In 2012, a novel betacoronavirus, designated Middle East respiratory syndrome coronavirus or MERS-CoV and associated with severe respiratory disease in humans, emerged in the Arabian Peninsula. To ...date, 108 human cases have been reported, including cases of human-to-human transmission. The availability of an animal disease model is essential for understanding pathogenesis and developing effective countermeasures. Upon a combination of intratracheal, ocular, oral, and intranasal inoculation with 7 × 10 ⁶ 50% tissue culture infectious dose of the MERS-CoV isolate HCoV-EMC/2012, rhesus macaques developed a transient lower respiratory tract infection. Clinical signs, virus shedding, virus replication in respiratory tissues, gene expression, and cytokine and chemokine profiles peaked early in infection and decreased over time. MERS-CoV caused a multifocal, mild to marked interstitial pneumonia, with virus replication occurring mainly in alveolar pneumocytes. This tropism of MERS-CoV for the lower respiratory tract may explain the severity of the disease observed in humans and the, up to now, limited human-to-human transmission.
The emergence of Middle East respiratory syndrome coronavirus (MERS-CoV) is of global concern: the virus has caused severe respiratory illness, with 111 confirmed cases and 52 deaths at the time of ...this article's publication. Therapeutic interventions have not been evaluated in vivo; thus, patient management relies exclusively on supportive care, which, given the high case-fatality rate, is not highly effective. The rhesus macaque is the only known model organism for MERS-CoV infection, developing an acute localized to widespread pneumonia with transient clinical disease that recapitulates mild to moderate human MERS-CoV cases. The combination of interferon-α2b and ribavirin was effective in reducing MERS-CoV replication in vitro; therefore, we initiated this treatment 8 h after inoculation of rhesus macaques. In contrast to untreated, infected macaques, treated animals did not develop breathing abnormalities and showed no or very mild radiographic evidence of pneumonia. Moreover, treated animals showed lower levels of systemic (serum) and local (lung) proinflammatory markers, in addition to fewer viral genome copies, distinct gene expression and less severe histopathological changes in the lungs. Taken together, these data suggest that treatment of MERS-CoV infected rhesus macaques with IFN-α2b and ribavirin reduces virus replication, moderates the host response and improves clinical outcome. As these two drugs are already used in combination in the clinic for other infections, IFN-α2b and ribavirin should be considered for the management of MERS-CoV cases.
The emergence of Middle East respiratory syndrome coronavirus (MERS-CoV) is of global concern: the virus has caused severe respiratory illness, with 111 confirmed cases and 52 deaths at the time of ...this article's publication. Therapeutic interventions have not been evaluated in vivo; thus, patient management relies exclusively on supportive care, which, given the high case-fatality rate, is not highly effective. The rhesus macaque is the only known model organism for MERS-CoV infection, developing an acute localized to widespread pneumonia with transient clinical disease that recapitulates mild to moderate human MERS-CoV cases. The combination of interferon- alpha 2b and ribavirin was effective in reducing MERS-CoV replication in vitro; therefore, we initiated this treatment 8 h after inoculation of rhesus macaques. In contrast to untreated, infected macaques, treated animals did not develop breathing abnormalities and showed no or very mild radiographic evidence of pneumonia. Moreover, treated animals showed lower levels of systemic (serum) and local (lung) proinflammatory markers, in addition to fewer viral genome copies, distinct gene expression and less severe histopathological changes in the lungs. Taken together, these data suggest that treatment of MERS-CoV infected rhesus macaques with IFN- alpha 2b and ribavirin reduces virus replication, moderates the host response and improves clinical outcome. As these two drugs are already used in combination in the clinic for other infections, IFN- alpha 2b and ribavirin should be considered for the management of MERS-CoV cases.
