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Insertion sequences are compact and pervasive transposable elements found in bacteria, which encode only the genes necessary for their mobilization and maintenance1. IS200- and IS605-family transposons undergo ‘peel-and-paste’ transposition catalysed by a TnpA transposase2, but they also encode diverse, TnpB- and IscB-family proteins that are evolutionarily related to the CRISPR-associated effectors Cas12 and Cas9, respectively3,4. Recent studies have demonstrated that TnpB and IscB function as RNA-guided DNA endonucleases5,6, but the broader biological role of this activity has remained enigmatic. Here we show that TnpB and IscB are essential to prevent permanent transposon loss as a consequence of the TnpA transposition mechanism. We selected a family of related insertion sequences from Geobacillus stearothermophilus that encode several TnpB and IscB orthologues, and showed that a single TnpA transposase was broadly active for transposon mobilization. The donor joints formed upon religation of transposon-flanking sequences were efficiently targeted for cleavage by RNA-guided TnpB and IscB nucleases, and co-expression of TnpB and TnpA led to substantially greater transposon retention relative to conditions in which TnpA was expressed alone. Notably, TnpA and TnpB also stimulated recombination frequencies, surpassing rates observed with TnpB alone. Collectively, this study reveals that RNA-guided DNA cleavage arose as a primal biochemical activity to bias the selfish inheritance and spread of transposable elements, which was later co-opted during the evolution of CRISPR–Cas adaptive immunity for antiviral defence. TnpB and IscB nucleases use transposon-encoded guide RNAs to target genomic sequences for cleavage, thereby favouring copying and spreading of transposable elements.
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Jump around: transposons in and out of the laboratory
Since Barbara McClintock’s groundbreaking discovery of mobile DNA sequences some 70 years ago, transposable elements have come to be recognized as important mutagenic agents impacting genome composition, genome evolution, and human health. Transposable elements are a major constituent of prokaryotic and eukaryotic genomes, and the transposition mechanisms enabling transposon proliferation over evolutionary time remain engaging topics for study, suggesting complex interactions with the host, both antagonistic and mutualistic. The impact of transposition is profound, as over 100 human heritable diseases have been attributed to transposon insertions. Transposition can be highly mutagenic, perturbing genome integrity and gene expression in a wide range of organisms. This mutagenic potential has been exploited in the laboratory, where transposons have long been utilized for phenotypic screening and the generation of defined mutant libraries. More recently, barcoding applications and methods for RNA-directed transposition are being used towards new phenotypic screens and studies relevant for gene therapy. Thus, transposable elements are significant in affecting biology both in vivo and in the laboratory, and this review will survey advances in understanding the biological role of transposons and relevant laboratory applications of these powerful molecular tools.
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- Award ID(s):
- 1902359
- PAR ID:
- 10162304
- Date Published:
- Journal Name:
- F1000Research
- Volume:
- 9
- ISSN:
- 2046-1402
- Page Range / eLocation ID:
- 135
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Introduction: Class II DNA transposable elements account for significant portions of eukaryotic genomes and contribute to genome evolution through their mobilization. To escape inactivating mutations and persist in the host genome over evolutionary time, these elements must be mobilized enough to result in additional copies. These elements utilize a “cut and paste” transposition mechanism that does not intrinsically include replication. However, elements such as the rice derived mPing element have been observed to increase in copy number over time. Methods: We used yeast transposition assays to test several parameters that could affect the excision and insertion of mPing and its related elements. This included development of novel strategies for measuring element insertion and sequencing insertion sites. Results: Increased transposase protein expression increased the mobilization frequency of a small (430 bp) element, while overexpression inhibition was observed for a larger (7,126 bp) element. Smaller element size increased both the frequency of excision and insertion of these elements. The effect of yeast ploidy on element excision, insertion, and copy number provided evidence that homology dependent repair allows for replicative transposition. These elements were found to preferentially insert into yeast rDNA repeat sequences. Discussion: Identifying the parameters that influence transposition of these elements will facilitate their use for gene discovery and genome editing. These insights in to the behavior of these elements also provide important clues into how class II transposable elements have shaped eukaryotic genomes.more » « less
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Abstract Protein translation is tightly and precisely controlled by multiple mechanisms including upstream open reading frames (uORFs), but the origins of uORFs and their role in maize are largely unexplored. In this study, an active transposition event was identified during the propagation of maize inbred line B73. The transposon, which was named BTA for ‘B73 active transposable element hAT’, creates a novel dosage-dependent hypomorphic allele of the hexose transporter gene ZmSWEET4c through insertion within the coding sequence in the first exon, and results in reduced kernel size. The BTA insertion does not affect transcript abundance but reduces protein abundance of ZmSWEET4c, probably through the introduction of a uORF. Furthermore, the introduction of BTA sequence in the exon of other genes can regulate translation efficiency without affecting their mRNA levels. A transposon capture assay revealed 79 novel insertions for BTA and BTA-like elements. These insertion sites have typical euchromatin features, including low levels of DNA methylation and high levels of H3K27ac. A putative autonomous element that mobilizes BTA and BTA-like elements was identified. Together, our results suggest a transposon-based origin of uORFs and document a new role for transposable elements to influence protein abundance and phenotypic diversity by affecting the translation rate.more » « less
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Arkhipova, Irina (Ed.)Abstract Horizontal transfer of transposable elements (TEs) is an important mechanism contributing to genetic diversity and innovation. Bats (order Chiroptera) have repeatedly been shown to experience horizontal transfer of TEs at what appears to be a high rate compared with other mammals. We investigated the occurrence of horizontally transferred (HT) DNA transposons involving bats. We found over 200 putative HT elements within bats; 16 transposons were shared across distantly related mammalian clades, and 2 other elements were shared with a fish and two lizard species. Our results indicate that bats are a hotspot for horizontal transfer of DNA transposons. These events broadly coincide with the diversification of several bat clades, supporting the hypothesis that DNA transposon invasions have contributed to genetic diversification of bats.more » « less
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Abstract The duplication of genes has long been recognized as a substrate for evolutionary novelty and adaptation, but the factors that govern fixation of paralogs soon after duplication are only partially understood. Duplication often leads to an increase in gene dosage, or the amount of functional gene product. For genes with which an increased dosage is harmful (i.e., triplosensitive genes), a dosage balancing mechanism needs to be present immediately after duplication if it is to evade negative selection. Previous research in vertebrates has demonstrated a potential role for epigenetic factors in allowing triplosensitive genes to increase in copy number by regulating their expression post-duplication. Here we expand this research by investigating the epigenetic landscape of duplicate genes inD. discoideum, a basal lineage separated from humans by over a billion years. We found that activating histone modifications are quickly lost in duplicate genes before gradually increasing in enrichment as paralogs age. For the repressive modification H3K9me3, we found it was enriched in the youngest paralogs, and that this enrichment was likely mediated by heterochromatin spread from transposable elements. We similarly found enrichment of H3K9me3 in young human duplicates, and again found transposable elements as a potential mediator. Finally, we leveraged recent genome-wide estimates of triplosensitivity in human genes to directly examine the relationship between this kind of dosage sensitivity and enrichment for repressive histone modifications. Interestingly, while we found no significant link between enrichment for the repressive mark H3K9me3 and triplosensitivity in human paralogs, we did find a significant association between triplosensitivity and transposon proximity. Our findings suggest that transposons may contribute to the epigenetic regulatory environment associated with dosage balancing of young duplicates in both protists and humans.more » « less
- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.12688/f1000research.21018.1
