scholarly journals Putative host-derived insertions in the genome of circulating SARS-CoV-2 variants

2022 ◽  
Author(s):  
Yiyan Yang ◽  
Keith Dufault-Thompson ◽  
Rafaela Salgado Fontenele ◽  
Xiaofang Jiang
Keyword(s):  
Direct Evidence ◽  
Viral Evolution ◽  
Genetic Material ◽  
Template Switching ◽  
Sequencing Data ◽  
Rna Dependent Rna Polymerase ◽  
Chimeric Rnas ◽  
Infected Cells ◽  
Host Genetic ◽  
Genomic Insertions

Insertions in the SARS-CoV-2 genome have the potential to drive viral evolution, but the source of the insertions is often unknown. Recent proposals have suggested that human RNAs could be a source of some insertions, but the small size of many insertions makes this difficult to confirm. Through an analysis of available direct RNA sequencing data from SARS-CoV-2 infected cells, we show that viral-host chimeric RNAs are formed through what are likely stochastic RNA-dependent RNA polymerase template switching events. Through an analysis of the publicly available GISAID SARS-CoV-2 genome collection, we then identified two genomic insertions in circulating SARS-CoV-2 variants that are identical to regions of the human 18S and 28S rRNAs. These results provide direct evidence of the formation of viral-host chimeric sequences and the integration of host genetic material into the SARS-CoV-2 genome, highlighting the potential importance of host-derived insertions in viral evolution.

scholarly journals One viral sequence for each host? – The neglected within-host diversity as the main stage of SARS-CoV-2 evolution

2021 ◽  
Author(s):  
Yongsen Ruan ◽  
Mei Hou ◽  
Jiarui Li ◽  
Yangzi Song ◽  
Hurng-YI Wang ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Early Detection ◽  
Viral Evolution ◽  
Infected Individual ◽  
Main Stage ◽  
Viral Sequence ◽  
Host Diversity ◽  
Infected Cells ◽  
Host Genetic ◽  
Total Diversity

The standard practice of presenting one viral sequence for each infected individual implicitly assumes low within-host genetic diversity. It places the emphasis on the viral evolution between, rather than within, hosts. To determine this diversity, we collect SARS-CoV-2 samples from the same patient multiple times. Our own data in conjunction with previous reports show that two viral samples collected from the same individual are often very different due to the substantial within-host diversity. Each sample captures only a small part of the total diversity that is transiently and locally released from infected cells. Hence, the global SARS-CoV-2 population is a meta-population consisting of the viruses in all the infected hosts, each of which harboring a genetically diverse sub-population. Advantageous mutations must be present first as the within-host diversity before they are revealed as between-host polymorphism. The early detection of such diversity in multiple hosts could be an alarm for potentially dangerous mutations. In conclusion, the main forces of viral evolution, i.e., mutation, drift, recombination and selection, all operate within hosts and should be studied accordingly. Several significant implications are discussed.

scholarly journals Identification of the cross-strand chimeric RNAs generated by fusions of bi-directional transcripts

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yuting Wang ◽  
Qin Zou ◽  
Fajin Li ◽  
Wenwei Zhao ◽  
Hui Xu ◽  
...  
Keyword(s):  
Rna Splicing ◽  
Sequencing Data ◽  
Normal Tissues ◽  
Rna Transcripts ◽  
Chimeric Rnas ◽  
Dna Strands ◽  
Comprehensive Survey ◽  
The Cross ◽  
Chimeric Rna ◽  
Cancerous Cells

AbstractA major part of the transcriptome complexity is attributed to multiple types of DNA or RNA fusion events, which take place within a gene such as alternative splicing or between different genes such as DNA rearrangement and trans-splicing. In the present study, using the RNA deep sequencing data, we systematically survey a type of non-canonical fusions between the RNA transcripts from the two opposite DNA strands. We name the products of such fusion events cross-strand chimeric RNA (cscRNA). Hundreds to thousands of cscRNAs can be found in human normal tissues, primary cells, and cancerous cells, and in other species as well. Although cscRNAs exhibit strong tissue-specificity, our analysis identifies thousands of recurrent cscRNAs found in multiple different samples. cscRNAs are mostly originated from convergent transcriptions of the annotated genes and their anti-sense DNA. The machinery of cscRNA biogenesis is unclear, but the cross-strand junction events show some features related to RNA splicing. The present study is a comprehensive survey of the non-canonical cross-strand RNA junction events, a resource for further characterization of the originations and functions of the cscRNAs.

