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Viruses Nov 2021The delivery of the HIV-1 genome into the nucleus is an indispensable step in retroviral infection of non-dividing cells, but the mechanism of HIV-1 nuclear import has... (Review)
Review
The delivery of the HIV-1 genome into the nucleus is an indispensable step in retroviral infection of non-dividing cells, but the mechanism of HIV-1 nuclear import has been a longstanding debate due to controversial experimental evidence. It was commonly believed that the HIV-1 capsid would need to disassemble (uncoat) in the cytosol before nuclear import because the capsid is larger than the central channel of nuclear pore complexes (NPCs); however, increasing evidence demonstrates that intact, or nearly intact, HIV-1 capsid passes through the NPC to enter the nucleus. With the protection of the capsid, the HIV-1 core completes reverse transcription in the nucleus and is translocated to the integration site. Uncoating occurs while, or after, the viral genome is released near the integration site. These independent discoveries reveal a compelling new paradigm of this important step of the HIV-1 life cycle. In this review, we summarize the recent studies related to HIV-1 nuclear import, highlighting the spatial-temporal relationship between the nuclear entry of the virus core, reverse transcription, and capsid uncoating.
Topics: Active Transport, Cell Nucleus; Capsid; Cell Nucleus; HIV Infections; HIV-1; Humans; Nuclear Pore; Reverse Transcription; Virus Integration; Virus Uncoating
PubMed: 34835048
DOI: 10.3390/v13112242 -
Human Gene Therapy Nov 2023Integration of naturally occurring adeno-associated viruses (AAV; wild-type AAV [wtAAV]) and those used in gene therapy (recombinant AAV [rAAV]) into host genomic DNA...
Integration of naturally occurring adeno-associated viruses (AAV; wild-type AAV [wtAAV]) and those used in gene therapy (recombinant AAV [rAAV]) into host genomic DNA has been documented for over two decades. Results from mouse and dog studies have raised concerns of insertional mutagenesis and clonal expansion following AAV exposure, particularly in the context of gene therapy. This study aimed to characterize the genomic location, abundance, and expansion of wtAAV and rAAV integrations in macaque and human genomes. Using an unbiased, next-generation sequencing-based approach, we identified the genome-wide integration loci in tissue samples (primarily liver) in 168 nonhuman primates (NHPs) and 85 humans naïve to rAAV exposure and 86 NHPs treated with rAAV in preclinical studies. Our results suggest that rAAV and wtAAV integrations exhibit similar, broad distribution patterns across species, with a higher frequency in genomic regions highly vulnerable to DNA damage or close to highly transcribed genes. rAAV exhibited a higher abundance of unique integration loci, whereas wtAAV integration loci were associated with greater clonal expansion. This expansive and detailed characterization of AAV integration in NHPs and humans provides key translational insights, with important implications for the safety of rAAV as a gene therapy vector.
Topics: Animals; Humans; Dependovirus; Genetic Therapy; Genetic Vectors; Liver; Macaca; Virus Integration
PubMed: 37930949
DOI: 10.1089/hum.2023.134 -
Viruses Jan 2023Bacteriophages are ubiquitous organisms that can be specific to one or multiple strains of hosts, in addition to being the most abundant entities on the planet. It is... (Review)
Review
Bacteriophages are ubiquitous organisms that can be specific to one or multiple strains of hosts, in addition to being the most abundant entities on the planet. It is estimated that they exceed ten times the total number of bacteria. They are classified as temperate, which means that phages can integrate their genome into the host genome, originating a prophage that replicates with the host cell and may confer immunity against infection by the same type of phage; and lytics, those with greater biotechnological interest and are viruses that lyse the host cell at the end of its reproductive cycle. When lysogenic, they are capable of disseminating bacterial antibiotic resistance genes through horizontal gene transfer. When professionally lytic-that is, obligately lytic and not recently descended from a temperate ancestor-they become allies in bacterial control in ecological imbalance scenarios; these viruses have a biofilm-reducing capacity. Phage therapy has also been advocated by the scientific community, given the uniqueness of issues related to the control of microorganisms and biofilm production when compared to other commonly used techniques. The advantages of using bacteriophages appear as a viable and promising alternative. This review will provide updates on the landscape of phage applications for the biocontrol of pathogens in industrial settings and healthcare.
