A Tunable Brake for HECT Ubiquitin Ligases.

The HECT E3 ligases ubiquitinate numerous transcription factors and signaling molecules, and their activity must be tightly controlled to prevent cancer, immune disorders, and other diseases. In this study, we have found unexpectedly that peptide linkers tethering WW domains in several HECT family members are key regulatory elements of their catalytic activities. Biochemical, structural, and cellular analyses have revealed that the linkers can lock the HECT domain in an inactive conformation and block the proposed allosteric ubiquitin binding site. Such linker-mediated autoinhibition of the HECT domain can be relieved by linker post-translational modifications, but complete removal of the brake can induce hyperactive autoubiquitination and E3 self destruction. These results clarify the mechanisms of several HECT protein cancer associated mutations and provide a new framework for understanding how HECT ubiquitin ligases must be finely tuned to ensure normal cellular behavior.

Global Dynamics of Porcine Enteric Coronavirus PEDV Epidemiology, Evolution, and Transmission.

With a possible origin from bats, the alphacoronavirus Porcine epidemic diarrhea virus (PEDV) causes significant hazards and widespread epidemics in the swine population. However, the ecology, evolution, and spread of PEDV are still unclear. Here, from 149,869 fecal and intestinal tissue samples of pigs collected in an 11-year survey, we identified PEDV as the most dominant virus in diarrheal animals. Global whole genomic and evolutionary analyses of 672 PEDV strains revealed the fast-evolving PEDV genotype 2 (G2) strains as the main epidemic viruses worldwide, which seems to correlate with the use of G2-targeting vaccines. The evolving pattern of the G2 viruses presents geographic bias as they evolve tachytely in South Korea but undergo the highest recombination in China. Therefore, we clustered six PEDV haplotypes in China, whereas South Korea held five haplotypes, including a unique haplotype G. In addition, an assessment of the spatiotemporal spread route of PEDV indicates Germany and Japan as the primary hubs for PEDV dissemination in Europe and Asia, respectively. Overall, our findings provide novel insights into the epidemiology, evolution, and transmission of PEDV, and thus may lay a foundation for the prevention and control of PEDV and other coronaviruses.

A Novel C Doped MoS2 /CoP/MoO2 Ternary Heterostructure Nanoflower for Hydrogen Evolution Reaction at Wide pH Range and Efficient Overall Water Splitting in Alkaline Media.

The development of low-cost and high-efficiency bifunctional catalysts is still a challenge for hydrogen production through overall water splitting. This paper reports the in-situ synthesis of C-doped MoS2 /CoP/MoO2 using bacterial cellulose (BC) as the reducing agent and the source of C and using BC (MoS2 /Co1.2 MoO4.2 ⋅ 1.2H2 O/BC) as the template. Heterogeneous structure for hydrogen evolution reaction (HER) and alkaline water electrolysis in a wide pH range. Due to the large number of defect sites caused by C doping and the synergy between these three active components (MoS2 , CoP and MoO2 ), the HER and oxygen evolution reaction (OER) activities of the catalyst have been greatly improved. Therefore, during HER, a small initial overpotential (27 mV) was achieved in 1.0 M KOH. In 0.5 M H2 SO4 , 0.1 M PBS and 1.0 M KOH, the current density reached 10 mA cm-2 at overpotentials of 123.4, 150, and 139 mV, respectively. For OER, an overpotential of 268 mV was required to achieve 10 mA cm-2 . The alkaline two-electrode device composed of C doped MoS2 /CoP/MoO2 delivers 10 mA cm-2 at a low potential of 1.51 V and can be easily driven by a single AA battery. This work provides a new design strategy of C doped ternary heterostructures for electrocatalysis and related energy applications.

Circular RNA hsa_circ_0018189 drives non-small cell lung cancer growth by sequestering miR-656-3p and enhancing xCT expression.

