The lncRNA NRON modulates HIV-1 replication in a NFAT-dependent manner and is differentially regulated by early and late viral proteins

A majority of the human genome is transcribed into noncoding RNAs, of which the functions of long noncoding RNAs (lncRNAs) are poorly understood. Many host proteins and RNAs have been characterized for their roles in HIV/AIDS pathogenesis, but there is only one lncRNA, NEAT1, which is shown to affect the HIV-1 life cycle. We profiled 90 disease-related lncRNAs and found NRON (noncoding repressor of Nuclear Factor of Activated T cells [NFAT]) to be one of several lncRNAs whose expression was significantly altered following HIV-1 infection. The regulation of NRON expression during the HIV-1 life cycle was complex; its levels were reduced by the early viral accessory protein Nef and increased by the late protein Vpu. Consequently, Nef and Vpu also modulated activity of the transcription factor NFAT. The knockdown of NRON enhanced HIV-1 replication through increased activity of NFAT and the viral LTR. Using siRNA-mediated NFAT knockdown, we show the effects of NRON on HIV-1 replication to be mediated by NFAT, and the viral Nef and Vpu proteins to modulate NFAT activity through their effects on NRON. These findings add the lncRNA, NRON to the vast repertoire of host factors utilized by HIV for infection and persistence

The Hepatitis E Virus ORF3 Protein Regulates the Expression of Liver-Specific Genes by Modulating Localization of Hepatocyte Nuclear Factor 4

The hepatitis E virus (HEV) is a small RNA virus and the cause of acute viral hepatitis E. The open reading frame 3 protein (pORF3) of HEV appears to be a pleiotropic regulatory protein that helps in the establishment, propagation and progression of viral infection. However, the global cellular effects of this protein remain to be explored. In the absence of traditional in vitro viral infection systems or efficient replicon systems, we made an adenovirus based ORF3 protein expression system to study its effects on host cell gene expression. We infected Huh7 hepatoma cells with recombinant adenoviruses expressing pORF3 and performed microarray-based gene expression analyses. Several genes down regulated in pORF3-expressing cells were found to be under regulation of the liver-enriched hepatocyte nuclear factor 4 (HNF4), which regulates hepatocyte-specific gene expression. While HNF4 localizes to the nucleus, its phosphorylation results in impaired nuclear localization of HNF4. Here we report that pORF3 increases HNF4 phosphorylation through the ERK and Akt kinases, which results in impaired nuclear translocation of HNF4 and subsequently the down modulation of HNF4-responsive genes in pORF3-expressing cells. We propose that modulation of several hepatocyte specific genes by pORF3 will create an environment favorable for viral replication and pathogenesis

Molecular virology of hepatitis E virus

This review details the molecular virology of the hepatitis E virus (HEV). While replicons and in vitro infection systems have recently become available, a lot of information on HEV has been generated through comparisons with better-studied positive-strand RNA viruses and through subgenomic expression of viral open reading frames. These models are now being verified with replicon and infection systems. We provide here the current knowledge on the HEV genome and its constituent proteins - ORF1, ORF2 and ORF3. Based on the available information, we also modify the existing model of the HEV life cycle

MicroRNA-150 is a potential biomarker of HIV/AIDS disease progression and therapy.

BACKGROUND: The surrogate markers of HIV/AIDS progression include CD4 T cell count and plasma viral load. But, their reliability has been questioned in patients on anti-retroviral therapy (ART). Five microRNAs (miRNAs) - miR-16, miR-146b-5p, miR-150, miR-191 and miR-223 in peripheral blood mononuclear cells (PBMCs) were earlier found to assign HIV/AIDS patients into groups with varying CD4 T cell counts and viral loads. In this pilot study, we profiled the expression of these five miRNAs in PBMCs, and two of these miRNAs (miR-146b-5p and miR-150) in the plasma of HIV/AIDS patients, including those on ART and those who developed ART resistance, to evaluate if these are biomarkers of disease progression and therapy. RESULTS: We quantified miRNA levels by quantitative reverse transcription polymerase chain reaction (qRT-PCR) using RNA isolated from PBMCs and plasma of healthy persons or HIV-infected patients who were (1) asymptomatic; (2) symptomatic and ART naïve; (3) on ART; and (4) failing ART. Our results show miR-150 (p<0.01) and to a lesser extent miR-146b-5p (p<0.05) levels in PBMCs to reliably distinguish between ART-naïve AIDS patients, those on ART, and those developing drug resistance and failing ART. The plasma levels of these two miRNAs also varied significantly between patients in these groups and between patients and healthy controls (p values <0.05). CONCLUSIONS: We report for the first time that PBMC and plasma levels of miR-150 and miR-146b-5p are predictive of HIV/AIDS disease progression and therapy

