Closing in on the causes of host shutoff

How influenza virus A drives virus-induced host shutoff When a virus enters a cell it relies on the molecular machinery of its host to help it replicate. In particular, the virus relies on the ribosomes in the host cell to translate viral messenger RNA (mRNA) into polypeptides. Many viruses also impair the translation of cellular mRNA, via a process termed “host shutoff”, in order to prevent the production of anti-viral, host defense proteins. For example, poliovirus does this by inactivating a translation factor that is required to load ribosomes onto host mRNAs, all of which have a type of “cap” called an “m7G cap”. Moreover, while the translation of these mRNAs is being suppressed, host ribosomes are involved in the translation of poliovirus mRNA, which does not have a cap: this is possible without the translation factor because poliovirus mRNA has an internal entry site for ribosomes. However, the mechanisms responsible for host shutoff in viruses that have mRNAs with m7G-caps, such as influenza A virus, have remained enigmatic.

A new paper in eLife shows that in Influenza virus-infected cells host and viral mRNAs were both translated with similar efficiencies, indicating that viral mRNAs were not preferentially translated relative to host mRNAs. Instead, Influenza virus-induced host shutoff primarily originates from a reduced abundance of cellular mRNA and from the high levels of viral mRNA in both the nucleus and cytoplasm. Fluorescence-based measurements confirmed these findings and revealed that the reduced abundance of cellular mRNA has its origins in the nucleus. This likely involves an RNA endoribonuclease called PA-X, which is encoded in the genome of Influenza virus, stimulating the decay of cellular mRNA.

A systematic view on influenza induced host shutoff. (2016) eLife 5: e18311. doi: 10.7554/eLife.18311
Host shutoff is a common strategy used by viruses to repress cellular mRNA translation and concomitantly allow the efficient translation of viral mRNAs. Here we use RNA-sequencing and ribosome profiling to explore the mechanisms that are being utilized by the Influenza A virus (IAV) to induce host shutoff. We show that viral transcripts are not preferentially translated and instead the decline in cellular protein synthesis is mediated by viral takeover on the mRNA pool. Our measurements also uncover strong variability in the levels of cellular transcripts reduction, revealing that short transcripts are less affected by IAV. Interestingly, these mRNAs that are refractory to IAV infection are enriched in cell maintenance processes such as oxidative phosphorylation. Furthermore, we show that the continuous oxidative phosphorylation activity is important for viral propagation. Our results advance our understanding of IAV-induced shutoff, and suggest a mechanism that facilitates the translation of genes with important housekeeping functions.

eLife: Virology: Closing in on the causes of host shutoff. doi: 10.7554/eLife.20755

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Latency in Herpes Simplex Virus 1

HSV-infected cell Herpes simplex virus 1 (HSV-1) is a neurotropic virus that establishes a life-long latent infection in the trigeminal ganglia (TG) of the infected human host. From time to time the virus asymptomatically or symptomatically reactivates from the latency stage producing epithelial lesions, most of the time on the face but also in the eye, inducing severe pathologies such as keratitis. HSV-1 infection is also associated with pathologies of the central nervous system such as encephalitis.

During latency the virus is in a transcriptionally restricted state. Of the about 80 genes transcribed during lytic infection, only a family of long non-coding RNAs is produced abundantly during latency. These latency associated transcripts (LATs) arise from the transcription of an 8.3 kb primary RNA that is processed in two major LATs of 1.5 kb and 2 kb and several microRNAs with cellular and viral targets. The precise role of LATs is a matter of debate; however, a point of convergence among the many studies of LATs is that their initial production would favor the survival of the infected neurons and the coordination of the infectious process towards the latency transcriptional program and reactivation. The lytic cycle is the alternative transcriptional program and is characterized by a temporarily regulated transcriptional program, which starts with the expression of immediate early (IE), then early (E), and finally late (L) genes.

