Cytomegalovirus microRNAs Facilitate Persistent Virus Infection in Salivary Glands

Micro (mi)RNAs are small non-coding RNAs that regulate the expression of their targets' messenger RNAs through both translational inhibition and regulation of target RNA stability. Recently, a number of viruses, particularly of the herpesvirus family, have been shown to express their own miRNAs to control both viral and cellular transcripts. Although some targets of viral miRNAs are known, their function in a physiologically relevant infection remains to be elucidated. As such, no in vivo phenotype of a viral miRNA knock-out mutant has been described so far. Here, we report on the first functional phenotype of a miRNA knock-out virus in vivo. During subacute infection of a mutant mouse cytomegalovirus lacking two viral miRNAs, virus production is selectively reduced in salivary glands, an organ essential for virus persistence and horizontal transmission. This phenotype depends on several parameters including viral load and mouse genetic background, and is abolished by combined but not single depletion of natural killer (NK) and CD4+ T cells. Together, our results point towards a miRNA-based immunoevasion mechanism important for long-term virus persistence

MicroRNA degradation by a conserved target RNA regulates animal behavior

International audiencemicroRNAs (miRNAs) repress target transcripts through partial complementarity. By contrast, highly complementary miRNA-binding sites within viral and artificially engineered transcripts induce miRNA degradation in vitro and in cell lines. Here, we show that a genome-encoded transcript harboring a near-perfect and deeply conserved miRNA-binding site for miR-29 controls zebrafish and mouse behavior. This transcript originated in basal vertebrates as a long noncoding RNA (lncRNA) and evolved to the protein-coding gene NREP in mammals, where the miR-29-binding site is located within the 3′ UTR. We show that the near-perfect miRNA site selectively triggers miR-29b destabilization through 3′ trimming and restricts its spatial expression in the cerebellum. Genetic disruption of the miR-29 site within mouse Nrep results in ectopic expression of cerebellar miR-29b and impaired coordination and motor learning. Thus, we demonstrate an endogenous target-RNA-directed miRNA degradation event and its requirement for animal behavio

Transcriptional Activation of the Adenoviral Genome Is Mediated by Capsid Protein VI

Gene expression of DNA viruses requires nuclear import of the viral genome. Human Adenoviruses (Ads), like most DNA viruses, encode factors within early transcription units promoting their own gene expression and counteracting cellular antiviral defense mechanisms. The cellular transcriptional repressor Daxx prevents viral gene expression through the assembly of repressive chromatin remodeling complexes targeting incoming viral genomes. However, it has remained unclear how initial transcriptional activation of the adenoviral genome is achieved. Here we show that Daxx mediated repression of the immediate early Ad E1A promoter is efficiently counteracted by the capsid protein VI. This requires a conserved PPxY motif in protein VI. Capsid proteins from other DNA viruses were also shown to activate the Ad E1A promoter independent of Ad gene expression and support virus replication. Our results show how Ad entry is connected to transcriptional activation of their genome in the nucleus. Our data further suggest a common principle for genome activation of DNA viruses by counteracting Daxx related repressive mechanisms through virion proteins

Degradation of cellular mir-27 by a novel, highly abundant viral transcript is important for efficient virus replication in vivo.

Cytomegaloviruses express large amounts of viral miRNAs during lytic infection, yet, they only modestly alter the cellular miRNA profile. The most prominent alteration upon lytic murine cytomegalovirus (MCMV) infection is the rapid degradation of the cellular miR-27a and miR-27b. Here, we report that this regulation is mediated by the ∼1.7 kb spliced and highly abundant MCMV m169 transcript. Specificity to miR-27a/b is mediated by a single, apparently optimized, miRNA binding site located in its 3'-UTR. This site is easily and efficiently retargeted to other cellular and viral miRNAs by target site replacement. Expression of the 3'-UTR of m169 by an adenoviral vector was sufficient to mediate its function, indicating that no other viral factors are essential in this process. Degradation of miR-27a/b was found to be accompanied by 3'-tailing and -trimming. Despite its dramatic effect on miRNA stability, we found this interaction to be mutual, indicating potential regulation of m169 by miR-27a/b. Most interestingly, three mutant viruses no longer able to target miR-27a/b, either due to miRNA target site disruption or target site replacement, showed significant attenuation in multiple organs as early as 4 days post infection, indicating that degradation of miR-27a/b is important for efficient MCMV replication in vivo

