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Nature Genetics Feb 2002Adeno-associated virus (AAV) vectors are currently being used in several clinical gene-therapy trials (see the NIH OBA Human Gene Transfer Clinical Trials Database);...
Adeno-associated virus (AAV) vectors are currently being used in several clinical gene-therapy trials (see the NIH OBA Human Gene Transfer Clinical Trials Database); however, little is known about the chromosomal effects of vector integration. Here we report that integrated vector proviruses are associated with chromosomal deletions and other rearrangements and are frequently located on chromosome 19 (although not at the wildtype AAV integration site).
Topics: Base Sequence; Chromosomes, Human, Pair 19; DNA; Dependovirus; Gene Transfer Techniques; Genetic Vectors; HeLa Cells; Humans; Molecular Sequence Data; Virus Integration
PubMed: 11799395
DOI: 10.1038/ng824 -
Retrovirology Jul 2016The NCI Retrovirus Integration Database is a MySql-based relational database created for storing and retrieving comprehensive information about retroviral integration...
UNLABELLED
The NCI Retrovirus Integration Database is a MySql-based relational database created for storing and retrieving comprehensive information about retroviral integration sites, primarily, but not exclusively, HIV-1. The database is accessible to the public for submission or extraction of data originating from experiments aimed at collecting information related to retroviral integration sites including: the site of integration into the host genome, the virus family and subtype, the origin of the sample, gene exons/introns associated with integration, and proviral orientation. Information about the references from which the data were collected is also stored in the database. Tools are built into the website that can be used to map the integration sites to UCSC genome browser, to plot the integration site patterns on a chromosome, and to display provirus LTRs in their inserted genome sequence. The website is robust, user friendly, and allows users to query the database and analyze the data dynamically.
AVAILABILITY
https://rid.ncifcrf.gov ; or http://home.ncifcrf.gov/hivdrp/resources.htm .
Topics: Databases, Genetic; Genome, Human; HIV-1; Host-Pathogen Interactions; Humans; Internet; Proviruses; Retroviridae; Virus Integration
PubMed: 27377064
DOI: 10.1186/s12977-016-0277-6 -
Human Gene Therapy Jun 2008Retroviral DNA integration into the host cell genome is an essential feature of the retroviral life cycle. The ability to integrate their DNA into the DNA of infected... (Review)
Review
Retroviral DNA integration into the host cell genome is an essential feature of the retroviral life cycle. The ability to integrate their DNA into the DNA of infected cells also makes retroviruses attractive vectors for delivery of therapeutic genes into the genome of cells carrying adverse mutations in their cellular DNA. Sequencing of the entire human genome has enabled identification of integration site preferences of both replication-competent retroviruses and retroviral vectors. These results, together with the unfortunate outcome of a gene therapy trial, in which integration of a retroviral vector in the vicinity of a protooncogene was associated with the development of leukemia, have stimulated efforts to elucidate the molecular mechanism underlying integration site selection by retroviral vectors, as well as the development of methods to direct integration to specific DNA sequences and chromosomal regions. This review outlines our current knowledge of the mechanism of integration site selection by retroviruses in vitro, in cultured cells, and in vivo; the outcome of several of the more recent gene therapy trials, which employed these vectors; and the efforts of several laboratories to develop vectors that integrate at predetermined sites in the human genome.
Topics: Base Sequence; Genetic Therapy; Genetic Vectors; Genome, Human; Humans; Retroviridae; Virus Integration
PubMed: 18533894
DOI: 10.1089/hum.2007.148 -
Current Opinion in Structural Biology Dec 2017Retroviral DNA integration takes place in the context of the intasome nucleoprotein complex. X-ray crystal structures of functional spumaviral intasomes were previously... (Review)
Review
Retroviral DNA integration takes place in the context of the intasome nucleoprotein complex. X-ray crystal structures of functional spumaviral intasomes were previously revealed to harbor a homotetramer of integrase, and it was generally believed that integrase tetramers catalyzed the integration of other retroviruses. The elucidation of new structures from four different retroviruses over the past year has however revealed this is not the case. The number of integrase molecules required to construct the conserved intasome core structure differs between viral species. While four subunits suffice for spumaviruses, α- and β-retroviruses require eight and the lentiviruses use up to sixteen. Herein we described these alternative architectures, highlighting both evolutionary and structural constraints that result in the different integrase-DNA stoichiometries across Retroviridae.
