ABSTRACT
[This corrects the article DOI: 10.1371/journal.pone.0169081.].
ABSTRACT
Important roles of microRNAs (miRNAs) in regulating the host response during viral infection have begun to be defined. However, little is known about the functional roles of miRNAs within an in vivo acute viral encephalitis model. We therefore identified global changes in miRNA expression during acute herpes simplex virus type 1 (HSV-1) encephalitis (HSVE) in mice. We found that many of the highly upregulated miRNAs (miR-155, miR-146a and miR-15b) detected in HSV-1 infected brain tissue are known regulators of inflammation and innate immunity. We also observed upregulation of 7 members belonging to the related group of miRNAs, the miR-200 family and miR-182 cluster (miR-200/182). Using in situ hybridization, we found that these miRNAs co-localized to regions of the brain with severe HSVE-related pathology and were upregulated in various cell types including neurons. Induction was apparent but not limited to cells in which HSV-1 was detected by immunohistochemistry, suggesting possible roles of these miRNAs in the host response to viral-induced tissue damage. Bioinformatic prediction combined with gene expression profiling revealed that the induced miR-200/182 members could regulate the biosynthesis of heparan sulfate proteoglycans. Using luciferase assays, we found that miR-96, miR-141, miR-183 and miR-200c all potentially targeted the syndecan-2 gene (Sdc2), which codes for a cell surface heparan sulfate proteoglycan involved in HSV-1 cellular attachment and entry.
Subject(s)
Brain/metabolism , Encephalitis, Herpes Simplex/genetics , MicroRNAs/genetics , Syndecan-2/genetics , Acute Disease , Animals , Brain/virology , Chlorocebus aethiops , Computational Biology , Encephalitis, Herpes Simplex/virology , Female , Gene Expression Profiling , Gene Expression Regulation, Viral , Heparan Sulfate Proteoglycans/metabolism , Herpesvirus 1, Human/genetics , Immunity, Innate , Inflammation , Mice , Real-Time Polymerase Chain Reaction , Transcriptome , Up-Regulation , Vero CellsABSTRACT
Second-generation high-throughput sequencing is a robust and inexpensive methodology that is becoming an increasingly common technique for the study of microRNA (miRNA) expression levels in the central nervous system. This method allows for the identification of both known and novel miRNAs, reporting on the qualitative and quantitative levels these RNA species represent in any given sample. Numerous bioinformatic programs are currently available to analyze deep sequencing data but many require at least a partial understanding of the command line interface. In this chapter, we describe a user-friendly computational workflow guiding the user through the process from the initial FASTQ deep sequencing file to the identification of known and potentially novel miRNAs in a given experiment, as well as the assessment of the differential expression of these miRNAs between experimental samples. Furthermore, programs that can predict potential targets for these miRNAs are also highlighted.
