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Nucleic Acids Research Sep 2023Dot1l is a histone methyltransferase without a SET domain and is responsible for H3K79 methylation, which marks active transcription. In contradiction, Dot1l also...
Dot1l is a histone methyltransferase without a SET domain and is responsible for H3K79 methylation, which marks active transcription. In contradiction, Dot1l also participates in silencing gene expression. The target regions and mechanism of Dot1l in repressing transcription remain enigmatic. Here, we show that Dot1l represses endogenous retroviruses in embryonic stem cells (ESCs). Specifically, the absence of Dot1l led to the activation of MERVL, which is a marker of 2-cell-like cells. In addition, Dot1l deletion activated the 2-cell-like state and predisposed ESCs to differentiate into trophectoderm lineage. Transcriptome analysis revealed activation of 2-cell genes and meiotic genes by Dot1l deletion. Mechanistically, Dot1l interacted with and co-localized with Npm1 on MERVL, and depletion of Npm1 similarly augmented MERVL expression. The catalytic activity and AT-hook domain of Dot1l are important to suppress MERVL. Notably, Dot1l-Npm1 restricts MERVL by regulating protein level and deposition of histone H1. Furthermore, Dot1l is critical for Npm1 to efficiently interact with histone H1 and inhibit ubiquitination of H1 whereas Npm1 is essential for Dot1l to interact with MERVL. Altogether, we discover that Dot1l represses MERVL through chaperoning H1 by collaborating with Npm1. Importantly, our findings shed light on the non-canonical transcriptional repressive role of Dot1l in ESCs.
Topics: Animals; Mice; Embryonic Stem Cells; Endogenous Retroviruses; Histone Methyltransferases; Histones; Methylation; Methyltransferases
PubMed: 37522386
DOI: 10.1093/nar/gkad640 -
Communications Biology Aug 2023Radiotherapy (RT) plus immunotherapy is a promising modality; however, the therapeutic effects are insufficient, and the molecular mechanism requires clarification to...
Radiotherapy (RT) plus immunotherapy is a promising modality; however, the therapeutic effects are insufficient, and the molecular mechanism requires clarification to further develop combination therapies. Here, we found that the RNA virus sensor pathway dominantly regulates the cellular immune response in NSCLC and ESCC cell lines. Notably, transposable elements (TEs), especially long terminal repeats (LTRs), functioned as key ligands for the RNA virus sensor RIG-I, and the mTOR-LTR-RIG-I axis induced the cellular immune response and dendritic cell and macrophage infiltration after irradiation. Moreover, RIG-I-dependent immune activation was observed in ESCC patient tissue. scRNA sequencing and spatial transcriptome analysis revealed that radiotherapy induced the expression of LTRs, and the RNA virus sensor pathway in immune and cancer cells; this pathway was also found to mediate tumour conversion to an immunological hot state. Here, we report the upstream and ligand of the RNA virus sensor pathway functions in irradiated cancer tissues.
Topics: Humans; Cell Line; DEAD Box Protein 58; DNA Transposable Elements; Macrophages
PubMed: 37543704
DOI: 10.1038/s42003-023-05080-x -
Genome Biology Nov 2023Transposable elements (TEs) have colonized the genomes of most metazoans, and many TE-embedded sequences function as cis-regulatory elements (CREs) for genes involved in...
BACKGROUND
Transposable elements (TEs) have colonized the genomes of most metazoans, and many TE-embedded sequences function as cis-regulatory elements (CREs) for genes involved in a wide range of biological processes from early embryogenesis to innate immune responses. Because of their repetitive nature, TEs have the potential to form CRE platforms enabling the coordinated and genome-wide regulation of protein-coding genes by only a handful of trans-acting transcription factors (TFs).
