Project description:While employing deep sequencing and de novo assembly to characterize the mRNA transcript profile of a cell line derived from the microbat Myotis velifer incautus, we serendipitously identified mRNAs encoding proteins with a high level of identity to herpesviruses. Next generation sequencing and de novo assembly of the viral genome from supernatants from Vero cells yielded a single contig of approximately 130 kilobases with at least 80 ORFs, predicted microRNAs and a gammaherpesvirus genomic organization. Phylogenetic analysis of the envelope glycoprotein (gB) and DNA polymerase (POLD1) revealed similarity to multiple gammaherpesvirus, including those from as yet uncultured viruses of the Rhadinovirus genus that were obtained by deep sequencing of bat tissues. Cumulatively, this study provides the first isolation and characterization of a replication competent bat gammaherpesvirus.
Project description:MicroRNA (miRNA) and endogenous siRNA (endo-siRNA) are two essential classes of small noncoding RNAs (sncRNAs) in eukaryotic organisms. The class of miRNA is diverse and there exist noncanonical miRNAs that bypass the canonical miRNA biogenesis pathway. In order to identify noncanonical miRNAs and endo-siRNAs responding to virus infection and study their potential function, we sequenced small-RNA species from cells lytically infected with murine gammaherpesvirus 68. In addition to 3 novel canonical miRNAs in mouse, two antisense miRNAs in virus and 25 novel noncanonical miRNAs, including miRNAs derived from tRNAs, snoRNAs and introns, in the host were identified. These noncanonical miRNAs exhibited features distinct from canonical miRNAs in the lengths and structures of miRNA hairpins as well as base pairings and first nucleotide preference. Many of the novel miRNAs are conserved in mammals. In addition to several known murine endo-siRNAs detected by the sequencing profiling, a novel locus in the mouse genome was identified to give rise to endo-siRNAs. This novel endo-siRNA locus is comprised of two tandem inverted B4 short interspersed nuclear elements (SINEs). Unexpectedly, the SINE-derived endo-siRNAs were found in a variety of sequencing data as well as virus-infected cells. Moreover, a murine miRNA was up-regulated more than 35 fold in infected than in mock-treated cells. The putative target genes of the viral and the up-regulated murine miRNAs were potentially involved in processes of gene transcription and protein phosphorylation and localized to membranes, suggesting their role in manipulating the host basal immune system during lytic infection. Our results extended the number of noncanonical miRNAs in mammals and shed new lights on their potential functions of lytic infection of MHV68. Mouse NIH 3T12 cells infectd with MHV68 (3 samples) and mock-treated (2 samples) were examined. Noncanonical microRNAs and endogenous siRNAs discovery in lytic infection of murine gammaherpesvirus MHV68 (NC_001826.2).
Project description:Bats are a major reservoir of zoonotic viruses, and there has been growing interest in characterizing bat-specific features of innate immunity and inflammation. Recent studies have revealed bat-specific adaptations affecting interferon (IFN) signaling and IFN-stimulated genes (ISGs), but we still have a limited understanding of the genetic mechanisms that have shaped the evolution of bat immunity. Here we investigated the transcriptional and epigenetic dynamics of transposable elements (TEs) during the type I IFN response in little brown bat (Myotis lucifugus) primary embryonic fibroblast cells, using RNA-seq and CUT&RUN. We found multiple bat-specific TEs that undergo both locus-specific and family-level transcriptional upregulation in response to IFN. Our transcriptome reassembly identified multiple ISGs that have acquired novel exons from bat-specific TEs, including NRLC5, SLNF5 and a previously unannotated isoform of the IFITM2 gene. We also identified examples of TE-derived regulatory elements, but did not find strong evidence supporting genome-wide epigenetic activation of TEs in response to IFN. Collectively, our study uncovers numerous TE-derived transcripts, proteins, and alternative isoforms that are induced by IFN in Myotis lucifugus cells, highlighting potential candidate loci that contribute to bat-specific immune function.
Project description:Bats are natural hosts for a wide diversity of viruses. While many of these viruses are highly pathogenic in humans, most do not appear to cause major symptoms in bats. These modern bat-specific characteristics are the result of past virus-host (co)evolution and virus-driven host adaptations. Innate immunity is the first line of defense against viruses in mammals, we aim at characterizing bat innate immunity in response to viruses. Using genome-wide and gene candidate evolutionary analyses, we found that many bat antiviral genes have undergone multiple duplication events in a lineage-specific manner, specifically in the Myotis bat lineage. We focus on Myotis yumanensis as a model in the Myotis lineage. We performed transcriptomic analyses and observed the upregulation of most mammalian genes implicated in the different steps of the innate immune response from sensing to interferon-stimulated genes (ISGs), showing the conservation of the core innate immunity. Our study will contribute to identifying adaptations that shaped bat innate immunity. These adaptations may contribute to the bat-virus specificity and influence viral emergence to another mammalian host