The invasion of tobacco mosaic virus RNA induces endoplasmic reticulum stress-related autophagy in HeLa cells

The ability of human cells to defend against viruses originating from distant species has long been ignored. Owing to the pressure of natural evolution and human exploration, some of these viruses may be able to invade human beings. If their ‘fresh’ host had no defences, the viruses could cause a serious pandemic, as seen with HIV, SARS (severe acute respiratory syndrome) and avian influenza virus that originated from chimpanzees, the common palm civet and birds, respectively. It is unknown whether the human immune system could tolerate invasion with a plant virus. To model such an alien virus invasion, we chose TMV (tobacco mosaic virus) and used human epithelial carcinoma cells (HeLa cells) as its ‘fresh’ host. We established a reliable system for transfecting TMV-RNA into HeLa cells and found that TMV-RNA triggered autophagy in HeLa cells as shown by the appearance of autophagic vacuoles, the conversion of LC3-I (light chain protein 3-I) to LC3-II, the up-regulated expression of Beclin1 and the accumulation of TMV protein on autophagosomal membranes. We observed suspected TMV virions in HeLa cells by TEM (transmission electron microscopy). Furthermore, we found that TMV-RNA was translated into CP (coat protein) in the ER (endoplasmic reticulum) and that TMV-positive RNA translocated from the cytoplasm to the nucleolus. Finally, we detected greatly increased expression of GRP78 (78 kDa glucose-regulated protein), a typical marker of ERS (ER stress) and found that the formation of autophagosomes was closely related to the expanded ER membrane. Taken together, our data indicate that HeLa cells used ERS and ERS-related autophagy to defend against TMV-RNA

Simulation and Design of a Prism-Type Ultra-Broadband Microwave Absorber Based on Magnetic Powder/Silica Gel Composites

Materials that absorb electromagnetic waves over an ultra-wide frequency band have great potential for military and civilian applications. In this study, a square-frustum-type metamaterial structure was designed and prepared using CI/silica gel composites and flake-shaped FeNi/silica gel composites as the filling substrate. The structural parameters of the square frustum were simulated and optimized using CST Studio Suite. The results show that the optimal performance was achieved when the base consisted of 50 vol.% CI/silica gel composites and 25 vol.% FeNi/silica gel composites with a cross-pattern distribution, the square frustum consisted of 50 vol.% CI/silica gel composites, and the total thickness, base thickness, base-edge length, and top-edge lengths were 5, 1.8, 2.5, and 1.5 mm, respectively. This arrangement can effectively absorb frequencies between 1.8 and 40 GHz, realizing ultra-broadband absorption. The excellent absorption performance of the absorber is attributed to multiple quarter-wavelength resonances and edge diffraction effects

Tandem catalytic synthesis of benzene from CO2 and H-2

Benzene as an important raw material for production of industrial chemicals is generally synthesized from petroleum and coal tar. Here, we realized the synthesis of benzene from greenhouse CO2 and H-2 with two connected reactors by a tandem catalysis reaction comprising CO2 methanation and CH4 aromatization. The Ni/SiO2 catalyst loaded in the first reactor was used to convert CO2 to CH4, and the formed CH4 was sequentially converted to benzene on the Mo/HZSM-5 catalyst in the second reactor. The benzene formation rate reaches 0.68 mu molg(-1) min(-1) accompanied by a high CO2 conversion of 92%. This concept will provide a new pathway for the direct synthesis of benzene and CO2 utilization

Design and fabrication of ultra-wideband power amplifier based on GaN HEMT

The research of an ultra-broadband power amplifier based on TGF2023-2-02 GaN HEMT which operates in the frequency ranging from 3 GHz to 8 GHz, is presented in this paper. The transistor of GaN HEMT is modeled and a frequency compensation and multi-side impedance matching approach are adopted for broadband impedance matching of amplifier. And a fan shaped micro strip line is implemented in the input matching network to achieve the wideband higher gain features. The measured results show that the amplifier module provided more than 37 dBm output power with minimum small signal gain of 9.8 dB over 3–8 GHz. The saturated output power is 38.3 dBm under DC bias of Vds = 28 V, Vgs = ?2.75 V at the frequency of 5 GHz