Topics: DNA, Viral; Integrases; Models, Molecular; Multiprotein Complexes; Nucleic Acid Conformation; Protein Binding; Protein Conformation; Retroviridae; Structure-Activity Relationship; Virus Integration
PubMed: 28458055
DOI: 10.1016/j.sbi.2017.04.005 -
Journal of Virology Jul 2022Integration of the reverse-transcribed genome is a critical step of the retroviral life cycle. Strand-transfer inhibitors (INSTIs) used for antiretroviral therapy...
Integration of the reverse-transcribed genome is a critical step of the retroviral life cycle. Strand-transfer inhibitors (INSTIs) used for antiretroviral therapy inhibit integration but can lead to resistance mutations in the integrase gene, the enzyme involved in this reaction. A significant proportion of INSTI treatment failures, particularly those with second-generation INSTIs, show no mutation in the integrase gene. Here, we show that replication of a selected dolutegravir-resistant virus with mutations in the 3'-PPT (polypurine tract) was effective, although no integrated viral DNA was detected, due to the accumulation of unintegrated viral DNA present as 1-LTR circles. Our results show that mutation in the 3'-PPT leads to 1-LTR circles and not linear DNA as classically reported. In conclusion, our data provide a molecular basis to explain a new mechanism of resistance to INSTIs, without mutation of the integrase gene and highlights the importance of unintegrated viral DNA in HIV-1 replication. Our work highlights the role of HIV-1 unintegrated viral DNA in viral replication. A virus, resistant to strand-transfer inhibitors, has been selected . This virus highlights a mutation in the 3'PPT region and not in the integrase gene. This mutation modifies the reverse transcription step leading to the accumulation of 1-LTR circles and not the linear DNA. This accumulation of 1-LTR circles leads to viral replication without integration of the viral genome.
Topics: DNA, Viral; HIV Infections; HIV-1; Humans; Mutation; Virus Integration; Virus Replication
PubMed: 35758669
DOI: 10.1128/jvi.00676-22 -
Clinical Microbiology and Infection :... Apr 2016
Topics: Gene Rearrangement; Host-Pathogen Interactions; Humans; Virus Integration; Virus Physiological Phenomena
PubMed: 26850825
DOI: 10.1016/j.cmi.2016.01.022 -
Journal of Medical Virology Apr 2024The clearance or transcriptional silencing of integrated HBV DNA is crucial for achieving a functional cure in patients with chronic hepatitis B and reducing the risk...
The clearance or transcriptional silencing of integrated HBV DNA is crucial for achieving a functional cure in patients with chronic hepatitis B and reducing the risk of hepatocellular carcinoma development. The PLC/PRF/5 cell line is commonly used as an in vitro model for studying HBV integration. In this study, we employed a range of multi-omics techniques to gain a panoramic understanding of the characteristics of HBV integration in PLC/PRF/5 cells and to reveal the transcriptional regulatory mechanisms of integrated HBV DNA. Transcriptome long-read sequencing (ONT) was conducted to analyze and characterize the transcriptional activity of different HBV DNA integration sites in PLC/PRF/5 cells. Additionally, we collected data related to epigenetic regulation, including whole-genome bisulfite sequencing (WGBS), histone chromatin immunoprecipitation sequencing (ChIP-seq), and assays for transposase-accessible chromatin using sequencing (ATAC-seq), to explore the potential mechanisms involved in the transcriptional regulation of integrated HBV DNA. Long-read RNA sequencing analysis revealed significant transcriptional differences at various integration sites in the PLC/PRF/5 cell line, with higher HBV DNA transcription levels at integration sites on chr11, chr13, and the chr13/chr5 fusion chromosome t (13:5). Combining long-read DNA and RNA sequencing results, we found that transcription of integrated HBV DNA generally starts downstream of the SP1, SP2, or XP promoters. ATAC-seq data confirmed that chromatin accessibility has limited influence on the transcription of integrated HBV DNA in the PLC/PRF/5 cell line. Analysis of WGBS data showed that the methylation intensity of integrated HBV DNA was highly negatively correlated with its transcription level (r = -0.8929, p = 0.0123). After AzaD treatment, the transcription level of integrated HBV DNA significantly increased, especially for the integration chr17, which had the highest level of methylation. Through ChIP-seq data, we observed the association between histone modification of H3K4me3 and H3K9me3 with the transcription of integrated HBV DNA. Our findings suggest that the SP1, SP2 and XP in integrated HBV DNA, methylation level of surrounding host chromosome, and histone modifications affect the transcription of integrated HBV DNA in PLC/PRF/5 cells. This provides important clues for future studies on the expression and regulatory mechanisms of integrated HBV.
