in self-fertilization in white maternal plants, which should lead to an increase in the frequency of white genes, according to modifier gene theory. Studies of geographical distributions indicate other, as yet undiscovered, disadvantages associated with the white phenotype. The ultimate goal of connecting ecology to molecular genetics through the medium of phenotype is yet to be attained, but this approach may represent a model for analyzing the translation between these two levels of biological organization.

Journal Article↗

Genomic and evolutionary analysis reveals dynamic variations of MKK3 gene, a key regulator for seed dormancy in barley.

Barley (Hordeum vulgare L.) is an important crop in the world, and its seed dormancy is primarily controlled by a mitogen-activated protein kinase kinase 3 (MKK3) gene. Although kinase activity of MKK3 and its roles in barley post-domestication have been widely studied, the pre-domestication evolution of MKK3 and the spread of nondormant alleles among global barley varieties remain largely unexplored. In this study, we analyzed MKK3 sequences in barley and its wild progenitor (Hordeum spontaneum K. Koch) and identified two polymorphic miniature inverted-repeat transposable elements (MITEs). Comparative analyses indicated that the insertions/excision of the MITEs predated the current estimates of barley domestication. Examination of the barley pangenomes coupled with droplet digital polymerase chain reaction revealed extensive copy number variation of MKK3 and suggested that transposons likely contributed to tandem amplification of the MKK3 gene on chromosome 5H. Additionally, approximately 1-Kb MKK3 sequences were found on chromosomes 1H and 6H. Further analysis indicated that these short MKK3 sequences were captured by a CACTA transposon that also contained fragments from four other expressed genes. The acquisition of MKK3 was estimated to be between 1.9 and 2.5 million years ago. Together, these findings illuminate the dynamic pre-domestication evolution of the MKK3 gene and identify three divergent MKK3 haplotype groups including a unique lineage predominant in Ethiopian germplasm. This study highlights the contribution of transposons to structural diversification and evolutionary differentiation of the MKK3 locus and provides helpful information for understanding the complex history of MKK3 gene in barley and also for improving preharvest sprouting tolerant varieties under distinct natural conditions.

Hordeum↗

An active DNA transposon family in rice.

The publication of draft sequences for the two subspecies of Oryza sativa (rice), japonica (cv. Nipponbare) and indica (cv. 93-11), provides a unique opportunity to study the dynamics of transposable elements in this important crop plant. Here we report the use of these sequences in a computational approach to identify the first active DNA transposons from rice and the first active miniature inverted-repeat transposable element (MITE) from any organism. A sequence classified as a Tourist-like MITE of 430 base pairs, called miniature Ping (mPing), was present in about 70 copies in Nipponbare and in about 14 copies in 93-11. These mPing elements, which are all nearly identical, transpose actively in an indica cell-culture line. Database searches identified a family of related transposase-encoding elements (called Pong), which also transpose actively in the same cells. Virtually all new insertions of mPing and Pong elements were into low-copy regions of the rice genome. Since the domestication of rice mPing MITEs have been amplified preferentially in cultivars adapted to environmental extremes-a situation that is reminiscent of the genomic shock theory for transposon activation.

Amino Acid Sequence↗

Isolation and characterization of mini-Tn5Km2 insertion mutants of Brucella abortus deficient in internalization and intracellular growth in HeLa cells.

Brucella spp. are facultative intracellular pathogens that have the ability to survive and multiply in professional and nonprofessional phagocytes and cause abortion in domestic animals and undulant fever in humans. The mechanism and factors of virulence are not fully understood. To identify genes related to internalization and multiplication in host cells, Brucella abortus was mutagenized by mini-Tn5Km2 transposon that carryied the kanamycin resistance gene, 4,400 mutants were screened, and HeLa cells were infected with each mutant. Twenty-three intracellular-growth-defective mutants were screened and were characterized for internalization and intracellular growth. From these results, we divided the mutants into the following three groups: class I, no internalization and intracellular growth within HeLa cells; class II, an internalization similar to that of the wild type but with no intracellular growth; and class III, internalization twice as high as the wild type but with no intracellular growth. Sequence analysis of DNA flanking the site of transposon showed various insertion sites of bacterial genes that are virulence-associated genes, including virB genes, an ion transporter system, and biosynthesis- and metabolism-associated genes. These internalization and intracellular-growth-defective mutants in HeLa cells also showed defective intracellular growth in macrophages. These results suggest that the virulence-associated genes isolated here contributed to the intracellular growth of both nonprofessional and professional phagocytes.