Mechanical Anisotropy Induced by Strain Path Change for AZ31 Mg Alloy Sheet

The variation of strain paths induces anisotropy during practical sheet forming processes, which is very important for the subsequent processing technology of anisotropic Mg alloys. In this study, two-step loading tests (tension-tension) were performed to clarify the effect of strain path changes on the evolution of anisotropy on rolled AZ31 sheet. Specimens were preloaded with tension along the rolling direction (RD) with 9% of prestrain. Then, second tension was conducted along 0°, 30°, 45°, 60° and 90° from the RD. It was found that yield strength during the second loading increased along the same direction compared to uniaxial tension without prestraining. For the second loading, the yield strength and flow stress decreased with the increase of the angle from the RD. It was found that the strain path change resulted in stronger anisotropy than that induced by texture. Moreover, it was found that the main deformation modes were basal and prismatic slips during the second loading based on visco-plastic self-consistent (VPSC) modeling. The relative activities of basal and prismatic slips were affected by the second loading direction due to texture evolution. The mechanical anisotropy induced by strain path changes was ascribed to the coupling of the heterogeneous distribution of dislocations and texture evolution induced by prestraining

Coxsackievirus A16 elicits incomplete autophagy involving the mTOR and ERK pathways.

Autophagy is an important homeostatic process for the degradation of cytosolic proteins and organelles and has been reported to play an important role in cellular responses to pathogens and virus replication. However, the role of autophagy in Coxsackievirus A16 (CA16) infection and pathogenesis remains unknown. Here, we demonstrated that CA16 infection enhanced autophagosome formation, resulting in increased extracellular virus production. Moreover, expression of CA16 nonstructural proteins 2C and 3C was sufficient to trigger autophagosome accumulation by blocking the fusion of autophagosomes with lysosomes. Interestingly, we found that Immunity-related GTPase family M (IRGM) was crucial for the activation of CA16 infection-induced autophagy; in turn, reducing IRGM expression suppressed autophagy. Expression of viral protein 2C enhanced IRGM promoter activation, thereby increasing IRGM expression and inducing autophagy. CA16 infection inhibited Akt/mTOR signaling and activated extracellular signal-regulated kinase (ERK) signaling, both of which are necessary for autophagy induction. In summary, CA16 can use autophagy to enhance its own replication. These results raise the possibility of targeting the autophagic pathway for the treatment of hand, foot, and mouth disease (HFMD)

Hepatitis B virus hijacks TSG101 to facilitate egress via multiple vesicle bodies.

Hepatitis B virus (HBV) chronically infects 296 million individuals and there is no cure. As an important step of viral life cycle, the mechanisms of HBV egress remain poorly elucidated. With proteomic approach to identify capsid protein (HBc) associated host factors and siRNA screen, we uncovered tumor susceptibility gene 101 (TSG101). Knockdown of TSG101 in HBV-producing cells, HBV-infected cells and HBV transgenic mice suppressed HBV release. Co-immunoprecipitation and site mutagenesis revealed that VFND motif in TSG101 and Lys-96 ubiquitination in HBc were essential for TSG101-HBc interaction. In vitro ubiquitination experiment demonstrated that UbcH6 and NEDD4 were potential E2 ubiquitin-conjugating enzyme and E3 ligase that catalyzed HBc ubiquitination, respectively. PPAY motif in HBc and Cys-867 in NEDD4 were required for HBc ubiquitination, TSG101-HBc interaction and HBV egress. Transmission electron microscopy confirmed that TSG101 or NEDD4 knockdown reduces HBV particles count in multivesicular bodies (MVBs). Our work indicates that TSG101 recognition for NEDD4 ubiquitylated HBc is critical for MVBs mediated HBV egress

Hepatitis B virus RNAs co-opt ELAVL1 for stabilization and CRM1-dependent nuclear export.