In the virus particle, HSV-1 genome is a 150-kb double stranded naked linear DNA. Upon entry into the host nucleus, the viral genome does not integrate, remaining as an extrachromosomal plasmid-like molecule. Chromatinization of the viral genome during latency plays a major regulatory role, and post-translational modifications of histones associated to key viral promoters determines the fate of the latency/reactivation process. Latent viral genomes are present in multiple copies within the nucleus of infected neurons in mouse models and human, and little is known about the molecular determinants that enable one neuron rather than another to sustain reactivation.

This paper describes a fluorescent in situ hybridization (FISH) approach combined with immunofluorescence to investigate the interaction between viral genomes and nuclear proteins within TG neurons of latently infected mice and during the whole process of latency establishment (from 4 to 28 days post infection, dpi). Viral genomes were found in neurons and satellite cells at 4 and 6 dpi, but only in neurons at > 6 dpi. In satellite cells, viral genomes showed only replication compartment (RC) patterns, whereas in neurons both RC and “multiple-acute” patterns were detected. From 4 to 14 dpi both patterns progressively disappeared, and transformed from14dpi onwards to the latency-associated “single” and “multiple-latency” patterns. Immuno-FISH analyses of human TG showed a close spatial distribution between latent HSV-1 genomes and PML protein in neurons, which suggests that, similar to the situation in the mouse model, HSV-1 latency in human is probably tightly linked to the activity of PML-NBs.

We have been trying for decades to understand the biology of latency in HSV. The strength of this approach is to point out that this cannot be achieved by looking at the virus alone, the biology of the host cell also has to be considered. This study describes the nuclear architecture and nuclear distribution of viral genomes as major determinants of HSV-1 latency. It confirms the close interrelation between PML-NBs and HSV-1 genomes in the establishment of latency through the formation of vDCP-NBs. Finally, it confirms that the spatial organization of HSV-1 genomes and PML is conserved in latently infected neurons in human TG, which indicates PML-NBs to be major HSV-1 genome interactants during latency and probably reactivation.

 

Latency Entry of Herpes Simplex Virus 1 Is Determined by the Interaction of Its Genome with the Nuclear Environment. (2016) PLoS Pathog 12(9): e1005834. doi: 10.1371/journal.ppat.1005834
Herpes simplex virus 1 (HSV-1) establishes latency in trigeminal ganglia (TG) sensory neurons of infected individuals. The commitment of infected neurons toward the viral lytic or latent transcriptional program is likely to depend on both viral and cellular factors, and to differ among individual neurons. In this study, we used a mouse model of HSV-1 infection to investigate the relationship between viral genomes and the nuclear environment in terms of the establishment of latency. During acute infection, viral genomes show two major patterns: replication compartments or multiple spots distributed in the nucleoplasm (namely “multiple-acute”). Viral genomes in the “multiple-acute” pattern are systematically associated with the promyelocytic leukemia (PML) protein in structures designated viral DNA-containing PML nuclear bodies (vDCP-NBs). To investigate the viral and cellular features that favor the acquisition of the latency-associated viral genome patterns, we infected mouse primary TG neurons from wild type (wt) mice or knock-out mice for type 1 interferon (IFN) receptor with wt or a mutant HSV-1, which is unable to replicate due to the synthesis of a non-functional ICP4, the major virus transactivator. We found that the inability of the virus to initiate the lytic program combined to its inability to synthesize a functional ICP0, are the two viral features leading to the formation of vDCP-NBs. The formation of the “multiple-latency” pattern is favored by the type 1 IFN signaling pathway in the context of neurons infected by a virus able to replicate through the expression of a functional ICP4 but unable to express functional VP16 and ICP0. Analyses of TGs harvested from HSV-1 latently infected humans showed that viral genomes and PML occupy similar nuclear areas in infected neurons, eventually forming vDCP-NB-like structures. Overall our study designates PML protein and PML-NBs to be major cellular components involved in the control of HSV-1 latency, probably during the entire life of an individual.

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Human Hookworm Infection

Hookworms Hookworm affects approximately 500 million people worldwide, yet its global economic and health impact is not well understood. A new study suggests that the health and economic burden of hookworm infection is estimated to exceed those of a number of diseases receiving greater attention and investment. Human hookworm infection confers a substantial global health and economic burden through loss of productivity, and years of life living with disability due to infection outcomes. While hookworm infection rarely results in death, it can lead to iron-deficiency anemia and malnutrition. Chronic health problems resulting from these conditions include lethargy, impaired physical and cognitive development and adverse pregnancy outcomes.