BACKGROUND: Non-small cell lung cancer (NSCLC) is one of the cancers with a high mortality rate. CircRNAs have emerged as an important regulatory factor in tumorigenesis in recent years. However, the detailed regulatory mechanism of a circular RNA cullin 2 (hsa_circ_0018189; hsa_circ_0018189) is still unclear in NSCLC. METHODS: RNA levels of hsa_circ_0018189, microRNA (miR)-656-3p, and Solute carrier family seven member 11 (SLC7A11, xCT) were analyzed by real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR), and protein level was assessed by Western blot and immunohistochemical assay. Enzyme-linked immunosorbent assay was conducted to detect cell glutamine metabolism. Effects of hsa_circ_0018189 on cell proliferation, apoptosis, migration, and invasion were analyzed by corresponding assays. Luciferase reporter assay and RNA-immunoprecipitation assay confirmed the target relationship between miR-656-3p and hsa_circ_0018189 or xCT. The in vivo function of hsa_circ_0018189 was verified by xenograft mouse models. RESULTS: Hsa_circ_0018189 abundance was overexpressed in NSCLC cells and samples. Deficiency of hsa_circ_0018189 lowered NSCLC cell proliferative, migrating, invading, and glutamine metabolism capacities, and hsa_circ_0018189 silencing inhibited the growth of tumors in vivo. Hsa_circ_0018189 could up-regulate xCT by sponging miR-656-3p. And miR-656-3p downregulation or xCT overexpression partly overturned hsa_circ_0018189 knockdown or miR-656-3p mimic-mediated repression of NSCLC cell malignancy. CONCLUSION: Hsa_circ_0018189 drove NSCLC growth by interacting with miR-656-3p and upregulating xCT.

[Application of Statistical Process Control in Evaluation of Performance for Beam-matched Medical Electron Linacs].

Clinically, beam matching can greatly improve the flexibility and efficiency of treating patients between different medical electron linacs. However, in addition to the regular quality assurance (QA) test of the machine performance of linacs, there is still a lack of comprehensive evaluation of the clinical radiotherapy performance of beam-matched linacs. In this paper, the performance of volumetric modulated arc therapy (VMAT) between three closely matched linacs was evaluated by statistical process control (SPC) technology. It was found that the average and median γ passing rates of the VMAT QA processes of the three linacs had little difference, but the process capability levels were at three different levels. The results show that SPC technology can effectively evaluate the performance of beam matching for medical electron linacs, improve the patient-specific VMAT QA processes, and guide clinical decision-making.

[Application of Virtual Monochromatic Images Reconstructed by Dual-energy Computed Tomography in Radiotherapy Treatment Planning System].

Virtual monochromatic images (VMI) that reconstructed on dual-energy computed tomography (DECT) have further application prospects in radiotherapy, and there is still a lack of clinical dose verification. In this study, GE Revolution CT scanner was used to perform conventional imaging and gemstone spectral imaging on the simulated head and body phantom. The CT images were imported to radiotherapy treatment planning system (TPS), and the same treatment plans were transplanted to compare the CT value and the dose distribution. The results show that the VMI can be imported into TPS for CT value-relative electron density conversion and dose calculation. Compared to conventional images, the VMI varies from 70 to 140 keV, has little difference in dose distribution of 6 MV photon treatment plan.

[Preliminary study on monitoring patient-specific volumetric modulated arc therapy quality assurance process with statistical process control methodology on the basis of TG-218 report].

Patient-specific volumetric modulated arc therapy (VMAT) quality assurance (QA) process is an important component of the implementation process of clinical radiotherapy. The tolerance limit and action limit of discrepancies between the calculated dose and the delivered radiation dose are the key parts of the VMAT QA processes as recognized by the AAPM TG-218 report, however, there is no unified standard for these two values among radiotherapy centers. In this study, based on the operational recommendations given in the AAPM TG-218 report, treatment site-specific tolerance limits and action limits of gamma pass rate in VMAT QA processes when using ArcCHECK for dose verification were established by statistical process control (SPC) methodology. The tolerance limit and action limit were calculated based on the first 25 in-control VMAT QA for each site. The individual control charts were drawn to continuously monitor the VMAT QA process with 287 VMAT plans and analyze the causes of VMAT QA out of control. The tolerance limits for brain, head and neck, abdomen and pelvic VMAT QA processes were 94.56%, 94.68%, 94.34%, and 92.97%, respectively, and the action limits were 93.82%, 92.54%, 93.23%, and 90.29%, respectively. Except for pelvic, the tolerance limits for the brain, head and neck, and abdomen were close to the universal tolerance limit of TG-218 (95%), and the action limits for all sites were higher than the universal action limit of TG-218 (90%). The out-of-control VMAT QAs were detected by the individual control chart, including one case of head and neck, two of the abdomen and two of the pelvic site. Four of them were affected by the setup error, and one was affected by the calibration of ArcCHECK. The results show that the SPC methodology can effectively monitor the IMRT/VMAT QA processes. Setting treatment site-specific tolerance limits is helpful to investigate the cause of out-of-control VMAT QA.