scholarly journals Genetic diversity of Collaborative Cross mice controls viral replication, clinical severity and brain pathology induced by Zika virus infection, independently of Oas1b

2019 ◽  
Author(s):  
Caroline Manet ◽  
Etienne Simon-Lorière ◽  
Grégory Jouvion ◽  
David Hardy ◽  
Matthieu Prot ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Viral Load ◽  
Viral Replication ◽  
Clinical Severity ◽  
Brain Pathology ◽  
Zikv Infection ◽  
Infected Cells ◽  
Host Genetic ◽  
Host Genes

ABSTRACTThe explosive spread of Zika virus (ZIKV) has been associated with major variations in severe disease and congenital afflictions among infected populations, suggesting an influence of host genes. We investigated how genome-wide variants could impact susceptibility to ZIKV infection in mice. We first describe that the susceptibility of Ifnar1 knockout mice is largely influenced by their genetic background. We then show that the broad genetic diversity of Collaborative Cross mice, which receptor to type I interferon (IFNAR) was blocked by anti-IFNAR antibody, expressed phenotypes ranging from complete resistance to severe symptoms and death with large variations in the peak and rate of decrease of plasma viral load, in brain viral load, in brain histopathology and in viral replication rate in infected cells. Differences of susceptibility between CC strains were correlated between Zika, Dengue and West Nile viruses. We identified highly susceptible and resistant mouse strains as new models to investigate the mechanisms of human ZIKV disease and other flavivirus infections. Genetic analyses revealed that phenotypic variations are driven by multiple genes with small effects, reflecting the complexity of ZIKV disease susceptibility in human population. Notably, our results rule out a role of the Oas1b gene in the susceptibility to ZIKV. Altogether, this study emphasizes the role of host genes in the pathogeny of ZIKV infection and lays the foundation for further genetic and mechanistic studies.IMPORTANCEIn recent outbreaks, ZIKV has infected millions of people and induced rare but potentially severe complications, including Guillain-Barré syndrome and encephalitis in adults. While several viral sequence variants were proposed to enhance the pathogenicity of ZIKV, the influence of host genetic variants in the clinical heterogeneity remains mostly unexplored. We have addressed this question using a mouse panel which models the genetic diversity of human population and a ZIKV strain from a recent clinical isolate. Through a combination of in vitro and in vivo approaches, we demonstrate that multiple host genetic variants determine viral replication in infected cells, and clinical severity, kinetics of blood viral load and brain pathology in mice. We describe new mouse models expressing high susceptibility or resistance to ZIKV and to other flaviviruses. These models will facilitate the identification and mechanistic characterization of host genes that influence ZIKV pathogenesis.

scholarly journals Comprehensive analysis of RNA-seq and whole genome sequencing data reveals no evidence for SARS-CoV-2 integrating into host genome

2021 ◽  
Author(s):  
Yu-Sheng Chen ◽  
Shuaiyao Lu ◽  
Bing Zhang ◽  
Tingfu Du ◽  
Wen-Jie Li ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Rna Virus ◽  
Comprehensive Analysis ◽  
Host Genome ◽  
Whole Genome Sequencing Data ◽  
Whole Genome ◽  
Rna Seq ◽  
Sequencing Data ◽  
Infected Cells

SARS-CoV-2, as the causation of severe epidemic of COVID-19, is one kind of positive single-stranded RNA virus with high transmissibility. However, whether or not SARS-CoV-2 can integrate into host genome needs thorough investigation. Here, we performed both RNA sequencing (RNA-seq) and whole genome sequencing on SARS-CoV-2 infected human and monkey cells, and investigated the presence of host-virus chimeric events. Through RNA-seq, we did detect the chimeric host-virus reads in the infected cells. But further analysis using mixed libraries of infected cells and uninfected zebrafish embryos demonstrated that these reads are falsely generated during library construction. In support, whole genome sequencing also didn't identify the existence of chimeric reads in their corresponding regions. Therefore, the evidence for SARS-CoV-2's integration into host genome is lacking.

scholarly journals Introduction of new genetic material into human myeloid leukemic blast stem cells by retroviral infection.