Hepatitis B virus (HBV) chronically infects 296 million people worldwide, posing a major global health threat. Export of HBV RNAs from the nucleus to the cytoplasm is indispensable for viral protein translation and genome replication, however the mechanisms regulating this critical process remain largely elusive. Here, we identify a key host factor embryonic lethal, abnormal vision, Drosophila-like 1 (ELAVL1) that binds HBV RNAs and controls their nuclear export. Using an unbiased quantitative proteomics screen, we demonstrate direct binding of ELAVL1 to the HBV pregenomic RNA (pgRNA). ELAVL1 knockdown inhibits HBV RNAs posttranscriptional regulation and suppresses viral replication. Further mechanistic studies reveal ELAVL1 recruits the nuclear export receptor CRM1 through ANP32A and ANP32B to transport HBV RNAs to the cytoplasm via specific AU-rich elements, which can be targeted by a compound CMLD-2. Moreover, ELAVL1 protects HBV RNAs from DIS3+RRP6+ RNA exosome mediated nuclear RNA degradation. Notably, we find HBV core protein is dispensable for HBV RNA-CRM1 interaction and nuclear export. Our results unveil ELAVL1 as a crucial host factor that regulates HBV RNAs stability and trafficking. By orchestrating viral RNA nuclear export, ELAVL1 is indispensable for the HBV life cycle. Our study highlights a virus-host interaction that may be exploited as a new therapeutic target against chronic hepatitis B

Measurement of the autophagic flux in HeLa cells infected with CA16.

<p>(A, B) Western blotting of cells with autophagy inhibited by CQ. HeLa cells were pretreated with CQ for 4 h, followed by infection with CA16 at an MOI of 2. After 1 h of virus absorption at 37°C, the cells were further cultured in maintenance medium in the absence or presence of CQ. At 24 h after infection with CA16, the cells were subjected to Western blotting using anti-LC3B and Vp1 antibodies. (C) Western blotting analysis of p62 protein expression in HeLa cells infected with CA16. Cells were infected with CA16 at an MOI of 1. After 1 h of virus absorption at 37°C, the cells were further cultured in maintenance medium. Cells were harvested at the indicated time points and detected with anti-p62 antibody compared to uninfected control cells. (D) HeLa cells transfected with ptfLC3 were infected with CA16 (MOI = 1) or treated with CQ or HBSS. The cells were collected, fixed, and visualized at 24 h postinfection. The graph shows the quantification of autophagosomes by calculating the average number of dots in 20 cells. Scale bar, 10μm. (E) Western blotting of autophagy-related proteins in cells transfected with the indicated shRNA and determination of CA16 replication in these transfected cells. HeLa cells were transfected with either specific shRNA targeting Beclin 1, Atg5 or scrambled shRNA. At 48 h after transfection, cells were infected with CA16 at an MOI of 2. Samples were collected at 12 h after infection with CA16 and detected with anti-Beclin 1,-Atg5 and-Vp1 antibodies. β-Actin was used as a protein loading control. The extracellular virus yields were determined at 48 hpi and expressed as lgTCID50/ml. Data are presented as the means from three independent experiments. Significance was analyzed with a two-tailed Student’s <i>t</i> test. *<i>P</i>< 0.05, **<i>P</i>< 0.01, ***<i>P</i>< 0.001. (F) Determination of CA16 replication in HeLa cells treated with CQ. HeLa cells were pretreated with CQ for 4 h followed by infection with CA16 at an MOI of 2. The extracellular virus yields were determined at 12 hpi and expressed as lgTCID50/ml. Data are presented as the means from three independent experiments. Significance was analyzed with a two-tailed Student’s <i>t</i> test. *<i>P</i>< 0.05.</p