Topics: Bacteriophages; Prophages; Lysogeny; Biofilms; Biotechnology
PubMed: 36851563
DOI: 10.3390/v15020349 -
Molecular Therapy : the Journal of the... Oct 2021Recombinant adeno-associated viral (rAAV) vectors are considered promising tools for gene therapy directed at the liver. Whereas rAAV is thought to be an episomal...
Recombinant adeno-associated viral (rAAV) vectors are considered promising tools for gene therapy directed at the liver. Whereas rAAV is thought to be an episomal vector, its single-stranded DNA genome is prone to intra- and inter-molecular recombination leading to rearrangements and integration into the host cell genome. Here, we ascertained the integration frequency of rAAV in human hepatocytes transduced either ex vivo or in vivo and subsequently expanded in a mouse model of xenogeneic liver regeneration. Chromosomal rAAV integration events and vector integrity were determined using the capture-PacBio sequencing approach, a long-read next-generation sequencing method that has not previously been used for this purpose. Chromosomal integrations were found at a surprisingly high frequency of 1%-3% both in vitro and in vivo. Importantly, most of the inserted rAAV sequences were heavily rearranged and were accompanied by deletions of the host genomic sequence at the integration site.
Topics: Animals; Cells, Cultured; Chromosomes; Dependovirus; Disease Models, Animal; Genetic Therapy; Genetic Vectors; Hepatocytes; Humans; Liver Regeneration; Mice; Transduction, Genetic; Virus Integration
PubMed: 34461297
DOI: 10.1016/j.ymthe.2021.08.031 -
Multiomics analyses reveal pathological mechanisms of HBV infection and integration in liver cancer.Journal of Medical Virology Aug 2023Hepatitis B virus (HBV) infection and integration are important for hepatocellular carcinoma (HCC) initiation and progression, while disease mechanisms are still largely...
Hepatitis B virus (HBV) infection and integration are important for hepatocellular carcinoma (HCC) initiation and progression, while disease mechanisms are still largely elusive. Here, we combined bulk and single-cell sequencing technologies to tackle the disease mechanisms of HBV-related HCC. We observed high HBV mutation rate and diversity only in tumors without HBV integration. We identified human somatic risk loci for HBV integration (VIMs). Transcription factors (TFs) enriched in VIMs were involved in DNA repair and androgen receptor (AR) signaling. Aberration of AR signaling was further observed by single-cell regulon analysis in HBV-infected hepatocytes, which showed remarkable interactions between AR and the complement system that, together with the X-linked ZXDB regulon that contains albumin (ALB), probably contribute to HCC male predominance. Complement system dysregulation caused by HBV infection was further confirmed by analyses of single-cell copy numbers and cell-cell communications. Finally, HBV infection-associated immune cells presented critical defects, including TXNIP in T cells, TYROBP in NK cells, and the X-linked TIMP1 in monocytes. We further experimentally validated our findings in multiple independent patient cohorts. Collectively, our work shed light on the pathogenesis of HBV-related HCC and other liver diseases that affect billions of people worldwide.
Topics: Male; Humans; Female; Hepatitis B virus; Liver Neoplasms; Carcinoma, Hepatocellular; Multiomics; Hepatitis B; Hepatitis B, Chronic; Virus Integration
PubMed: 37522289
DOI: 10.1002/jmv.28980 -
Advanced Science (Weinheim,... May 2021Approximately 15% of human cancers are estimated to be attributed to viruses. Virus sequences can be integrated into the host genome, leading to genomic instability and...