RESULTS
Here, we directly test this hypothesis through mathematical modeling and demonstrate that differences in expression at protein-coding genes alone are sufficient to estimate the magnitude and significance of TE-contributed cis-regulatory activities, even in contexts where TE-derived transcription fails to do so. We leverage hundreds of overexpression experiments and estimate that, overall, gene expression is influenced by TE-embedded CREs situated within approximately 500 kb of promoters. Focusing on the cis-regulatory potential of TEs within the gene regulatory network of human embryonic stem cells, we find that pluripotency-specific and evolutionarily young TE subfamilies can be reactivated by TFs involved in post-implantation embryogenesis. Finally, we show that TE subfamilies can be split into truly regulatorily active versus inactive fractions based on additional information such as matched epigenomic data, observing that TF binding may better predict TE cis-regulatory activity than differences in histone marks.
CONCLUSION
Our results suggest that TE-embedded CREs contribute to gene regulation during and beyond gastrulation. On a methodological level, we provide a statistical tool that infers TE-dependent cis-regulation from RNA-seq data alone, thus facilitating the study of TEs in the next-generation sequencing era.
Topics: Humans; DNA Transposable Elements; Gene Expression Regulation; Gene Regulatory Networks; Promoter Regions, Genetic; Protein Binding
PubMed: 37950299
DOI: 10.1186/s13059-023-03085-7 -
Trends in Genetics : TIG May 2024Helitrons, classified as DNA transposons, employ rolling-circle intermediates for transposition. Distinguishing themselves from other DNA transposons, they leave the... (Review)
Review
Helitrons, classified as DNA transposons, employ rolling-circle intermediates for transposition. Distinguishing themselves from other DNA transposons, they leave the original template element unaltered during transposition, which has led to their characterization as 'peel-and-paste elements'. Helitrons possess the ability to capture and mobilize host genome fragments, with enormous consequences for host genomes. This review discusses the current understanding of Helitrons, exploring their origins, transposition mechanism, and the extensive repercussions of their activity on genome structure and function. We also explore the evolutionary conflicts stemming from Helitron-transposed gene fragments and elucidate their domestication for regulating responses to environmental challenges. Looking ahead, further research in this evolving field promises to bring interesting discoveries on the role of Helitrons in shaping genomic landscapes.
Topics: DNA Transposable Elements; Genome; Animals; Evolution, Molecular; Genomics; Humans
PubMed: 38429198
DOI: 10.1016/j.tig.2024.02.002 -
Cell Reports Jul 2023The type V-K CRISPR-associated transposons (CASTs) allow RNA-guided DNA integration and have great potential as a programmable site-specific gene insertion tool....
The type V-K CRISPR-associated transposons (CASTs) allow RNA-guided DNA integration and have great potential as a programmable site-specific gene insertion tool. Although all core components have been independently characterized structurally, the mechanism of how the transposase TnsB associates with AAA+ ATPase TnsC and catalyzes donor DNA cleavage and integration remains ambiguous. In this study, we demonstrate that TniQ-dCas9 fusion can direct site-specific transposition by TnsB/TnsC in ShCAST. TnsB is a 3'-5' exonuclease that specifically cleaves donor DNA at the end of the terminal repeats and integrates the left end prior to the right end. The nucleotide preference and the cleavage site of TnsB are markedly different from those of the well-documented MuA. We also find that TnsB/TnsC association is enhanced in a half-integration state. Overall, our results provide valuable insights into the mechanism and application expansion of CRISPR-mediated site-specific transposition by TnsB/TnsC.
Topics: Escherichia coli Proteins; Escherichia coli; DNA Transposable Elements; DNA-Binding Proteins; Mutagenesis, Insertional; Transposases
PubMed: 37379212
DOI: 10.1016/j.celrep.2023.112698 -
RNA (New York, N.Y.) Oct 2023RNA-directed transposon silencing operates in the mammalian soma and germline to safeguard genomic integrity. The piRNA pathway and the HUSH complex identify active...
