During the emergence of novel coronavirus 2019 (nCoV) outbreak in Wuhan city, China at the end of 2019, there was movement of many airline travelers between Wuhan and Japan, suggesting that the ...Japanese population was at high risk of infection by the virus. Hence, we urgently developed diagnostic systems for detection of 2019 nCoV. Two nested RT-PCR and two real-time RT-PCR assays were adapted for use in Japan. As of February 8, 2020, these assays have successfully detected 25 positive cases of infection in Japan.
Rubella virus (RuV) causes a systemic infection, and transplacental fetal infection causes congenital rubella syndrome. In this study, we showed that treatment of cells with sphingomyelinase ...inhibited RuV infection. Assays using inhibitors of serine palmitoyl transferase and ceramide transport protein demonstrated the contribution of sphingomyelin (SM) to RuV infection. Compelling evidence for direct binding of RuV to lipid membranes at neutral pH was obtained using liposome coflotation assays. The absence of either SM or cholesterol (Chol) abrogated the RuV-liposome interaction. SM and Chol (SM/Chol) were also critical for RuV binding to erythrocytes and lymphoid cells. Removal of Ca
from the assay buffer or mutation of RuV envelope E1 protein Ca
-binding sites abrogated RuV binding to liposomes, erythrocytes, and lymphoid cells. However, RuV bound to various nonlymphoid adherent cell lines independently of extracellular Ca
or SM/Chol. Even in these adherent cell lines, both the E1 protein Ca
-binding sites and cellular SM/Chol were essential for the early stage of RuV infection, possibly affecting envelope-membrane fusion in acidic compartments. Myelin oligodendrocyte glycoprotein (MOG) has recently been identified as a cellular receptor for RuV. However, RuV bound to MOG-negative cells in a Ca
-independent manner. Collectively, our data demonstrate that RuV has two distinct binding mechanisms: one is Ca
dependent and the other is Ca
independent. Ca
-dependent binding observed in lymphoid cells occurs by the direct interaction between E1 protein fusion loops and SM/Chol-enriched membranes. Clarification of the mechanism of Ca
-independent RuV binding is an important next step in understanding the pathology of RuV infection.
Rubella has a significant impact on public health as infection during early pregnancy can result in babies being born with congenital rubella syndrome. Even though effective rubella vaccines are available, rubella outbreaks still occur in many countries. We studied the entry mechanism of rubella virus (RuV) and found that RuV binds directly to the host plasma membrane in the presence of Ca
at neutral pH. This Ca
-dependent binding is specifically directed to membranes enriched in sphingomyelin and cholesterol and is critical for RuV infection. Importantly, RuV also binds to many cell lines in a Ca
-independent manner. An unidentified RuV receptor(s) is involved in this Ca
-independent binding. We believe that the data presented here may aid the development of the first anti-RuV drug.
Two viral nonstructural proteins, p150 and p90, are expressed in rubella virus (RUBV)-infected cells and mediate viral genome replication, presumably using various host machineries. Molecular ...chaperones are critical host factors for the maintenance of cellular proteostasis, and certain viral proteins use this chaperone system. The RUBV p150 and p90 proteins are generated from a precursor polyprotein, p200, via processing by the protease activity of its p150 region. This processing is essential for RUBV genome replication. Here we show that heat shock protein 90 (HSP90), a molecular chaperone, is an important host factor for RUBV genome replication. The treatment of RUBV-infected cells with the HSP90 inhibitors 17-allylamino-17-desmethoxygeldanamycin (17-AAG) and ganetespib suppressed RUBV genome replication. HSP90α physically interacted with p150, but not p90. Further analyses into the mechanism of action of the HSP90 inhibitors revealed that HSP90 activity contributes to p150 functional integrity and promotes p200 processing. Collectively, our data demonstrate that RUBV p150 is a client of the HSP90 molecular chaperone and that HSP90 functions as a key host factor for RUBV replication.
Accumulating evidence indicates that RNA viruses use numerous host factors during replication of their genomes. However, the host factors involved in rubella virus (RUBV) genome replication are largely unknown. In this study, we demonstrate that the HSP90 molecular chaperone is needed for the efficient replication of the RUBV genome. Further, we reveal that HSP90 interacts with RUBV nonstructural protein p150 and its precursor polyprotein, p200. HSP90 contributes to the stability of p150 and the processing of p200 via its protease domain in the p150 region. We conclude that the cellular molecular chaperone HSP90 is a key host factor for functional maturation of nonstructural proteins for RUBV genome replication. These findings provide novel insight into this host-virus interaction.
Hepatitis E virus (HEV) is a causative agent of acute hepatitis. The crystal structure of HEV-like particles (HEV-LP) consisting of capsid protein was determined at 3.5-Å resolution. The capsid ...protein exhibited a quite different folding at the protruding and middle domains from the members of the families of Caliciviridae and Tombusviridae, while the shell domain shared the common folding. Tyr-288 at the 5-fold axis plays key roles in the assembly of HEV-LP, and aromatic amino acid residues are well conserved among the structurally related viruses. Mutational analyses indicated that the protruding domain is involved in the binding to the cells susceptive to HEV infection and has some neutralization epitopes. These structural and biological findings are important for understanding the molecular mechanisms of assembly and entry of HEV and also provide clues in the development of preventive and prophylactic measures for hepatitis E.