The Global Economic and Health Burden of Human Hookworm Infection. (2016) PLoS Negl Trop Dis 10(9): e0004922. doi: 10.1371/journal.pntd.0004922
Even though human hookworm infection is highly endemic in many countries throughout the world, its global economic and health impact is not well known. Without a better understanding of hookworm’s economic burden worldwide, it is difficult for decision makers such as funders, policy makers, disease control officials, and intervention manufacturers to deter- mine how much time, energy, and resources to invest in hookworm control. We developed a computational simulation model to estimate the economic and health bur- den of hookworm infection in every country, WHO region, and globally, in 2016 from the societal perspective. Globally, hookworm infection resulted in a total 2,126,280 DALYs using 2004 disability weight estimates and 4,087,803 DALYs using 2010 disability weight estimates (excluding cognitive impairment outcomes). Including cognitive impairment did not significantly increase DALYs worldwide. Total productivity losses varied with the proba- bility of anemia and calculation method used, ranging from $7.5 billion to $138.9 billion annually using gross national income per capita as a proxy for annual wages and ranging from $2.5 billion to $43.9 billion using minimum wage as a proxy for annual wages. Even though hookworm is classified as a neglected tropical disease, its economic and health burden exceeded published estimates for a number of diseases that have received comparatively more attention than hookworm such as rotavirus. Additionally, certain large countries that are transitioning to higher income countries such as Brazil and China, still face considerable hookworm burden.

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The debt we owe to retroviruses

It was already known that genes inherited from ancient retroviruses are essential to the formation of the placenta in mammals. Now it appears that genes with retrovirus origins may also be responsible for the more developed muscle mass seen in males.

Retroviruses carry proteins on their surface that are able to mediate fusion of their envelope with the membrane of a target cell. Once released inside that cell, their genetic material becomes integrated in the host’s chromosomes. In the rare cases where the infected cell is involved in reproduction, the viral genes may be transmitted to progeny. Nearly 8% of the mammalian genome is made up of vestiges of retroviruses, or “endogenous” retroviruses. Most of them are inactive, but some remain capable of producing proteins. This is the case of syncytins, proteins that are present in all mammals and encoded by genes inherited from retroviruses captured by their ancestors. Five years ago it was discovered that syncytins contribute to formation of the placenta in mammals.

New research has revealed an additional and unexpected effect of these proteins: they endow males with more muscle mass than females. Like the syncytiotrophoblast, muscle mass develops from fused stem cells. In the genetically-modified male mice, these fibers were 20% smaller and displayed 20% fewer nuclei than in standard males; they were then similar to those seen in females, as was their total muscle mass. It therefore appears that the inactivation of syncytins leads to a fusion deficit during muscle growth, but only in males. Researchers observed the same phenomenon in the case of muscle regeneration following a lesion: the male mice incapable of producing syncytins experienced less effective regeneration than the other males, but it was comparable to that seen in females. Furthermore, the regenerating muscle fibers produced syncytin – once again, only in males.

If this discovery were to be confirmed in other mammals, it might account for the muscle dimorphism observed between males and females, a difference that is not seen so systematically in egg laying animals. By cultivating muscle stem cells from different mammalian species (mouse, sheep, dog, human), the scientists have advanced some way along the path: they indeed showed that syncytins contributed to the formation of muscle fibers in all the species tested. It is now necessary to demonstrate whether, in these species as well, the action of syncytins is also male-specific.