The mechanism of OTUB1-mediated inhibition of ubiquitination.

Histones are ubiquitinated in response to DNA double-strand breaks (DSB), promoting recruitment of repair proteins to chromatin. UBC13 (also known as UBE2N) is a ubiquitin-conjugating enzyme (E2) that heterodimerizes with UEV1A (also known as UBE2V1) and synthesizes K63-linked polyubiquitin (K63Ub) chains at DSB sites in concert with the ubiquitin ligase (E3), RNF168 (ref. 3). K63Ub synthesis is regulated in a non-canonical manner by the deubiquitinating enzyme, OTUB1 (OTU domain-containing ubiquitin aldehyde-binding protein 1), which binds preferentially to the UBC13∼Ub thiolester. Residues amino-terminal to the OTU domain, which had been implicated in ubiquitin binding, are required for binding to UBC13∼Ub and inhibition of K63Ub synthesis. Here we describe structural and biochemical studies elucidating how OTUB1 inhibits UBC13 and other E2 enzymes. We unexpectedly find that OTUB1 binding to UBC13∼Ub is allosterically regulated by free ubiquitin, which binds to a second site in OTUB1 and increases its affinity for UBC13∼Ub, while at the same time disrupting interactions with UEV1A in a manner that depends on the OTUB1 N terminus. Crystal structures of an OTUB1-UBC13 complex and of OTUB1 bound to ubiquitin aldehyde and a chemical UBC13∼Ub conjugate show that binding of free ubiquitin to OTUB1 triggers conformational changes in the OTU domain and formation of a ubiquitin-binding helix in the N terminus, thus promoting binding of the conjugated donor ubiquitin in UBC13∼Ub to OTUB1. The donor ubiquitin thus cannot interact with the E2 enzyme, which has been shown to be important for ubiquitin transfer. The N-terminal helix of OTUB1 is positioned to interfere with UEV1A binding to UBC13, as well as with attack on the thiolester by an acceptor ubiquitin, thereby inhibiting K63Ub synthesis. OTUB1 binding also occludes the RING E3 binding site on UBC13, thus providing a further component of inhibition. The general features of the inhibition mechanism explain how OTUB1 inhibits other E2 enzymes in a non-catalytic manner.

Engineering the Band Gap States of the Rutile TiO2 (110) Surface by Modulating the Active Heteroatom.

Introducing band gap states to TiO2 photocatalysts is an efficient strategy for expanding the range of accessible energy available in the solar spectrum. However, few approaches are able to introduce band gap states and improve photocatalytic performance simultaneously. Introducing band gap states by creating surface disorder can incapacitate reactivity where unambiguous adsorption sites are a prerequisite. An alternative method for introduction of band gap states is demonstrated in which selected heteroatoms are implanted at preferred surface sites. Theoretical prediction and experimental verification reveal that the implanted heteroatoms not only introduce band gap states without creating surface disorder, but also function as active sites for the CrVI reduction reaction. This promising approach may be applicable to the surfaces of other solar harvesting materials where engineered band gap states could be used to tune photophysical and -catalytic properties.

Enzymatic Analysis of PTEN Ubiquitylation by WWP2 and NEDD4-1 E3 Ligases.