Mucosal sites are continuously exposed to pathogenic microorganisms and are therefore equipped to control respiratory infections. Type 3 innate lymphoid cells (ILC3) are key players in antimicrobial ...defense in intestinal mucosa, through interleukin 17 and interleukin 22 (IL-22) production. The present study aimed at analyzing the distribution and function of ILC3 in the respiratory tract. We first observed that lung mucosa harbors a discrete population of ILC3 expressing CD127, CD90, CCR6, and the transcriptional factor RORγt. In addition, lung ILC3 were identified as a major source of IL-22 in response to interleukin 23 stimulation. During Streptococcus pneumoniae infection, ILC3 rapidly accumulated in the lung tissue to produce IL-22. In response to S. pneumoniae, dendritic cells and MyD88, an important adaptor of innate immunity, play critical functions in IL-22 production by ILC3. Finally, administration of the Toll-like receptor 5 agonist flagellin during S. pneumoniae challenge exacerbated IL-22 production by ILC3, a process that protects against lethal infection. In conclusion, boosting lung ILC3 might represent an interesting strategy to fight respiratory bacterial infections.
The high mobility group protein A1 (HMGA1) is a master regulator of chromatin structure mediating its major gene regulatory activity by direct interactions with A/T-rich DNA sequences located in the ...promoter and enhancer regions of a large variety of genes. HMGA1 DNA-binding through three AT-hook motifs results in an open chromatin structure and subsequently leads to changes in gene expression. Apart from its significant expression during development, HMGA1 is over-expressed in virtually every cancer, where HMGA1 expression levels correlate with tumor malignancy. The exogenous overexpression of HMGA1 can lead to malignant cell transformation, assigning the protein a key role during cancerogenesis. Recent studies have unveiled highly specific competitive interactions of HMGA1 with cellular and viral RNAs also through an AT-hook domain of the protein, significantly impacting the HMGA1-dependent gene expression. In this review, we discuss the structure and function of HMGA1-RNA complexes during transcription and epigenomic regulation and their implications in HMGA1-related diseases.
Primary aldosteronism is the most prevalent form of secondary hypertension. To explore molecular mechanisms of autonomous aldosterone secretion, we performed exome sequencing of aldosterone-producing ...adenomas (APAs). We identified somatic hotspot mutations in the ATP1A1 (encoding an Na(+)/K(+) ATPase α subunit) and ATP2B3 (encoding a Ca(2+) ATPase) genes in three and two of the nine APAs, respectively. These ATPases are expressed in adrenal cells and control sodium, potassium and calcium ion homeostasis. Functional in vitro studies of ATP1A1 mutants showed loss of pump activity and strongly reduced affinity for potassium. Electrophysiological ex vivo studies on primary adrenal adenoma cells provided further evidence for inappropriate depolarization of cells with ATPase alterations. In a collection of 308 APAs, we found 16 (5.2%) somatic mutations in ATP1A1 and 5 (1.6%) in ATP2B3. Mutation-positive cases showed male dominance, increased plasma aldosterone concentrations and lower potassium concentrations compared with mutation-negative cases. In summary, dominant somatic alterations in two members of the ATPase gene family result in autonomous aldosterone secretion.
Virulence factors generally enhance a pathogen's fitness and thereby foster transmission. However, most studies of pathogen fitness have been performed by averaging the phenotypes over large ...populations. Here, we have analyzed the fitness costs of virulence factor expression by Salmonella enterica subspecies I serovar Typhimurium in simple culture experiments. The type III secretion system ttss-1, a cardinal virulence factor for eliciting Salmonella diarrhea, is expressed by just a fraction of the S. Typhimurium population, yielding a mixture of cells that either express ttss-1 (TTSS-1(+) phenotype) or not (TTSS-1(-) phenotype). Here, we studied in vitro the TTSS-1(+) phenotype at the single cell level using fluorescent protein reporters. The regulator hilA controlled the fraction of TTSS-1+ individuals and their ttss-1 expression level. Strikingly, cells of the TTSS-1(+) phenotype grew slower than cells of the TTSS-1(-) phenotype. The growth retardation was at least partially attributable to the expression of TTSS-1 effector and/or translocon proteins. In spite of this growth penalty, the TTSS-1(+) subpopulation increased from <10% to approx. 60% during the late logarithmic growth phase of an LB batch culture. This was attributable to an increasing initiation rate of ttss-1 expression, in response to environmental cues accumulating during this growth phase, as shown by experimental data and mathematical modeling. Finally, hilA and hilD mutants, which form only fast-growing TTSS-1(-) cells, outcompeted wild type S. Typhimurium in mixed cultures. Our data demonstrated that virulence factor expression imposes a growth penalty in a non-host environment. This raises important questions about compensating mechanisms during host infection which ensure successful propagation of the genotype.