 

Genetic Evidence That Captured Retroviral Envelope syncytins Contribute to Myoblast Fusion and Muscle Sexual Dimorphism in Mice. (2016) PLoS Genet 12(9): e1006289. doi: 10.1371/journal.pgen.1006289
Syncytins are envelope genes from endogenous retroviruses, “captured” for a role in placentation. They mediate cell-cell fusion, resulting in the formation of a syncytium (the syncytiotrophoblast) at the fetomaternal interface. These genes have been found in all placental mammals in which they have been searched for. Cell-cell fusion is also pivotal for muscle fiber formation and repair, where the myotubes are formed from the fusion of mononucleated myoblasts into large multinucleated structures. Here we show, taking advantage of mice knocked out for syncytins, that these captured genes contribute to myoblast fusion, with a >20% reduction in muscle mass, mean muscle fiber area and number of nuclei per fiber in knocked out mice for one of the two murine syncytin genes. Remarkably, this reduction is only observed in males, which subsequently show muscle quantitative traits more similar to those of females. In addition, we show that syncytins also contribute to muscle repair after cardiotoxin-induced injury, with again a male-specific effect on the rate and extent of regeneration. Finally, ex vivo experiments carried out on murine myoblasts demonstrate the direct involvement of syncytins in fusion, with a >40% reduction in fusion index upon addition of siRNA against both syncytins. Importantly, similar effects are observed with primary myoblasts from sheep, dog and human, with a 20–40% reduction upon addition of siRNA against the corresponding syncytins. Altogether, these results show a direct contribution of the fusogenic syncytins to myogenesis, with a demonstrated male-dependence of the effect in mice, suggesting that these captured genes could be responsible for the muscle sexual dimorphism observed in placental mammals.

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Fighting Rabies in China

Rabies China has the second highest number of reported rabies cases in the world, but numbers of human deaths have been decreasing. According to the WHO, the Chinese authorities are forecasting national rabies elimination by 2025. Most of the cases are reported from the country’s Southeastern provinces. A new study reports on ongoing transmission of rabies in Northwestern China, highlighting challenges and opportunities pertinent to the elimination efforts.

Rabies virus continues to cross carnivorous species and to infect humans and livestock in China. Rabies vaccination of the principal reservoir animals is even being neglected in most regions of China, resulting in continuous expansion of rabies epidemics. Since there is no oral vaccine for stray dogs and wild animals and no inactivated vaccine for large domestic animals, rabies is not currently controlled in China. Rabies outbreaks are caused by bites of dogs and wild foxes and the long-term effects on protection against rabies using canine inactivated vaccines in domestic camels and cattle. At least three separate phylogenetic groups of rabies virus consistently exist and spread throughout Northwest China. Local canine vaccine products can be used to induce levels of virus neutralizing antibodies indicative of protection against rabies in cattle and camels, however licensed oral and inactivated vaccines for reservoir carnivores and large domestic animals are urgently needed for elimination of rabies in China.

Rabies Outbreaks and Vaccination in Domestic Camels and Cattle in Northwest China. (2016) PLoS Negl Trop Dis 10(9): e0004890. doi:10.1371/journal.pntd.0004890
In contrast to many countries where rabies has been well controlled in humans and livestock, even in wildlife, rabies is still endemic in almost regions of China. In Northwest China, rabies transmitted by stray dogs and wild foxes has caused heavy economic losses to local herdsmen, as well as causing numbers of human cases. In this study, as part of an investigation of ways to prevent rabies epidemics in livestock, we report an analysis of domestic cattle and camel rabies cases in Ningxia Hui (NHAR) and Inner Mongolia Autonomous Region (IMAR) and the immune efficacy of canine inactivated rabies vaccines in these ani- mals. We found that rabies viruses from these animals are closely related to dog-hosted China I and fox-associated China III lineages, respectively, indicating that the infections originated from two different sources (dogs and wild foxes). As well as the previously reported Arctic and Arctic-related China IV lineage in IMAR, at least three separate phylogenetic groups of rabies virus consistently exist and spread throughout Northwest China. Since there is no licensed oral vaccine for wild foxes and no inactivated vaccine for large livestock, local canine inactivated vaccine products were used for emergency immunization of beef and milk cattle and bactrian (two-humped) camels in local farms. Compared with a single injection with one (low-efficacy) or three doses (high-cost), a single injection of a double dose of canine vaccine provided low-price and convenience for local veterinarians while inducing levels of virus neutralizing antibodies indicative of protection against rabies for at least 1 year in the cattle and camels. However, licensed vaccines for wildlife and large domestic animals are still needed in China.