RNA-directed transposon silencing operates in the mammalian soma and germline to safeguard genomic integrity. The piRNA pathway and the HUSH complex identify active transposons through recognition of their nascent transcripts, but mechanistic understanding of how these distinct pathways evolved is lacking. TASOR is an essential component of the HUSH complex. TASOR's DUF3715 domain adopts a pseudo-PARP structure and is required for transposon silencing in a manner independent of complex assembly. TEX15, an essential piRNA pathway factor, also contains the DUF3715 domain. Here, we show that TASOR's and TEX15's DUF3715 domain share extensive structural homology. We found that the DUF3715 domain arose in early eukaryotes and that in vertebrates it is restricted to TEX15, TASOR, and TASORB orthologs. While TASOR-like proteins are found throughout metazoa, TEX15 is vertebrate-specific. The branching of TEX15 and the TASOR-like DUF3715 domain likely occurred in early metazoan evolution. Remarkably, despite this vast evolutionary distance, the DUF3715 domain from divergent TEX15 sequences can functionally substitute the DUF3715 domain of TASOR and mediates transposon silencing. We have thus termed this domain of unknown function as the RNA-directed pseudo-PARP transposon silencing (RDTS) domain. In summary, we show an unexpected functional link between these critical transposon silencing pathways.
Topics: Animals; RNA, Small Interfering; Poly(ADP-ribose) Polymerase Inhibitors; RNA Interference; Genome; Argonaute Proteins; Piwi-Interacting RNA; Mammals; DNA Transposable Elements; Drosophila Proteins; Drosophila melanogaster
PubMed: 37433650
DOI: 10.1261/rna.079693.123 -
International Journal of Molecular... Dec 2023Genetic diversity is a key factor for plant breeding. The birth of novel genic and genomic variants is also crucial for plant adaptation in nature. Therefore, the... (Review)
Review
Genetic diversity is a key factor for plant breeding. The birth of novel genic and genomic variants is also crucial for plant adaptation in nature. Therefore, the genomes of almost all living organisms possess natural mutagenic mechanisms. Transposable elements (TEs) are a major mutagenic force driving genetic diversity in wild plants and modern crops. The relatively rare TE transposition activity during the thousand-year crop domestication process has led to the phenotypic diversity of many cultivated species. The utilization of TE mutagenesis by artificial and transient acceleration of their activity in a controlled mode is an attractive foundation for a novel type of mutagenesis called TE-mediated biological mutagenesis. Here, I focus on TEs as mutagenic sources for plant breeding and discuss existing and emerging transgene-free approaches for TE activation in plants. Furthermore, I also review the non-randomness of TE insertions in a plant genome and the molecular and epigenetic factors involved in shaping TE insertion preferences. Additionally, I discuss the molecular mechanisms that prevent TE transpositions in germline plant cells (e.g., meiocytes, pollen, egg and embryo cells, and shoot apical meristem), thereby reducing the chances of TE insertion inheritance. Knowledge of these mechanisms can expand the TE activation toolbox using novel gene targeting approaches. Finally, the challenges and future perspectives of plant populations with induced novel TE insertions (iTE plant collections) are discussed.
Topics: Plant Breeding; DNA Transposable Elements; Genome, Plant; Crops, Agricultural; Mutagenesis; Evolution, Molecular
PubMed: 38069377
DOI: 10.3390/ijms242317054 -
Andrology Jul 2023The germline perpetuates genetic information across generations. To maintain the integrity of the germline, transposable elements in the genome must be silenced, as... (Review)
Review
BACKGROUND
The germline perpetuates genetic information across generations. To maintain the integrity of the germline, transposable elements in the genome must be silenced, as these mobile elements would otherwise engender widespread mutations passed on to subsequent generations. There are several well-established mechanisms that are dedicated to providing defense against transposable elements, including DNA methylation, RNA interference, and the PIWI-interacting RNA pathway.
