Project description:Small RNA libraries were constructed from total RNA from Jasminum sambac plants exhibiting virus-like symptoms. After sequencing, small RNAs were assembled into contigs with MetaVelvet and assembled contigs were aligned against the NR database of NCBI using BLASTx. Top hits that reported a virus as subject were considered putative viral sequences. Based on such alignments, the whole genome of a virus, we tentatively name Jasmine Virus H was recovered and cloned. Two more small RNA libraries were made in a confirmatory experiment. One from Jasminum sambac and another one from Nicotiana benthamiana plants infected with the newly-cloned virus. The small RNA libraries were aligned against the full-length sequence of Jasmine Virus H to determine the spacial distribution of virus-derived small RNAs along the virus genome.
Project description:We report the application of high throughput sequencing technology for investigating the transcriptional regulatory network of the human innate immune response. With ChIP-seq, we generated genome-wide virus-activated transcription factor and transcription machinery maps of infected and uninfected human Namalwa B cells. Analysis of ChIP-seq data reveals extensive collaboration of IRF3 and NF-κB with Mediator throughout the human genome, and implicates additional transcription factor partners in antiviral responses. Moreover, analysis of Pol II occupancy and elongation status during virus infection indicates that IRF3 and NF-κB drive both de novo polymerase recruitment and mediate polymerase pause-release at their target sites, stimulating the expression of a variety of protein-coding, non-coding, and unannotated loci. Examination of 6 different proteins before and after virus infection in 1 cell type.
Project description:Long-range RNA-RNA pairing impacts the genome structure and function of SARS-CoV-2 variants. To understand structure and function relationships of different SARS-CoV-2 variants that have emerged during the COVID-19 pandemic, we performed high throughput structure probing and modelling of the genomic structures of the wildtype (WT), Alpha, Beta, Delta and Omicron variants of the SARS-CoV-2. We observed that genomes of SARS-CoV-2 variants are generally structurally conserved, and that single nucleotide variations (SNVs) and interactions with RNA binding proteins (RBPs) can impact RNA structures across the viruses. Importantly, using proximity ligation sequencing, we identified many conserved ultra-long-range RNA-RNA interactions, including one that spans more than 17kb in both the WT virus and Omicron variant. We showed that mutations that disrupt this 17kb long-range interacting structure reduce virus fitness at late stages of its infection cycle, while compensatory mutations partially restore virus fitness. Additionally, we showed that this ultra-long-range RNA-RNA interaction binds directly to ADAR1 to alter the RNA editing levels on the viral genome. These studies deepen our understanding of RNA structures in SARS-CoV-2 genome and their ability to interact with host factors to facilitate virus infectivity.
Project description:Grapevine line pattern virus (GLPV) was described 30 years ago from Hungary, and in the lack of its sequence until now no additional information about its presence was reported. However High-Throughput Sequencing (HTS) applied on dsRNAs extracts recovered from a grapevine plant (accession Baco22A) infected with GLPV Grapevine line pattern virus (GLPV) allowed us to sequence it with different High-Throughput Sequencing (HTS) methods andthe assembleing of the full genome sequence of this virus. The availability of the sequence allowed us to validate the presence of the virus bot with RT-PCR and with Northern blot hybridization. These methods were also used to test its graft and seed transmission. In accordance as it was originally suggested its genome was found to comprise three RNA segments.Its RNA1 (3.160 bp), RNA2 (2.493 bp) and RNA3 (2.529 bp), encode four proteins, denoted 1a (Methyltransferase, helicase), 2a (RNA-dependent RNA Polymerase), 3a (Movement protein, MP) and 3b (Coat protein, CP). GLPV showed the highest amino acid identity (92%–99%) with all domains of Hop yellow virus (HYV), which is a tentative member of the genus Anulavirus of the family Bromoviridae. The phylogenetic trees constructed based on the amino acid sequences of 2a and 3b also confirmed the belongingness of GLPV to the genus Anulavirus, allocating it in one cluster together with the anulaviruses, and close to HYV. The very high sequence identity found between GLPV and HYV leaves no doubt that both are two isolates of the same viral species.