Knockdown of NEAT1 restricts dengue virus replication by augmenting interferon alpha-inducible protein 27 via the RIG-I pathway.

The lncRNA NEAT1 plays a vital role in mitochondrial function and antiviral response. We have previously identified NEAT1 as dysregulated lncRNAs and found an inverse correlation with interferon alpha-inducible protein 27 (IFI27) expression associated with developing dengue severity. However, the role of NEAT1 in dengue virus (DV) infection remains elusive. Here, we undertook a study to evaluate the functional consequences of NEAT1 and IFI27 modulation on antiviral response and viral replication in dengue infection. We observed that the knockdown of NEAT1 augmented IFI27 expression and antiviral response via the RIG-I pathway. Increased antiviral response leads to a decrease in dengue viral replication. Further study suggested that the knockdown of IFI27 augmented expression of the activating transcription factor 3 (ATF3), a negative regulator of antiviral response, and increased dengue virus replication suggesting an important role played by IFI27 in mediating antiviral response. RNA sequencing study confirmed several mitochondrial genes significantly altered upon knockdown of NEAT1 in DV-infected cells. We further verified the effect of NEAT1 knockdown on mitochondrial functions. We observed a reduced level of phospho-DRP1(S616) expression along with elongated mitochondria in DV2-infected cells. Further, NEAT1 knockdown or ectopic expression of IFI27 increased mitochondrial ROS production and cell death via activation of caspase 3. Our study points to the crucial role of NEAT1 and IFI27 in mediating antiviral response and mitochondrial dysfunction in dengue infection.

High-Throughput RNA Sequencing Analysis of Plasma Samples Reveals Circulating microRNA Signatures with Biomarker Potential in Dengue Disease Progression.

The circulating microRNA (miRNA) profile has been widely used for identifying potential biomarkers against viral infections. However, data on circulating microRNA expression patterns in dengue patients are scanty. Considering the impact of severity caused by dengue infection, circulating miRNA profiles in plasma of dengue patients may prove to be valuable for developing early prognostic markers for the disease severity. Here, we described an in-depth analytical study of small RNA sequencing data obtained from the plasma of 39 dengue patients. Integrating bioinformatics and in vitro studies, we identified differentially expressed miRNAs (DEMs) (log2 fold change ≥1.5, P < 0.05) associated with dengue disease progression. In comparing miRNA expression pattern with the follow-up samples, nine miRNAs were found to exhibit an altered expression that could distinguish between severe dengue and the convalescent patients. To understand the abundance and specificity of the DEMs in the context of dengue infection and disease progression, eight top-hit DEMs were further validated in the dengue virus-infected cell lines as well as in the patient's plasma and peripheral blood mononuclear cells (PBMCs) using the quantitative reverse transcription-PCR (qRT-PCR) method. Importantly, receiver operating curve analysis further confirmed that the plasma expression pattern of hsa-miR-122-5p could differentiate between different stages of dengue infection (area under the concentration-time curve [AUC] = 0.792), and dengue-negative patients with other febrile illnesses (AUC = 0.984). The in silico analysis of DEM target genes suggested an enrichment of the pathways associated with metabolism and inflammation. Our study gives a global view of miRNA expression in the plasma from dengue patients and provides a precious resource of candidate miRNAs involved in dengue infection and disease progression.IMPORTANCE Dengue virus (DENV) infection usually causes dengue fever (DF) with flu-like illness affecting infants, young children, and adults. The DF occasionally evolves into a potentially lethal complication called dengue severe (DS) leading to a rapid fall in platelet count along with plasma leakage, fluid accumulation, respiratory distress, and severe bleeding. The diverse clinical spectrum of dengue disease, as well as its significant similarity to other febrile viral illnesses, makes early identification more challenging in this high-risk group. microRNAs (miRNAs) are small (∼19 to 21 nucleotides [nt] in length), noncoding RNAs, extremely stable and easily detectable in the plasma; thus, they have potential as biomarkers for diagnosing and monitoring human diseases. This study provides a comprehensive analysis of miRNAs circulating in plasma of dengue virus-infected patients and identifies the miRNA signatures that have biomarker potential for dengue infection and disease progression.