Relative plasma levels of miRNAs.

<p>The expression levels (mean ± SEM) of plasma miR-150 (a, b) and miR-146b-5p (c, d) normalized to miR-16 (a, c) and cel-miR-39 (b, d) in different groups of HIV patients and healthy controls are shown. The asterisks denote statistically significant differences (** p<0.01, * p<0.05) for the indicated groups (Student's T test).</p

Correlation of miRNA levels with CD4 counts and viral load.

<p>The correlations are shown for (a) miR-150 in PBMCs with absolute CD4+ T-cell counts and (b) miR-146b-5p in PBMCs with HIV viral loads. A positive significant correlation was noted between miR-150 and CD4 cell count (r = 0.64; p<0.01) and an inverse correlation between miR-146b-5p and HIV viral loads (r = −0.34; p<0.05).</p

Differential expression of the indicated miRNAs in the PBMCs of healthy controls and HIV/AIDS patients at different stages of disease.

<p>The values in the bar graphs are given as mean ± SEM, and asterisks denote statistically significant differences (** p<0.01, * p<0.05) for the indicated groups (Student's T test).</p

PBMC miRNAs as classifiers for different sets of patients.

<p>Receiver operating characteristic (ROC) curve analysis is shown for the expression ratio of (a) miR-150/RNU44 as classifier of CD4+ T-cell counts, (b) miR-146b-5p/RNU44 as classifier of CD4+ T-cell counts, and (c) miR-16/RNU44 as a classifier of viral load. Comparisons are made between Set-A (healthy, HIV+ asymptomatic and patients on ART) versus Set-B (HIV+ symptomatic and ART resistance patients), as described in the text.</p

miRNAs as classifiers for patients without and with ART.

<p>Receiver operating characteristic (ROC) curve analysis of expression ratio of (a) PBMC miR-150/RNU-44, (b) plasma miR-150/miR-16, (c) PBMC miR-146b-5p/RNU-44 and (d) plasma miR-146b-5p/miR-16, as classifier of Set-C (HIV+; asymptomatic and symptomatic) versus Set-D (patients on ART) shown on left panels; and Set-D versus Set-E (ART resistance patients) shown on right panels. Insets of plots show area under curve (AUC), standard error (SE) and significance (p value).</p

Chemically activatable viral capsid functionalized for cancer targeting.

AimTo design a theranostic capsule using the virus-like nanoparticle of the hepatitis E virus modified to display breast cancer cell targeting functional group (LXY30).MethodsFive surface-exposed residues were mutated to cysteine to allow conjugation to maleimide-linked chemical groups via thiol-selective linkages. Engineered virus-like nanoparticles were then covalently conjugated to a breast cancer recognized ligand, LXY30 and an amine-coupled near-infrared fluorescence dye.ResultsLXY30-HEV VLP was checked for its binding and entry to a breast cancer cell line and for tumor targeting in vivo to breast cancer tissue in mice. The engineered virus-like nanoparticle not only targeted cancer cells, but also appeared immune silent to native hepatitis E virus antibodies due to epitope disruption at the antibody-binding site.ConclusionThese results demonstrate the production of a theranostic capsule suitable for cancer diagnostics and therapeutics based on surface modification of a highly stable virus-like nanoparticle