Regulation of microRNA biogenesis and turnover by animals and their viruses

Item does not contain fulltextMicroRNAs (miRNAs) are a ubiquitous component of gene regulatory networks that modulate the precise amounts of proteins expressed in a cell. Despite their small size, miRNA genes contain various recognition elements that enable specificity in when, where and to what extent they are expressed. The importance of precise control of miRNA expression is underscored by functional studies in model organisms and by the association between miRNA mis-expression and disease. In the last decade, identification of the pathways by which miRNAs are produced, matured and turned-over has revealed many aspects of their biogenesis that are subject to regulation. Studies in viral systems have revealed a range of mechanisms by which viruses target these pathways through viral proteins or non-coding RNAs in order to regulate cellular gene expression. In parallel, a field of study has evolved around the activation and suppression of antiviral RNA interference (RNAi) by viruses. Virus encoded suppressors of RNAi can impact miRNA biogenesis in cases where miRNA and small interfering RNA pathways converge. Here we review the literature on the mechanisms by which miRNA biogenesis and turnover are regulated in animals and the diverse strategies that viruses use to subvert or inhibit these processes

The Lid Domain of Caenorhabditis elegans Hsc70 Influences ATP Turnover, Cofactor Binding and Protein Folding Activity

Hsc70 is a conserved ATP-dependent molecular chaperone, which utilizes the energy of ATP hydrolysis to alter the folding state of its client proteins. In contrast to the Hsc70 systems of bacteria, yeast and humans, the Hsc70 system of C. elegans (CeHsc70) has not been studied to date

Experimental investigations of buckling of steel, pressurised spherical shells

Cienkie powłoki stalowe w postaci wycinka sfery obciążone ciśnieniem zewnętrznym wykazują tendencje do utraty stateczności w fazie poprzedzającej wyczerpanie nośności plastycznej. Zjawisko wyboczenia powłok sferycznych było przedmiotem badań wielu autorów, a wyniki dotychczasowych badań teoretycznych i doświadczalnych zostały wykorzystane w zaleceniach projektowych wydanych w 2008 roku przez ECCS [9]. W pracy przedstawiono badania eksperymentalne wykonane na specjalnie w tym celu zaprojektowanym stanowisku badawczym. Próbki do badań zostały wykonane techniką wyoblania z blach przeznaczonych do głębokiego tłoczenia. Zinwentaryzowano geometrię początkową każdej z powłok za pomocą skanowania 3D i wykonano badania materiałowe stali użytej do wykonania powłok. Ciśnienie wymuszano za pomocą kompresora powietrza, a pomiary przemieszczeń były wykonywane przy zastosowaniu optycznego systemu pomiarowego Pontos-Aramis. Rejestrowano formy deformacji całej powłoki w trakcie stopniowego wzrostu ciśnienia także po utracie stateczności. Wszystkie dane pomiarowe rejestrowano automatycznie za pomocą dwóch komputerów. Wynikiem każdego z eksperymentów była nieliniowa ścieżka równowagi w postaci zależności ciśnienia od wybranego parametru przemieszczeniowego. W artykule zaprezentowano porównanie otrzymanych wartości ciśnień krytycznych z wynikami symulacji numerycznych wykonanych systemem COSMOS/M dla geometrii nominalnej oraz porównanie z wartościami nośności wyboczeniowych wynikających z zapisów EDR5th ECCS [9] oraz z propozycji przedstawionej w pracy [3].Thin steel shells in a form of spherical segment loaded by external pressure show a tendency to sudden buckling for a lower pressure than their plastic resistance. The buckling phenomenon of spherical shells was subject of investigations of many authors and hitherto results of theoretical and experimental research were used in European Design Recommendations published in 2008 by ECCS [9]. Experimental investigations performed on the deliberately designed and fabricated stand were presented in this paper. Specimens were manufactured by a metal spinning technique from steel sheets used usually to a deep drawing and stamping of metal objects. The actual shape of every specimen was determined by 3D scanning technique. Material investigations of steel sheets were performed as well. The pressure was exerted by the air compressor and displacement measurements were accomplished by optical measurement system Pontos-Aramis. Forms of deformations of the whole specimen were registered precisely at every step of pressure exertion till the final postbuckling stage. All measured data were registered automatically by means of two computers. The result of every experiment was presented in the form of equilibrium path as the relationship of the pressure p as a function of the chosen displacement parameter. The comparison of critical pressures obtained in experiments with results received in preliminary numerical simulations done by COSMOS/M was presented in the paper. Experimental results were compared also with buckling resistances proposed in [9] and [3]