Approximately 15% of human cancers are estimated to be attributed to viruses. Virus sequences can be integrated into the host genome, leading to genomic instability and carcinogenesis. Here, a new deep convolutional neural network (CNN) model is developed with attention architecture, namely DeepVISP, for accurately predicting oncogenic virus integration sites (VISs) in the human genome. Using the curated benchmark integration data of three viruses, hepatitis B virus (HBV), human herpesvirus (HPV), and Epstein-Barr virus (EBV), DeepVISP achieves high accuracy and robust performance for all three viruses through automatically learning informative features and essential genomic positions only from the DNA sequences. In comparison, DeepVISP outperforms conventional machine learning methods by 8.43-34.33% measured by area under curve (AUC) value enhancement in three viruses. Moreover, DeepVISP can decode -regulatory factors that are potentially involved in virus integration and tumorigenesis, such as HOXB7, IKZF1, and LHX6. These findings are supported by multiple lines of evidence in literature. The clustering analysis of the informative motifs reveales that the representative k-mers in clusters could help guide virus recognition of the host genes. A user-friendly web server is developed for predicting putative oncogenic VISs in the human genome using DeepVISP.
Topics: Cluster Analysis; Deep Learning; Genome, Human; Genomic Instability; Hepatitis B virus; Herpesvirus 1, Human; Herpesvirus 4, Human; Humans; Neoplasms; Oncogenic Viruses; Reproducibility of Results; Virus Integration
PubMed: 33977077
DOI: 10.1002/advs.202004958 -
MSystems Apr 2022Temperate phages (prophages) are ubiquitous in nature and persist as dormant components of host cells (lysogenic stage) before activating and lysing the host (lytic...
Temperate phages (prophages) are ubiquitous in nature and persist as dormant components of host cells (lysogenic stage) before activating and lysing the host (lytic stage). Actively replicating prophages contribute to central community processes, such as enabling bacterial virulence, manipulating biogeochemical cycling, and driving microbial community diversification. Recent advances in sequencing technology have allowed for the identification and characterization of diverse phages, yet no approaches currently exist for identifying if a prophage has activated. Here, we present PropagAtE (Prophage Activity Estimator), an automated software tool for estimating if a prophage is in the lytic or lysogenic stage of infection. PropagAtE uses statistical analyses of prophage-to-host read coverage ratios to decipher actively replicating prophages, irrespective of whether prophages were induced or spontaneously activated. We demonstrate that PropagAtE is fast, accurate, and sensitive, regardless of sequencing depth. Application of PropagAtE to prophages from 348 complex metagenomes from human gut, murine gut, and soil environments identified distinct spatial and temporal prophage activation signatures, with the highest proportion of active prophages in murine gut samples. In infants treated with antibiotics or infants without treatment, we identified active prophage populations correlated with specific treatment groups. Within time series samples from the human gut, 11 prophage populations, some encoding the sulfur metabolism gene or a -like virulence factor, were consistently present over time but not active. Overall, PropagAtE will facilitate accurate representations of viruses in microbiomes by associating prophages with their active roles in shaping microbial communities in nature. Viruses that infect bacteria are key components of microbiomes and ecosystems. They can kill and manipulate microorganisms, drive planetary-scale processes and biogeochemical cycling, and influence the structures of entire food networks. Prophages are viruses that can exist in a dormant state within the genome of their host (lysogenic stage) before activating in order to replicate and kill the host (lytic stage). Recent advances have allowed for the identification of diverse viruses in nature, but no approaches exist for characterizing prophages and their stages of infection (prophage activity). We develop and benchmark an automated approach, PropagAtE, to identify the stages of infection of prophages from genomic data. We provide evidence that active prophages vary in identity and abundance across multiple environments and scales. Our approach will enable accurate and unbiased analyses of viruses in microbiomes and ecosystems.