1988 ◽  
Vol 8 (2) ◽  
pp. 974-977 ◽  
Author(s):  
L J Smith ◽  
S Benchimol
Keyword(s):  
Stem Cells ◽  
Clonogenic Assay ◽  
Genetic Material ◽  
Retroviral Vector ◽  
Self Renewal ◽  
Retroviral Infection ◽  
Resistant Phenotype ◽  
Blast Cells ◽  
Infected Cells

An amphotropic retroviral vector containing the bacterial neomycin phosphotransferase gene (neo) was used to infect blast cells from patients with acute myeloblastic leukemia. The infected cells acquired a G418-resistant phenotype that was stable as measured in a clonogenic assay and in long-term suspension culture. Thus, gene transfer into stem cells was accomplished by this procedure. This approach for manipulating gene expression in blast stem cells provides a means to assess the roles of a variety of genes in self-renewal, differentiation, and leukemogenesis.

scholarly journals The structure of a dimeric form of SARS-CoV-2 polymerase

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Florian A. Jochheim ◽  
Dimitry Tegunov ◽  
Hauke S. Hillen ◽  
Jana Schmitzová ◽  
Goran Kokic ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Catalytic Subunit ◽  
Template Switching ◽  
Rna Dependent Rna Polymerase ◽  
Genomic Rnas ◽  
Dimeric Form

AbstractThe coronavirus SARS-CoV-2 uses an RNA-dependent RNA polymerase (RdRp) to replicate and transcribe its genome. Previous structures of the RdRp revealed a monomeric enzyme composed of the catalytic subunit nsp12, two copies of subunit nsp8, and one copy of subunit nsp7. Here we report an alternative, dimeric form of the enzyme and resolve its structure at 5.5 Å resolution. In this structure, the two RdRps contain only one copy of nsp8 each and dimerize via their nsp7 subunits to adopt an antiparallel arrangement. We speculate that the RdRp dimer facilitates template switching during production of sub-genomic RNAs.

scholarly journals Populations of Genomic RNAs Devoted to the Replication or Spread of a Bipartite Plant Virus Differ in Genetic Structure

2009 ◽  
Vol 83 (24) ◽  
pp. 12973-12983 ◽  
Author(s):  
Gloria Lozano ◽  
Ana Grande-Pérez ◽  
Jesús Navas-Castillo
Keyword(s):  
Genetic Structure ◽  
Detailed Analysis ◽  
Rna Viruses ◽  
Control Strategies ◽  
Cooperative Behavior ◽  
Viral Evolution ◽  
Continuous Process ◽  
Viral Quasispecies ◽  
Sequencing Data ◽  
Genomic Rnas

ABSTRACT RNA viruses within a host exist as dynamic distributions of closely related mutants and recombinant genomes. These closely related mutants and recombinant genomes, which are subjected to a continuous process of genetic variation, competition, and selection, act as a unit of selection, termed viral quasispecies. Characterization of mutant spectra within hosts is essential for understanding viral evolution and pathogenesis resulting from the cooperative behavior of viral mutants within viral quasispecies. Furthermore, a detailed analysis of viral variability within hosts is needed to design control strategies, because viral quasispecies are reservoirs of viral variants that potentially can emerge with increased virulence or altered tropism. In this work, we report a detailed analysis of within-host viral populations in 13 field isolates of the bipartite Tomato chlorosis virus (ToCV) (genus Crinivirus, family Closteroviridae). The intraisolate genetic structure was analyzed based on sequencing data for 755 molecular clones distributed in four genomic regions within the RNA-dependent RNA polymerase (RNA1) and Hsp70h, CP, and CPm (RNA2) open reading frames. Our results showed that populations of ToCV within a host plant have a heterogeneous and complex genetic structure similar to that described for animal and plant RNA viral quasispecies. Moreover, the structures of these populations clearly differ depending on the RNA segment considered, being more complex for RNA1 (encoding replication-associated proteins) than for RNA2 (encoding encapsidation-, systemic-movement-, and insect transmission-relevant proteins). These results support the idea that, in multicomponent RNA viruses, function can generate profound differences in the genetic structures of the different genomic segments.