A special symposium titled ‘Towards Elimination of Rubella’ was held at the 23rd Annual Meeting of the Japanese Society for Vaccinology in Tokyo, Japan on November 30, 2019. Representatives from the ...Japanese national government, a local government, and a private company presented their efforts to control rubella, and the group discussed the challenges of eliminating rubella in Japan.
Measles outbreaks have occurred in some countries despite supplementary immunization activities (SIA) using measles-containing vaccine with high vaccination coverage. We conducted a cross-sectional ...seroprevalence survey to estimate population immunity in Lao People's Democratic Republic where repeated mass immunization has failed to eliminate measles.
In this nationwide multistage cluster sampling survey conducted in 2014 based on probability proportionate to size sampling, blood samples were collected from 2,135 children and adults living in 52 randomly selected villages. Anti-measles and anti-rubella IgG were measured, and IgG prevalence was calculated. We applied mathematical modelling to estimate the number of cases of congenital rubella syndrome (CRS) in 2013 that were averted by the 2011 SIA. A stability testing was applied to the MR vaccine at 4°C, 25°C, and 35°C to examine stability differences between measles and rubella vaccine components. Measles IgG prevalence was significantly lower in the target age groups (5-21 years) of the 2011 SIA using a combination vaccine for measles and rubella vaccine (MR vaccine) than in young adults (22-39 years) (86.8% 95% CI: 83.0-90.6 vs. 99.0% 98.3-99.8; p<0.001), whereas rubella IgG prevalence was significantly higher (88.2% 84.5-91.8 vs. 74.6% 70.7-78.5; p<0.001). In the SIA target age groups, prevalence of measles IgG, but not rubella IgG, increased with age. CRS cases prevented in 2013 ranged from 16 0-50 to 92 32-180 if the force of infection had remained unchanged or had been reduced by 75%, respectively. In freeze-dried conditions, the measles vaccine component was more heat sensitive than the rubella component.
Inconsistent IgG prevalence between measles and rubella in Lao PDR can be partly explained by different stability of the measles and rubella vaccine components under heat exposure. Suboptimal vaccine handling may cause insufficient immunogenicity for measles, which subsequently leads to an outbreak despite high SIA coverage, while direct evidence is lacking. Temperature monitoring of the vaccine should be conducted.
Retroperitoneal sarcoma (RPS) is a rare disease. RPS invading the abdominal aorta is exceedingly rare and has a poor prognosis. There have been scattered cases of RPS treated with combined abdominal ...aortic replacement. However, the average survival time for these cases was only 8 months, with a 2-year survival rate of 21%, indicating a poor prognosis. In this case study, a 44-year-old man presented to our hospital complaining of abdominal pain. Multiple imaging findings suggested a retroperitoneal mass that was diagnosed as a malignant tumor. The patient underwent tumor resection with abdominal aortic replacement due to an RPS tumor invading the abdominal aorta. The histopathological grade was determined to be grade 3, the most malignant grade tumor, according to the Fédération Nationale des Centres de Lutte Contre le Cancer grading system. Postoperative chemotherapy with doxorubicin and ifosfamide was administered for five cycles. The patient has been alive for over 8 years after the operation without any recurrence. This case presents a long-term survival of RPS requiring abdominal aortic replacement.
The lipid composition of the host cell membrane is one of the key determinants of the entry of enveloped viruses into cells. To elucidate the detailed mechanisms behind the cell entry of rubella ...virus (RuV), one of the enveloped viruses, we searched for host factors involved in such entry by using CRISPR/Cas9 genome-wide knockout screening, and we found sphingomyelin synthase 1 (SMS1), encoded by the
gene, as a candidate. RuV growth was strictly suppressed in
-knockout cells and was completely recovered by the overexpression of enzymatically active SMS1 and partially recovered by that of SMS2, another member of the SMS family, but not by that of enzymatically inactive SMS1. An entry assay using pseudotyped vesicular stomatitis virus possessing RuV envelope proteins revealed that sphingomyelin generated by SMSs is crucial for at least RuV entry. In
-knockout cells, lipid mixing between the RuV envelope membrane and the membrane of host cells occurred, but entry of the RuV genome from the viral particles into the cytoplasm was strongly inhibited. This indicates that sphingomyelin produced by SMSs is essential for the formation of membrane pores after hemifusion occurs during RuV entry.
Infection with rubella virus during pregnancy causes congenital rubella syndrome in infants. Despite its importance in public health, the detailed mechanisms of rubella virus cell entry have only recently become somewhat clearer. The E1 protein of rubella virus is classified as a class II fusion protein based on its structural similarity, but it has the unique feature that its activity is dependent on calcium ion binding in the fusion loops. In this study, we found another unique feature, as cellular sphingomyelin plays a critical role in the penetration of the nucleocapsid into the cytoplasm after hemifusion by rubella virus. This provides important insight into the entry mechanism of rubella virus. This study also presents a model of hemifusion arrest during cell entry by an intact virus, providing a useful tool for analyzing membrane fusion, a biologically important phenomenon.
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