Topics: Humans; Animals; Mice; Prophages; Metagenome; Lysogeny; Bacteriophages; Microbiota
PubMed: 35323045
DOI: 10.1128/msystems.00084-22 -
Communications Biology Jul 2023Hepatitis B virus (HBV) may integrate into the genome of infected cells and contribute to hepatocarcinogenesis. However, the role of HBV integration in hepatocellular...
Hepatitis B virus (HBV) may integrate into the genome of infected cells and contribute to hepatocarcinogenesis. However, the role of HBV integration in hepatocellular carcinoma (HCC) development remains unclear. In this study, we apply a high-throughput HBV integration sequencing approach that allows sensitive identification of HBV integration sites and enumeration of integration clones. We identify 3339 HBV integration sites in paired tumour and non-tumour tissue samples from 7 patients with HCC. We detect 2107 clonally expanded integrations (1817 in tumour and 290 in non-tumour tissues), and a significant enrichment of clonal HBV integrations in mitochondrial DNA (mtDNA) preferentially occurring in the oxidative phosphorylation genes (OXPHOS) and D-loop region. We also find that HBV RNA sequences are imported into the mitochondria of hepatoma cells with the involvement of polynucleotide phosphorylase (PNPASE), and that HBV RNA might have a role in the process of HBV integration into mtDNA. Our results suggest a potential mechanism by which HBV integration may contribute to HCC development.
Topics: Humans; Hepatitis B virus; Carcinoma, Hepatocellular; Liver Neoplasms; DNA, Mitochondrial; Virus Integration; Mitochondria
PubMed: 37400627
DOI: 10.1038/s42003-023-05017-4 -
Molecular Therapy : the Journal of the... Dec 2021
Topics: Dependovirus; Genetic Vectors; HeLa Cells; Humans; Virus Integration
PubMed: 34758291
DOI: 10.1016/j.ymthe.2021.10.024 -
Viruses Jan 2021Human hepatitis B virus (HBV) can cause chronic, lifelong infection of the liver that may lead to persistent or episodic immune-mediated inflammation against... (Review)
Review
Human hepatitis B virus (HBV) can cause chronic, lifelong infection of the liver that may lead to persistent or episodic immune-mediated inflammation against virus-infected hepatocytes. This immune response results in elevated rates of killing of virus-infected hepatocytes, which may extend over many years or decades, lead to fibrosis and cirrhosis, and play a role in the high incidence of hepatocellular carcinoma (HCC) in HBV carriers. Immune-mediated inflammation appears to cause oxidative DNA damage to hepatocytes, which may also play a major role in hepatocarcinogenesis. An additional DNA damaging feature of chronic infections is random integration of HBV DNA into the chromosomal DNA of hepatocytes. While HBV DNA integration does not have a role in virus replication it may alter gene expression of the host cell. Indeed, most HCCs that arise in HBV carriers contain integrated HBV DNA and, in many, the integrant appears to have played a role in hepatocarcinogenesis. Clonal expansion of hepatocytes, which is a natural feature of liver biology, occurs because the hepatocyte population is self-renewing and therefore loses complexity due to random hepatocyte death and replacement by proliferation of surviving hepatocytes. This process may also represent a risk factor for the development of HCC. Interestingly, during chronic HBV infection, hepatocyte clones detected using integrated HBV DNA as lineage-specific markers, emerge that are larger than those expected to occur by random death and proliferation of hepatocytes. The emergence of these larger hepatocyte clones may reflect a survival advantage that could be explained by an ability to avoid the host immune response. While most of these larger hepatocyte clones are probably not preneoplastic, some may have already acquired preneoplastic changes. Thus, chronic inflammation in the HBV-infected liver may be responsible, at least in part, for both initiation of HCC via oxidative DNA damage and promotion of HCC via stimulation of hepatocyte proliferation through immune-mediated killing and compensatory division.
Topics: Animals; DNA, Viral; Hepatitis B virus; Hepatitis B, Chronic; Hepatocytes; Humans; Liver; Virus Integration
PubMed: 33573130
DOI: 10.3390/v13020210