Importin α1 is required for nuclear import of herpes simplex virus proteins and capsid assembly in fibroblasts and neurons.

Herpesviruses are large DNA viruses which depend on many nuclear functions, and therefore on host transport factors to ensure specific nuclear import of viral and host components. While some import cargoes bind directly to certain transport factors, most recruit importin ß1 via importin α. We identified importin α1 in a small targeted siRNA screen to be important for herpes simplex virus (HSV-1) gene expression. Production of infectious virions was delayed in the absence of importin α1, but not in cells lacking importin α3 or importin α4. While nuclear targeting of the incoming capsids, of the HSV-1 transcription activator VP16, and of the viral genomes were not affected, the nuclear import of the HSV-1 proteins ICP4 and ICP0, required for efficient viral transcription, and of ICP8 and pUL42, necessary for DNA replication, were reduced. Furthermore, quantitative electron microscopy showed that fibroblasts lacking importin α1 contained overall fewer nuclear capsids, but an increased proportion of mature nuclear capsids indicating that capsid formation and capsid egress into the cytoplasm were impaired. In neurons, importin α1 was also not required for nuclear targeting of incoming capsids, but for nuclear import of ICP4 and for the formation of nuclear capsid assembly compartments. Our data suggest that importin α1 is specifically required for the nuclear localization of several important HSV1 proteins, capsid assembly, and capsid egress into the cytoplasm, and may become rate limiting in situ upon infection at low multiplicity or in terminally differentiated cells such as neurons.

KIF13A mediates trafficking of influenza A virus ribonucleoproteins.

Influenza A is a rapidly evolving virus that is successful in provoking periodic epidemics and occasional pandemics in humans. Viral assembly is complex as the virus incorporates an eight-partite genome of RNA (in the form of viral ribonucleoproteins, vRNPs), and viral genome assembly - with its implications to public health - is not completely understood. It has previously been reported that vRNPs are transported to the cell surface on Rab11-containing vesicles by using microtubules but, so far, no molecular motor has been assigned to the process. Here, we have identified KIF13A, a member of the kinesin-3 family, as the first molecular motor to efficiently transport vRNP-Rab11 vesicles during infection with influenza A. Depletion of KIF13A resulted in reduced viral titers and less accumulation of vRNPs at the cell surface, without interfering with the levels of other viral proteins at sites of viral assembly. In addition, when overexpressed and following two separate approaches to displace vRNP-Rab11 vesicles, KIF13A increased levels of vRNP at the plasma membrane. Together, our results show that KIF13A plays an important role in the transport of influenza A vRNPs, a crucial step for viral assembly.This article has an associated First Person interview with the first author of the paper.

Human gut microbiome and metabolite dynamics under simulated microgravity.

The Artificial Gravity Bed Rest - European Space Agency (AGBRESA) study was the first joint bed rest study by ESA, DLR, and NASA that examined the effect of simulated weightlessness on the human body and assessed the potential benefits of artificial gravity as a countermeasure in an analog of long-duration spaceflight. In this study, we investigated the impact of simulated microgravity on the gut microbiome of 12 participants during a 60-day head-down tilt bed rest at the :envihab facilities. Over 60 days of simulated microgravity resulted in a mild change in the gut microbiome, with distinct microbial patterns and pathway expression in the feces of the countermeasure group compared to the microgravity simulation-only group. Additionally, we found that the countermeasure protocols selectively increased the abundance of beneficial short-chain fatty acids in the gut, such as acetate, butyrate, and propionate. Some physiological signatures also included the modulation of taxa reported to be either beneficial or opportunistic, indicating a mild adaptation in the microbiome network balance. Our results suggest that monitoring the gut microbial catalog along with pathway clustering and metabolite profiling is an informative synergistic strategy to determine health disturbances and the outcome of countermeasure protocols for future space missions.

The future of spaceflight will involve missions beyond the International Space Station or the Moon and astronaut's health will be challenged by a harsh space environment for longer periods. In the last decade, the intestine has gained importance in dictating overall physiology and we explore it as an additional indicator of health during our ground-based bed rest study simulating microgravity for 60 days. Through the analysis of fecal proteins, we compile the catalog of microbes colonizing the gut of the 12 participants along with the implicated biological activity of the proteins and another 9 lipid analytes. We found specific microbes associated with recovery or healthy status in our subjects to be increased during spaceflight countermeasure conditions and inverse observations in subjects subjected to perilous spaceflight simulation. Our approach improves the functional characterization of the gut by the use of noninvasive methodology correlating the microbial composition of human stool samples with physiological status.

Spaceflight Virology: What Do We Know about Viral Threats in the Spaceflight Environment?

Viruses constitute a significant part of the human microbiome, so wherever humans go, viruses are brought with them, even on space missions. In this mini review, we focus on the International Space Station (ISS) as the only current human habitat in space that has a diverse range of viral genera that infect microorganisms from bacteria to eukaryotes. Thus, we have reviewed the literature on the physical conditions of space habitats that have an impact on both virus transmissibility and interaction with their host, which include UV radiation, ionizing radiation, humidity, and microgravity. Also, we briefly comment on the practices used on space missions that reduce virus spread, that is, use of antimicrobial surfaces, spacecraft sterilization practices, and air filtration. Finally, we turn our attention to the health threats that viruses pose to space travel. Overall, even though efforts are taken to ensure safe conditions during human space travel, for example, preflight quarantines of astronauts, we reflect on the potential risks humans might be exposed to and how those risks might be aggravated in extraterrestrial habitats.

The interferon-inducible GTPase MxB promotes capsid disassembly and genome release of herpesviruses.

Host proteins sense viral products and induce defence mechanisms, particularly in immune cells. Using cell-free assays and quantitative mass spectrometry, we determined the interactome of capsid-host protein complexes of herpes simplex virus and identified the large dynamin-like GTPase myxovirus resistance protein B (MxB) as an interferon-inducible protein interacting with capsids. Electron microscopy analyses showed that cytosols containing MxB had the remarkable capability to disassemble the icosahedral capsids of herpes simplex viruses and varicella zoster virus into flat sheets of connected triangular faces. In contrast, capsids remained intact in cytosols with MxB mutants unable to hydrolyse GTP or to dimerize. Our data suggest that MxB senses herpesviral capsids, mediates their disassembly, and thereby restricts the efficiency of nuclear targeting of incoming capsids and/or the assembly of progeny capsids. The resulting premature release of viral genomes from capsids may enhance the activation of DNA sensors, and thereby amplify the innate immune responses.

Acid ceramidase of macrophages traps herpes simplex virus in multivesicular bodies and protects from severe disease.

Macrophages have important protective functions during infection with herpes simplex virus type 1 (HSV-1). However, molecular mechanisms that restrict viral propagation and protect from severe disease are unclear. Here we show that macrophages take up HSV-1 via endocytosis and transport the virions into multivesicular bodies (MVBs). In MVBs, acid ceramidase (aCDase) converts ceramide into sphingosine and increases the formation of sphingosine-rich intraluminal vesicles (ILVs). Once HSV-1 particles reach MVBs, sphingosine-rich ILVs bind to HSV-1 particles, which restricts fusion with the limiting endosomal membrane and prevents cellular infection. Lack of aCDase in macrophage cultures or in Asah1-/- mice results in replication of HSV-1 and Asah1-/- mice die soon after systemic or intravaginal inoculation. The treatment of macrophages with sphingosine enhancing compounds blocks HSV-1 propagation, suggesting a therapeutic potential of this pathway. In conclusion, aCDase loads ILVs with sphingosine, which prevents HSV-1 capsids from penetrating into the cytosol.