PTEN is a lipid phosphatase that converts phosphatidylinositol 3,4,5-phosphate (PIP3) to phosphatidylinositol 4,5-phosphate (PIP2) and plays a critical role in the regulation of tumor growth. PTEN is subject to regulation by a variety of post-translational modifications, including phosphorylation on a C-terminal cluster of four Ser/Thr residues (380, 382, 383, and 385) and ubiquitylation by various E3 ligases, including NEDD4-1 and WWP2. It has previously been shown that C-terminal phosphorylation of PTEN can increase its cellular half-life. Using in vitro ubiquitin transfer assays, we show that WWP2 is more active than NEDD4-1 in ubiquitylating unphosphorylated PTEN. The mapping of ubiquitylation sites in PTEN by mass spectrometry showed that both NEDD4-1 and WWP2 can target a broad range of Lys residues in PTEN, although NEDD4-1 versus WWP2 showed a stronger preference for ubiquitylating PTEN's C2 domain. Whereas tetraphosphorylation of PTEN did not significantly affect its ubiquitylation by NEDD4-1, it inhibited PTEN ubiquitylation by WWP2. Single-turnover and pull-down experiments suggested that tetraphosphorylation of PTEN appears to weaken its interaction with WWP2. These studies reveal how the PTEN E3 ligases WWP2 and NEDD4-1 exhibit distinctive properties in Lys selectivity and sensitivity to PTEN phosphorylation. Our findings also provide a molecular mechanism for the connection between PTEN Ser/Thr phosphorylation and PTEN's cellular stability.

Identification and pathogenicity of a variant porcine epidemic diarrhea virus field strain with reduced virulence.

BACKGROUND: Since 2010, a variant Porcine epidemic diarrhea virus (PEDV), which causes an acute, highly contagious, and devastating viral enteric disease with a high mortality rate in suckling pigs, broke out in China and spread rapidly to neighboring countries, even to the North America. This virus gradually became the main subtype of PEDV worldwide. However, there were no reports of mild pathogenicity of a variant porcine epidemic diarrhea virus in China. FINDINGS: In 2013, a PEDV-positive sample from a sow with very mild clinical sign was used to inoculate in Vero cells to isolate the virus. This PEDV field strain, designated FL2013 strain, was successfully propagated and genetically characterized. The phylogenetic trees based upon either the complete genome or S gene showed that the FL2013 strain belongs to the genogroup G2b. The S gene of FL2013 has a 7-aa deletion (FEKVHVQ) in the C-terminus comparison with the other G2 PEDV sequences. Further comparative pathology study indicated that the FL2013 strain had reduced virulence to newborn piglets. CONCLUSIONS: A novel variant PEDV strain FL2013 with reduced virulence, as determined by the pathological study, was identified from east China. This strain is closely related to the genogroup-2 PEDV strains prevalent in the U.S. and China currently, but had a short deletion at the 3'-end of the spike gene.

Sequence and phylogenetic analysis of surface protein genes of emerging H9N2 influenza viruses isolated from poultry in two geographical regions of China.

Subtype H9N2 avian influenza viruses (AIVs) circulating in China have aroused increasing concerns for their impact on poultry and risk to public health. The present study was an attempt to elucidate the phylogenetic relationship of H9N2 AIVs in two geographically distinct regions of China where vaccination is routinely practiced. A total of 18 emerging H9N2 isolates were identified and genetically characterized. Phylogenetic analysis of hemagglutinin (HA) and neuraminidase (NA) genes confirmed that the isolates belonged to the Y280 lineage. Based on the HA genes, the isolates were subdivided into two subgroups. The viruses from Zhejiang Province were clustered together in Group I, while the isolates from Guangdong Province were clustered together in Group II. Antigenic characterization showed that the tested viruses were antigenically different when compared to the current used vaccine strain. It was notable that 14 out of total 18 isolates had an amino acid exchange (Q→L) at position 216 (226 by H3 Numbering) in the receptor-binding site, which indicated that the virus had potential affinity of binding to human like receptor. These results suggest that the emerging viruses have potential risk to public health than previously thought. Therefore, continuous surveillance studies of H9N2 influenza virus are very important to the prognosis and control of future influenza pandemics.

Haspin kinase binds to a nucleosomal DNA supergroove.