Topics: Humans; Hepatitis B virus; Epigenesis, Genetic; Virus Integration; DNA, Viral; Transcription, Genetic; Cell Line; DNA Methylation; Cell Line, Tumor; Histones; Multiomics
PubMed: 38647071
DOI: 10.1002/jmv.29614 -
Journal of Neuroimmune Pharmacology :... Sep 2010Over the past decade, antiretroviral therapy targeting the viral entry process, reverse transcriptase, integrase, and protease, has prolonged the lives of people... (Review)
Review
Over the past decade, antiretroviral therapy targeting the viral entry process, reverse transcriptase, integrase, and protease, has prolonged the lives of people infected with human immunodeficiency virus type 1 (HIV-1). However, despite the development of more effective therapeutic strategies, reservoirs of viral infection remain. This review discusses molecular mechanisms surrounding the development of latency from the site of integration to pre- and post-integration maintenance of latency, including epigenetic factors. In addition, an overview of innate and adaptive cells important to HIV-1 infection are examined from the viewpoint of cytokines released and cytokines that act on these cells to explore an overall understanding of HIV-1 proviral genome activation. Finally, this review is discussed from the viewpoint of how an understanding of the interplay of all of these factors will help guide the next generation of therapies.
Topics: Anti-HIV Agents; CD4-Positive T-Lymphocytes; Cytokines; Epigenesis, Genetic; HIV Infections; HIV Long Terminal Repeat; HIV-1; Humans; Immunity, Innate; Virus Activation; Virus Integration; Virus Latency
PubMed: 20387125
DOI: 10.1007/s11481-010-9207-x -
Annals of the New York Academy of... Apr 2015Single-stranded DNA (ssDNA) viruses are widespread in the environment and include economically, medically, and ecologically important pathogens. Recently, it has been... (Review)
Review
Single-stranded DNA (ssDNA) viruses are widespread in the environment and include economically, medically, and ecologically important pathogens. Recently, it has been discovered that ssDNA virus genomes are also prevalent in the chromosomes of their bacterial, archaeal, and eukaryotic hosts. Sequences originating from viruses of the families Parvoviridae, Circoviridae, and Geminiviridae are particularly widespread in the genomes of eukaryotes, where they are often fossilized as endogenous viral elements. ssDNA viruses have evolved diverse mechanisms to invade cellular genomes, and these principally vary between viruses infecting bacteria/archaea and eukaryotes. Filamentous bacteriophages (Inoviridae) use at least three major mechanisms of integration. Some of these phages encode integrases of serine or tyrosine recombinase superfamilies, while others utilize DDE transposases of the IS3, IS30, or IS110/IS492 families, whereas some inoviruses, and possibly certain members of the Microviridae, hijack the host XerCD recombination machinery. By contrast, eukaryotic viruses for integration rely on the endonuclease activity of their rolling-circle replication-initiation proteins, mimicking the mechanisms used by some bacterial transposons. Certain bacterial and eukaryotic ssDNA viruses have embraced a transposon-like means of propagation, with occasionally dramatic effects on host genome evolution. Here, we review the diversity of experimentally verified and hypothetical mechanisms of genome integration employed by ssDNA viruses, and consider the evolutionary implications of these processes, particularly in the emergence of novel virus groups.
Topics: Archaea; Bacteria; DNA Viruses; DNA, Single-Stranded; Eukaryotic Cells; Evolution, Molecular; Genome, Viral; Host-Pathogen Interactions; Models, Genetic; Virus Integration
PubMed: 25675979
DOI: 10.1111/nyas.12675 -
Journal of Virology Aug 2006Preferential integration into transcriptionally active regions of genomes has been observed for retroviral vectors based on gamma-retroviruses and lentiviruses. However,...
Preferential integration into transcriptionally active regions of genomes has been observed for retroviral vectors based on gamma-retroviruses and lentiviruses. However, differences in the integration site preferences were detected, which might be explained by differences in viral components of the preintegration complexes. Viral determinants of integration site preferences have not been defined. Therefore, integration sites of simian immunodeficiency virus (SIV)-based vectors produced in the absence of accessory genes or lacking promoter and enhancer elements were compared. Similar integration patterns for the different SIV vectors indicate that vif, vpr, vpx, nef, env, and promoter or enhancer elements are not required for preferential integration of SIV into transcriptionally active regions of genomes.
Topics: Cells, Cultured; Chromosome Mapping; Genetic Vectors; Genome, Human; Humans; Simian Immunodeficiency Virus; Virus Integration
PubMed: 16873270
DOI: 10.1128/JVI.00373-06