ATP-Binding Cassette Transporters↗

Transposable elements reveal the impact of introgression, rather than transposition, in Pisum diversity, evolution, and domestication.

The genetic structure and evolutionary history of the genus Pisum were studied exploiting our germplasm collection to compare the contribution of different mechanisms to the generation of diversity. We used sequence-specific amplification polymorphism (SSAP) markers to assess insertion site polymorphism generated by a representative of each of the two major groups of LTR-containing retrotransposons, PDR1 (Ty1/copia-like) and Cyclops (Ty3/gypsy-like), together with Pis1, a member of the En/Spm transposon superfamily. The analysis of extended sets of the four main Pisum species, P. fulvum, P. elatius, P. abyssinicum, and P. sativum, together with the reference set, revealed a distinct pattern of the NJ (Neighbor-Joining) tree for each basic lineage, which reflects the different evolutionary history of each species. The SSAP markers showed that Pisum is exceptionally polymorphic for an inbreeding species. The patterns of phylogenetic relationships deduced from different transposable elements were in general agreement. The retrotransposon-derived markers gave a clearer separation of the main lineages than the Pis1 markers and were able to distinguish the truly wild form of P. elatius from the antecedents of P. sativum. There were more species-specific and unique PDR1 markers than Pis1 markers in P. fulvum and P. elatius, pointing to PDR1 activity during speciation and diversification, but the proportion of these markers is low. The overall genetic diversity of Pisum and the extreme polymorphism in all species, except P. abyssinicum, indicate a high contribution of recombination between multiple ancestral lineages compared to transposition within lineages. The two independently domesticated pea species, P. abyssinicum and P. sativum, arose in contrasting ways from the common processes of hybridization, introgression, and selection without associated transpositional activity.

DNA Transposable Elements↗

Domestication of wild Saccharomyces cerevisiae is accompanied by changes in gene expression and colony morphology.

Although colonies from Saccharomyces cerevisiae laboratory strains are smooth, those isolated from nature exhibit a structured fluffy pattern. Environmental scanning electron microscopy shows that the cells within wild fluffy colonies are connected by extracellular matrix (ECM) material. This material contains a protein of about 200 kDa unrelated to the flocculins, proteins involved in cell-cell adhesion in liquid media. The matrix material binds to concanavalin A. Within a few passages on rich agar medium, the wild strains switch from the fluffy to the smooth colony morphology. This domestication is accompanied by loss of the ECM and by extensive changes in gene expression as detected by DNA microarrays. The expression of about 320 genes was changed in smooth colonies. The major changes comprise carbohydrate metabolism, cell wall, water channels, Ty-transposons and subtelomeric genes, iron homeostasis, vitamin metabolism and cell cycle and polarity. The growth in fluffy colonies may represent a metabolic strategy for survival of yeast under unfavourable conditions that is switched off under felicitous laboratory conditions.

Computational Biology↗

Unraveling Brucella genomics and pathogenesis in immunocompromised IRF-1-/- mice.

PROBLEM: Brucellosis causes abortion in domestic animals and Malta fever in humans. Comparison of Brucella species genomes may reveal potential virulence mechanisms. Engineering bioluminescent Brucella would permit monitoring bacterial dissemination. METHOD OF STUDY: Microarray of the B. melitensis genome allowed comparison of gene content from six Brucella species. Bioluminescent B. melitensis strains were developed using transposon mutagenesis permitting the study of pathogenic Brucella in mice. Monitoring bacterial dissemination as well as organ localization permits evaluating the role of genes and genomic islands in mutant bacteria. RESULTS: Comparative genomic analysis revealed 217 ORFs altered in five Brucella species and were often found in islands. Bioluminescent bacteria disseminated from the injection site to liver, spleen, inguinal lymph nodes, testes and submanibular region. CONCLUSIONS: Genomic islands contribute to Brucella pathogenicity. Biophotonic imaging suggests that Brucella dissemination in mice parallels acute and chronic infections of humans.

Animals↗