Phosphorylation of histone H3 threonine 3 (H3T3) by Haspin recruits the chromosomal passenger complex to the inner centromere and ensures proper cell cycle progression through mitosis. The mechanism by which Haspin binds to nucleosomes to phosphorylate H3T3 is not known. We report here cryo-EM structures of the Haspin kinase domain bound to a nucleosome. In contrast with previous structures of histone-modifying enzymes, Haspin solely contacts the nucleosomal DNA, inserting into a supergroove formed by apposing major grooves of two DNA gyres. This unique binding mode provides a plausible mechanism by which Haspin can bind to nucleosomes in a condensed chromatin environment to phosphorylate H3T3. We identify key basic residues in the Haspin kinase domain that are essential for phosphorylation of nucleosomal histone H3 and binding to mitotic chromatin. Our structure is the first of a kinase domain bound to a nucleosome and is the first example of a histone-modifying enzyme that binds to nucleosomes solely through DNA contacts.

Promising therapeutic targets for ischemic stroke identified from plasma and cerebrospinal fluid proteomes: a multicenter Mendelian randomization study.

BACKGROUND: Ischemic stroke (IS) is more common every year, the condition is serious, and have a poor prognosis. New, efficient, and safe therapeutic targets are desperately needed as early treatment especially prevention and reperfusion is the key to lowering the occurrence of poorer prognosis. Generally circulating proteins are attractive therapeutic targets, this study aims to identify potential pharmacological targets among plasma and cerebrospinal fluid (CSF) proteins for the prevention and treatment of IS using a multicenter Mendelian randomization (MR) approach. METHODS: First, the genetic instruments of 734 plasma and 151 CSF proteins were assessed for causative connections with IS from MEGASTROKE consortium by MR to identify prospective therapeutic targets. Then, for additional validation, plasma proteins from the deCODE consortium and the Fenland consortium, as well as IS GWAS data from the FinnGen cohort, the ISGC consortium and UK biobank, were employed. A thorough evaluation of the aforementioned possible pharmacological targets was carried out using meta-analysis. The robustness of MR results was then confirmed through sensitivity analysis using several techniques, such as bidirectional MR analysis, Steiger filtering, and Bayesian colocalization. Finally, methods like Protein-Protein Interaction (PPI) Networking were utilized to investigate the relationship between putative drug targets and therapeutic agents. RESULTS: The authors discovered three proteins that may function as promising therapeutic targets for IS and meet the Bonferroni correction ( P <0.05/885=5.65×10 -5 ). Prekallikrein (OR=0.41, 95% CI: 0.27-0.63, P =3.61×10 -5 ), a protein found in CSF, has a 10-fold protective impact in IS, while the plasma proteins SWAP70 (OR=0.85, 95% CI: 0.80-0.91, P =1.64×10 -6 ) and MMP-12 (OR=0.92, 95% CI: 0.89-0.95, P =4.49×10 -6 ) of each SD play a protective role in IS. Prekallikrein, MMP-12, SWAP70 was replicated in the FinnGen cohort and ISGC database. MMP-12 (OR=0.93, 95% CI: 0.91-0.94, P <0.001), SWAP70 (OR=0.92, 95% CI: 0.90-0.94, P <0.001), and prekallikrein (OR=0.53, 95% CI: 0.33-0.72, P <0.001) may all be viable targets for IS, according to the combined meta-analysis results. Additionally, no evidence of reverse causality was identified, and Bayesian colocalization revealed MMP-12 (PPH 4 =0.995), SWAP70 (PPH 4 =0.987), and prekallikrein (PPH 4 =0.894) shared the same variant with IS, supporting the robustness of the aforementioned causation. Prekallikrein and MMP-12 were associated with the target protein of the current treatment of IS. Among them, Lanadelumab, a new drug whose target protein is a prekallikrein, may be a promising new drug for the treatment of IS. CONCLUSION: The prekallikrein, MMP-12, and SWAP70 are causally associated with the risk of IS. Moreover, MMP-12 and prekallikrein may be treated as promising therapeutic targets for medical intervention of IS.

Complete genome sequence of a variant porcine epidemic diarrhea virus strain isolated in China.

Since October 2010, an outbreak of porcine epidemic diarrhea (PED) has been observed in some provinces of China. Here we report the complete genome sequence of porcine epidemic diarrhea virus (PEDV) strain LC, which was recently isolated from sucking piglets that suffered from severe watery diarrhea in Guangdong. It will help in understanding the epidemiological and molecular characteristics of PEDV in China.

Complete genome sequence of porcine circovirus 2d strain GDYX.

