Insertional Mutagenesis Using Tnt1 Retrotransposon in Potato

Cancer arises from the accumulation of genetic and epigenetic alterations. Even in the era of precision oncology, carcinogens contributing to neoplastic process are still an important focus of research. Comprehensive genomic analyses have revealed various combinations of base substitutions, referred to as the mutational signatures, in cancer. Each mutational signature is believed to arise from specific DNA damage and repair processes, including carcinogens. However, as a type of carcinogen, tumor viruses increase the cancer risk by alternative mechanisms, including insertional mutagenesis, viral oncogenes, and immunosuppression. In this review, we summarize virus-driven carcinogenesis to provide a framework for the control of malignant cell proliferation. We first provide a brief overview of oncogenic viruses and describe their implication in virus-related tumors. Next, we describe tumor viruses (HPV, Human papilloma virus; HBV, Hepatitis B virus; HCV, Hepatitis C virus; EBV, Epstein–Barr virus; Kaposi sarcoma herpesvirus; MCV, Merkel cell polyoma virus; HTLV-1, Human T-cell lymphotropic virus, type-1) and tumor virus-related cancers. Lastly, we introduce emerging tumor virus candidates, human cytomegalovirus (CMV), human herpesvirus-6 (HHV-6) and adeno-associated virus-2 (AAV-2). We expect this review to be a hub in a complex network of data for virus-associated carcinogenesis.

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Fgf10 is an oncogene activated by MMTV insertional mutagenesis in mouse mammary tumors and overexpressed in a subset of human breast carcinomas

Oncogene ◽

10.1038/sj.onc.1207816 ◽

2004 ◽

Vol 23 (36) ◽

pp. 6047-6055 ◽

Cited By ~ 43

Author(s):

Vassiliki Theodorou ◽

Mandy Boer ◽

Britta Weigelt ◽

Jos Jonkers ◽

Martin van der Valk ◽

...

Keyword(s):

Insertional Mutagenesis ◽

Mammary Tumors ◽

Breast Carcinomas ◽

Human Breast ◽

Human Breast Carcinomas ◽

Mouse Mammary Tumors

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Genome-wide integration site detection using Cas9 enriched amplification-free long-range sequencing

Nucleic Acids Research ◽

10.1093/nar/gkaa1152 ◽

2020 ◽

Author(s):

Joost van Haasteren ◽

Altar M Munis ◽

Deborah R Gill ◽

Stephen C Hyde

Keyword(s):

Long Range ◽

Insertional Mutagenesis ◽

Lentiviral Vector ◽

Integration Site ◽

Low Cost ◽

Safety Evaluation ◽

Read Length ◽

Chromosomal Dna ◽

Wide Range

Abstract The gene and cell therapy fields are advancing rapidly, with a potential to treat and cure a wide range of diseases, and lentivirus-based gene transfer agents are the vector of choice for many investigators. Early cases of insertional mutagenesis caused by gammaretroviral vectors highlighted that integration site (IS) analysis was a major safety and quality control checkpoint for lentiviral applications. The methods established to detect lentiviral integrations using next-generation sequencing (NGS) are limited by short read length, inadvertent PCR bias, low yield, or lengthy protocols. Here, we describe a new method to sequence IS using Amplification-free Integration Site sequencing (AFIS-Seq). AFIS-Seq is based on amplification-free, Cas9-mediated enrichment of high-molecular-weight chromosomal DNA suitable for long-range Nanopore MinION sequencing. This accessible and low-cost approach generates long reads enabling IS mapping with high certainty within a single day. We demonstrate proof-of-concept by mapping IS of lentiviral vectors in a variety of cell models and report up to 1600-fold enrichment of the signal. This method can be further extended to sequencing of Cas9-mediated integration of genes and to in vivo analysis of IS. AFIS-Seq uses long-read sequencing to facilitate safety evaluation of preclinical lentiviral vector gene therapies by providing IS analysis with improved confidence.

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Methylation of rRNA as a host defense against rampant group II intron retrotransposition

Mobile DNA ◽

10.1186/s13100-021-00237-z ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Justin M. Waldern ◽

Dorie Smith ◽

Carol Lyn Piazza ◽

E. Jake Bailey ◽

Nicholas J. Schiraldi ◽

...

Keyword(s):

Insertional Mutagenesis ◽

Fold Increase ◽

Group Ii Intron ◽

In Vivo Experiments ◽

Cellular Factors ◽

Group Ii ◽

Self Splicing ◽

Native Host

Abstract Background Group II introns are mobile retroelements, capable of invading new sites in DNA. They are self-splicing ribozymes that complex with an intron-encoded protein to form a ribonucleoprotein that targets DNA after splicing. These molecules can invade DNA site-specifically, through a process known as retrohoming, or can invade ectopic sites through retrotransposition. Retrotransposition, in particular, can be strongly influenced by both environmental and cellular factors. Results To investigate host factors that influence retrotransposition, we performed random insertional mutagenesis using the ISS1 transposon to generate a library of over 1000 mutants in Lactococcus lactis, the native host of the Ll.LtrB group II intron. By screening this library, we identified 92 mutants with increased retrotransposition frequencies (RTP-ups). We found that mutations in amino acid transport and metabolism tended to have increased retrotransposition frequencies. We further explored a subset of these RTP-up mutants, the most striking of which is a mutant in the ribosomal RNA methyltransferase rlmH, which exhibited a reproducible 20-fold increase in retrotransposition frequency. In vitro and in vivo experiments revealed that ribosomes in the rlmH mutant were defective in the m3Ψ modification and exhibited reduced binding to the intron RNA. Conclusions Taken together, our results reinforce the importance of the native host organism in regulating group II intron retrotransposition. In particular, the evidence from the rlmH mutant suggests a role for ribosome modification in limiting rampant retrotransposition.

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