scholarly journals Integration of viral transcriptome sequencing with structure and sequence motifs predicts novel regulatory elements in SARS-CoV-2

2020 ◽  
Author(s):  
Brian J. Cox
Keyword(s):  
Regulatory Elements ◽  
Viral Gene ◽  
Template Switching ◽  
Sequence Motif ◽  
Sequence Motifs ◽  
Human Pathogens ◽  
Sequencing Data ◽  
Stem Loop ◽  
Conserved Sequence ◽  
Splice Junctions

SummaryIn the last twenty years, three separate coronaviruses have left their typical animal hosts and became human pathogens. An area of research interest is coronavirus transcription regulation that uses an RNA-RNA mediated template-switching mechanism. It is not known how different transcriptional stoichiometries of each viral gene are generated. Analysis of SARS-CoV-2 RNA sequencing data from whole RNA transcriptomes identified TRS dependent and independent transcripts. Integration of transcripts and 5’-UTR sequence motifs identified that the pentaloop and the stem-loop 3 were also located upstream of spliced genes. TRS independent transcripts were detected as likely non-polyadenylated. Additionally, a novel conserved sequence motif was discovered at either end of the TRS independent splice junctions. While similar both SARS viruses generated similar TRS independent transcripts they were more abundant in SARS-CoV-2. TRS independent gene regulation requires investigation to determine its relationship to viral pathogenicity.

scholarly journals Securing the exchange of synthetic genetic constructs using digital signatures

2019 ◽  
Author(s):  
Jenna E Gallegos ◽  
Diptendu M. Kar ◽  
Indrakshi Ray ◽  
Indrajit Ray ◽  
Jean Peccoud
Keyword(s):  
Supply Chain ◽  
Digital Signature ◽  
Genetic Material ◽  
Digital Signatures ◽  
Data Encryption ◽  
Reference Sequence ◽  
Sequencing Data ◽  
Encrypted Data ◽  
Genetic Constructs ◽  
Extract Information

AbstractSynthetic biology relies on an ever-growing supply chain of synthetic genetic material. Technologies to secure the exchange of this material are still in their infancy. Solutions proposed thus far have focused on watermarks, a dated security approach that can be used to claim authorship, but is subject to counterfeit, and does not provide any information about the integrity of the genetic material itself. We describe how data encryption and digital signature algorithms can be used to ensure the integrity and authenticity of synthetic genetic constructs. Using a pilot software that generates digital signatures and other encrypted data for plasmids, we demonstrate that we can predictably extract information about the author, the identity, and the integrity of plasmid sequences from sequencing data alone without a reference sequence, all without compromising the function of the plasmids. We discuss how this technology can be improved, applied, and expanded to support the new bioeconomy.

scholarly journals Structural basis of ribosomal frameshifting during translation of the SARS-CoV-2 RNA genome

2020 ◽  
Author(s):  
Pramod R. Bhatt ◽  
Alain Scaiola ◽  
Gary Loughran ◽  
Marc Leibundgut ◽  
Annika Kratzel ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Viral Proteins ◽  
Viral Rna ◽  
Structural Basis ◽  
Rna Dependent Rna Polymerase ◽  
Ribosomal Frameshifting ◽  
Infected Cells ◽  
Rna Genome ◽  
Exit Tunnel ◽  
Viral Polyprotein

AbstractProgrammed ribosomal frameshifting is the key event during translation of the SARS-CoV-2 RNA genome allowing synthesis of the viral RNA-dependent RNA polymerase and downstream viral proteins. Here we present the cryo-EM structure of the mammalian ribosome in the process of translating viral RNA paused in a conformation primed for frameshifting. We observe that the viral RNA adopts a pseudoknot structure lodged at the mRNA entry channel of the ribosome to generate tension in the mRNA that leads to frameshifting. The nascent viral polyprotein that is being synthesized by the ribosome paused at the frameshifting site forms distinct interactions with the ribosomal polypeptide exit tunnel. We use biochemical experiments to validate our structural observations and to reveal mechanistic and regulatory features that influence the frameshifting efficiency. Finally, a compound previously shown to reduce frameshifting is able to inhibit SARS-CoV-2 replication in infected cells, establishing coronavirus frameshifting as target for antiviral intervention.

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