OBJECTIVES
Recently, several studies have provided evidence that transposon defense is not only provided by factors dedicated to this purpose but also factors with other roles, including in germline development. Many of these are transcription factors. Our objective is to summarize what is known about these "bi-functional" transcriptional regulators.
MATERIALS AND METHODS
Literature search.
RESULTS AND CONCLUSION
We summarize the evidence that six transcriptional regulators-GLIS3, MYBL1, RB1, RHOX10, SETDB1, and ZBTB16-are both developmental regulators and transposable element-defense factors. These factors act at different stages of germ cell development, including in pro-spermatogonia, spermatogonial stem cells, and spermatocytes. Collectively, the data suggest a model in which specific key transcriptional regulators have acquired multiple functions over evolutionary time to influence developmental decisions and safeguard transgenerational genetic information. It remains to be determined whether their developmental roles were primordial and their transposon defense roles were co-opted, or vice versa.
Topics: Male; Humans; DNA Transposable Elements; Gene Expression Regulation, Developmental; Spermatogonia; Transcription Factors; Spermatocytes; RNA, Small Interfering; Germ Cells
PubMed: 36895139
DOI: 10.1111/andr.13427 -
Science (New York, N.Y.) Nov 2023CRISPR-associated transposases (CASTs) repurpose nuclease-deficient CRISPR effectors to catalyze RNA-guided transposition of large genetic payloads. Type V-K CASTs offer...
CRISPR-associated transposases (CASTs) repurpose nuclease-deficient CRISPR effectors to catalyze RNA-guided transposition of large genetic payloads. Type V-K CASTs offer potential technology advantages but lack accuracy, and the molecular basis for this drawback has remained elusive. Here, we reveal that type V-K CASTs maintain an RNA-independent, "untargeted" transposition pathway alongside RNA-dependent integration, driven by the local availability of TnsC filaments. Using cryo-electron microscopy, single-molecule experiments, and high-throughput sequencing, we found that a minimal, CRISPR-less transpososome preferentially directs untargeted integration at AT-rich sites, with additional local specificity imparted by TnsB. By exploiting this knowledge, we suppressed untargeted transposition and increased type V-K CAST specificity up to 98.1% in cells without compromising on-target integration efficiency. These findings will inform further engineering of CAST systems for accurate, kilobase-scale genome engineering applications.
Topics: CRISPR-Associated Proteins; CRISPR-Cas Systems; Cryoelectron Microscopy; DNA Transposable Elements; Transposases; Cyanobacteria; Single Molecule Imaging; Gene Editing
PubMed: 37972161
DOI: 10.1126/science.adj8543 -
Virulence Dec 2024Enterohemorrhagic (EHEC) is an important zoonotic pathogen that is a major cause of foodborne diseases in most developed and developing countries and can cause... (Review)
Review
Enterohemorrhagic (EHEC) is an important zoonotic pathogen that is a major cause of foodborne diseases in most developed and developing countries and can cause uncomplicated diarrhoea, haemorrhagic colitis, and haemolytic uraemic syndrome. O islands (OIs), which are unique genomic islands in EHEC O157:H7, are composed of 177 isolated genomic features and harbour 26% of the total genes that are absent in the non-pathogenic K-12 genome. In the last twenty years, many OI-encoded proteins have been characterized, including proteins regulating virulence, motility, and acid resistance. Given the critical role of regulatory proteins in the systematic and hierarchical regulation of bacterial biological processes, this review summarizes the OI-encoded regulatory proteins in EHEC O157:H7 characterized to date, emphasizing OI-encoded regulatory proteins for bacterial virulence, motility, and acid resistance. This summary will be significant for further exploration and understanding of the virulence and pathogenesis of EHEC O157:H7.
Topics: Humans; Genomic Islands; Escherichia coli O157; Transcription Factors; Enterohemorrhagic Escherichia coli; Virulence; Escherichia coli Infections; Escherichia coli Proteins
PubMed: 38357901
DOI: 10.1080/21505594.2024.2313407