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The Ins and Outs of Multipartite Plant Viruses

Ins and Outs of Multipartite Plant Viruses Viruses possessing a non-segmented genome require a specific recognition of their nucleic acid to ensure its protection in a capsid. A similar feature exists for viruses having a segmented genome, usually consisting of viral genomic segments joined together into one viral entity. While this appears as a rule for animal viruses, the majority of segmented plant viruses package their genomic segments individually. To ensure a productive infection, all viral particles and thereby all segments have to be present in the same cell. Progression of the virus within the plant requires as well a concerted genome preservation to avoid loss of function. This review discusses the replication of chosen phytoviruses and argue for the existence of RNA-RNA interactions that drive the preservation of viral genome integrity while the virus progresses in the plant.

 

Ins and Outs of Multipartite Positive-Strand RNA Plant Viruses: Packaging versus Systemic Spread. Viruses 2016, 8(8), 228; doi: 10.3390/v8080228

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The Buzz about Honey Bee Viruses

The Buzz about Honey Bee Viruses This short review presents current understanding of the role of viruses on honey bee health and address some overarching questions in honey bee virology.

  • Why Should I Be Concerned about Honey Bee Colony Losses and What Is Colony Collapse Disorder (CCD)?
  • What Are the Most Common Viruses Infecting Honey Bees and How Do They Impact Bee Health?
  • How Are Bee Viruses Transmitted?
  • What Are the Mechanisms of Honey Bee Antiviral Defense?
  • What Is the Future of Honey Bee Virology?

 

The Buzz about Honey Bee Viruses. (2016) PLoS Pathog 12 (8): e1005757. doi: 10.1371/journal.ppat.1005757

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Assessing the global threat from Zika virus

Mosquito-carried diseases First discovered in 1947, Zika virus (ZIKV) infection remained a little-known tropical disease until 2015, when its apparent association with a considerable increase in the incidence of microcephaly in Brazil raised alarms worldwide. There is limited information on the key factors that determine the extent of the global threat from ZIKV infection and resulting complications. This review describes what is known about the epidemiology, natural history, and public health effects of ZIKV infection, the empirical basis for this knowledge, and the critical knowledge gaps that need to be filled.

“The evidence highlighted in this review is both encouraging and disheartening. On the one hand, the speed with which the global community has collected and disseminated clinical, epidemiologic, and laboratory information on ZIKV after identification of the threat is impressive. But the development of therapeutics and diagnostics is hampered by our ignorance, despite knowing of ZIKV’s existence for more than half a century. Consequently, we have been able to do little to contain the virus’s rapid spread across the Americas. New threats from infectious diseases may emerge from unexpected places, and we need strategies in place that we can roll out to rapidly gain an understanding of the transmission, pathogenesis, and control of previously little-known pathogens to protect global public health.”

 

Assessing the global threat from Zika virus. (2016) Science 353(6300): aaf8160. doi: 10.1126/science.aaf8160

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Where are things inside a bacterial cell?

Bacteria Bacterial cells are intricately organized, despite the lack of membrane-bounded organelles. The extremely crowded cytoplasm promotes macromolecular self-assembly and formation of distinct subcellular structures, which perform specialized functions. For example, the cell poles act as hubs for signal transduction complexes, thus providing a platform for the coordination of optimal cellular responses to environmental cues. Distribution of macromolecules is mostly mediated via specialized transport machineries, including the MreB cytoskeleton. Recent evidence shows that RNAs also specifically localize within bacterial cells, raising the possibility that gene expression is spatially organized. This review describes the current understanding of where things are in bacterial cells and discuss emerging questions that need to be addressed in the future.

  • Bacterial cells are intricately organized, with many proteins and RNAs being specifically localized.
  • The poles of rod-shaped bacterial cells are emerging as subcellular regions of importance for sensing and signaling.
  • The MreB cytoskeletal system plays an important role in cellular trafficking of macromolecules.
  • Subcellular domains in bacterial cells are connected through a complex network of interactions.

 

Where are things inside a bacterial cell? (2016) Current Opinion in Microbiology 33: 83–90. doi: 10.1016/j.mib.2016.07.003

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