Porcine circovirus type 2 (PCV2) is the etiologic agent of porcine circovirus-associated disease, and it is mainly divided into five genotypes. Here, we report the complete genome sequence of PCV2 strain GDYX, which belongs to PCV2d and has a unique amino acid variation at position 169 (S to G).

Complete genome sequence of a porcine orthoreovirus from southern China.

Porcine orthoreoviruses belong to the family Reoviridae and cause mainly mild enteritis in piglets. We present here the complete genome sequence of a novel porcine orthoreovirus strain (GD-1) isolated from a piglet in southern China. Our data will facilitate future investigations of the molecular characteristics and epidemiology of porcine orthoreoviruses.

Complete genome sequence of a novel porcine Sapelovirus strain YC2011 isolated from piglets with diarrhea.

Sapelovirus is a member of the family Picornaviridae and is emerging as an enteric porcine, simian, and avian pathogen. Here, we report the genome sequence of a novel porcine sapelovirus strain YC2011 isolated from piglets with severe diarrhea. The availability of the genome sequence is helpful to further investigations of molecular characteristics and epidemiology of porcine sapelovirus.

Supplementation of xanthophylls increased antioxidant capacity and decreased lipid peroxidation in hens and chicks.

The present study investigated the effects of xanthophyll supplementation on production performance, antioxidant capacity (measured by glutathione peroxidase, superoxide dismutase (SOD), catalase, total antioxidant capacity (T-AOC), and reduced glutathione:oxidised glutathione ratio (GSH:GSSG)) and lipid peroxidation (measured by malondialdehyde (MDA)) in breeding hens and chicks. In Expt 1, 432 hens were fed diets supplemented with 0 (control group), 20 or 40 mg xanthophyll/kg diet. Blood samples were taken at 7, 14, 21, 28 and 35 d of the trial. Liver and jejunal mucosa were sampled at 35 d. Both xanthophyll groups improved serum SOD at 21 and 28 d, serum T-AOC at 21 d and liver T-AOC, and serum GSH:GSSG at 21, 28 and 35 d and liver GSH:GSSG. Xanthophylls also decreased serum MDA at 21 d in hens. Expt 2 was a 2 × 2 factorial design. Male chicks hatched from 0 or 40 mg in ovo xanthophyll/kg diet of hens were fed a diet containing either 0 or 40 mg xanthophyll/kg diet. Liver samples were collected at 0, 7, 14 and 21 d after hatching. Blood samples were also collected at 21 d. In ovo-deposited xanthophylls increased antioxidant capacity and decreased MDA in the liver mainly within 1 week after hatching. Maternal effects gradually vanished during 1-2 weeks after hatching. Dietary xanthophylls increased antioxidant capacity and decreased MDA in the liver and serum mainly from 2 weeks onwards. Data suggested that xanthophyll supplementation enhanced antioxidant capacity and reduced lipid peroxidation in different tissues of hens and chicks.

Molecular characterization and phylogenetic analysis of infectious bursal disease viruses isolated from chicken in South China in 2011.

Infectious bursal disease virus (IBDV) is a double-stranded RNA virus that causes immunosuppressive disease in young chickens. Thousands of cases of IBDV infection are reported each year in South China, and these infections can result in considerable economic losses to the poultry industry. To monitor variations of the virus during the outbreaks, 30 IBDVs were identified from vaccinated chicken flocks from nine provinces in South China in 2011. VP2 fragments from different virus strains were sequenced and analyzed by comparison with the published sequences of IBDV strains from China and around the world. Phylogenetic analysis of hypervariable regions of the VP2 (vVP2) gene showed that 29 of the isolates were very virulent (vv) IBDVs, and were closely related to vvIBDV strains from Europe and Asia. Alignment analysis of the deduced amino acid (aa) sequences of vVP2 showed the 29 vv isolates had high uniformity, indicated low variability and slow evolution of the virus. The non-vvIBDV isolate JX2-11 was associated with higher than expected mortality, and had high deduced aa sequence similarity (99.2 %) with the attenuated vaccine strain B87 (BJ). The present study has demonstrated the continued circulation of IBDV strains in South China, and emphasizes the importance of reinforcing IBDV surveillance.