For years, GluN3A was solely considered to be a dominant-negative modulator of NMDARs, since its incorporation into receptors alters hallmark features of conventional NMDARs composed of GluN1/GluN2 ...subunits. Only recently, increasing evidence has accumulated that GluN3A plays a more diversified role. It is considered to be critically involved in the maturation of glutamatergic synapses, and it might act as a molecular brake to prevent premature synaptic strengthening. Its expression pattern supports a putative role during neural development, since GluN3A is predominantly expressed in early pre- and postnatal stages. In this study, we used RNA interference to efficiently knock down GluN3A in 46C-derived neural stem cells (NSCs) both at the mRNA and at the protein level. Global gene expression profiling upon GluN3A knockdown revealed significantly altered expression of a multitude of neural genes, including genes encoding small GTPases, retinal proteins, and cytoskeletal proteins, some of which have been previously shown to interact with GluN3A or other iGluR subunits. Canonical pathway enrichment studies point at important roles of GluN3A affecting key cellular pathways involved in cell growth, proliferation, motility, and survival, such as the mTOR pathway. This study for the first time provides insights into transcriptome changes upon the specific knockdown of an NMDAR subunit in NSCs, which may help to identify additional functions and downstream pathways of GluN3A and GluN3A-containing NMDARs.
The transactivating response element （TAR） structure of the nascent HIV-1 transcript is critically involved in the recruitment of inactive positive transcription elongation factor b （P-TEFb） to the ...promoter proximal paused RNA polymerase II. The viral transactivator Tat is responsible for subsequent P-TEFb activation in order to start efficient viral transcription elongation. In the absence of the viral transactivator of transcription （Tat）, e.g., during latency or in early stages of HIV tran- scription, TAR mediates an interaction of P-TEFb with its inhibitor hexamethylene bis-aeetamide- inducible protein 1 （HEXIM1）, keeping P-TEFb in its inactive form. In this study, we address the function of HIV-1 TAR in the absence of Tat by analyzing consequences of HIV-1 TAR overexpres- sion on host cellular gene expression. An RNA chimera consisting of Epstein-Barr virus-expressed RNA 2 （EBER2） and HIV-1 TAR was developed to assure robust overexpression of TAR in HEK293 cells. The overexpression results in differential expression of more than 800 human genes. A significant proportion of these genes is involved in the suppression of cellular immune responses, including a significant set of 7SK-responsive P-TEFb target genes. Our findings identify a novel role for HIV-1 TAR in the absence of Tat, involving the interference with host cellular immune responses by targeting 7SK RNA-mediated gene expression and P-TEFb inactivation.
High-throughput molecular profiling and computational biology are changing the face of virology, providing a new appreciation of the importance of the host in viral pathogenesis and offering ...unprecedented opportunities for better diagnostics, therapeutics and vaccines. Here, we provide a snapshot of the evolution of systems virology, from global gene expression profiling and signatures of disease outcome, to geometry-based computational methods that promise to yield novel therapeutic targets, personalized medicine and a deeper understanding of how viruses cause disease. To realize these goals, pipettes and Petri dishes need to join forces with the powers of mathematics and computational biology.
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