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Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) is one of the major viral pathogens in silkworm. Its infection often results in significant losses to sericulture. Studies have demonstrated that RNAi is one of the important anti-viral mechanisms in organisms. In this study, three dsRNAs targeting the RNA-dependent RNA polymerase (RDRP) gene of BmCPV were designed and synthesized with 2′-F modification to explore their interference effects on BmCPV replication in silkworm larvae. The results showed that injecting dsRNA in the dosage of 4–6 ng per mg body weight into the 5th instar larvae can interfere with the BmCPV-RDRP expression by 93% after virus infection and by 99.9% before virus infection. In addition, the expression of two viral structural protein genes (genome RNA segments 1 and 5) was also decreased with the decrease of RDRP expression, suggesting that RNAi interference of BmCPV-RDRP expression could affect viral replication. The study provides an effective method for investigating virus replication as well as the virus–host interactions in the silkworm larvae using dsRNA. [ABSTRACT FROM AUTHOR]

Abstract: The immunotoxicity of tributyltin (TBT) on marine gastropods has been comparatively little studied although risks to wildlife associated with this compound are well known. In this study, a 30-day trial was conducted to evaluate the immunotoxic effects on abalone (Haliotis diversicolor supertexta) by exposing a range of doses of TBT (0, 2, 10, and 50ng/L). Innate immune parameters, including phagocytic ability (PA), lysozyme activity, phenoloxidase (PO) level and superoxide dismutase (SOD) activity were monitored at intervals of 5, 15 and 30 days. Haemolymph protein expression profile was also examined at the end of the experiment. The results showed that PA value, lysozyme activity and PO level significantly decreased compared with the controls (P <0.05), which indicated that TBT exposure markedly suppressed non-specific immune competence. Exposure to TBT also caused variation in protein expression patterns of haemolymph. Among the protein spots of differential expressions, seven proteins from the haemolymph of TBT-treated abalone were successfully identified by MALDI-TOF-MS analysis. Three protein spots increased and were identified as carrier-like peptide, peroxidase 21 precursor and creatine phosphokinase. These proteins are believed to up-regulate in expression as a response to detoxification and antioxidative stress mechanisms. The other four protein spots that down-regulated in TBT-treated groups were identified as aromatase-like protein, protein kinase C, ceruloplasmin and microtubule-actin crosslinking factor 1, and these proteins play an important role in endocrine regulation and immune defense. Taken together, the results demonstrate that TBT impair abalone immunological ability and is a potential immune disruptor. [ABSTRACT FROM AUTHOR]

Abstract: The contamination of marine ecosystems by endocrine disrupting compounds (EDCs) is of great concern. Protein expression profile maybe a good method to help us understand the molecular mechanisms of EDCs-toxicity to aquatic organisms. In this study, the abalone (Haliotis diversicolor supertexta), was selected as the target organism. Toxicological effects of two reference endocrine disruptors: diallyl phthalate (DAP, 50μgL−1) and bisphenol-A (BPA, 100μgL−1) were investigated after a three months static-renewal exposure on abalones using proteomics to analyze their hepatopancreas tissues. Some enzyme activity parameters of hepatopancreas extracts were also performed, including Na+–K+-ATPase, Ca2+–Mg2+-ATPase, peroxidase (POD) and malondialdehyde (MDA) production. After analyzing the proteomics profile of hepatopancreas by 2D gel electrophoresis, we found that 24 spots significantly increased or decreased at protein expression level (⩾2-fold difference) in the 2D-maps from the treatment groups. Eighteen out of 24 protein spots were successfully identified by matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI–TOF-MS). These proteins can be roughly categorized into diverse functional classes such as detoxification, oxidative stress, hormone regulating, cellular metabolism and innate immunity. In addition, the enzymatic results indicated that DAP/BPA exposure affected the oxidative stress status and the cellular homeostasis, which partly corroborated the proteomics’ results. Taken together, these data demonstrate that proteomics is a powerful tool to provide valuable insights into possible mechanisms of toxicity of EDCs contaminants in aquatic species. Additionally, the results highlight the potential of abalone as a valuable candidate for investigating EDCs impacts on marine ecosystems. [Copyright &y& Elsevier]

Abstract: S-(1,1,2,2-Tetrafluoroethyl)-l-cysteine (TFEC), a major metabolite of the industrial gas tetrafluoroethylene, has been shown to mediate nephrotoxicity by necrosis. TFEC-induced cell death is associated with an early covalent modification of specific intramitochondrial proteins; including aconitase, α-ketoglutarate dehydrogenase (KGDH) subunits, HSP60 and HSP70. Previous studies have indicated that the TAMH line accurately models TFEC-induced in vivo cell death with dose- and time-dependent inhibitions of both KGDH and aconitase activities. Here, we show that the molecular pathway leading to TFEC-mediated cell death is associated with an early cytosolic to mitochondrial translocation of BAX, a pro-apoptotic member of the BCL-2 family. Immunoblot analyses indicated movement of BAX (21kDa) to the mitochondrial fraction after exposure to a cytotoxic concentration of TFEC (250μM). Subsequent cytochrome c release from mitochondria was also demonstrated, but only a modest increase in caspase activities was observed, suggesting a degeneration of early apoptotic signals into secondary necrosis. Significantly, TAMH cells overexpressing BCL-xL preserved cell viability even to supratoxicological concentrations of TFEC (≤600μM), and this cytoprotection was associated with decreased HSP70i upregulation, indicating suppression of TFEC-induced proteotoxicity. Hence, TFEC-induced necrotic cell death in the TAMH cell line is mediated by BAX and antagonized by the anti-apoptotic BCL-2 family member, BCL-xL. [Copyright &y& Elsevier]

[Display omitted] • First report whole genome wide comparative analysis of drought tolerant wild barley vs cultivated barley. • Drought tolerance in Tibetan wild barley attributes to candidate genes with unique genetic variants and expression patterns. • Wild barley employs effective regulators to activate various responsive pathways to adapted to water deficit conditions. • Novel functional genes HvZIFL2 and HvPOD11 positively regulate drought tolerance. Frequent climate change-induced drought events are detrimental environmental stresses affecting global crop production and ecosystem health. Several efforts have facilitated crop breeding for resilient varieties to counteract stress. However, progress is hampered due to the complexity of drought tolerance; a greater variety of novel genes are required across varying environments. Tibetan annual wild barley is a unique and precious germplasm that is well adapted to abiotic stress and can provide elite genes for crop improvement in drought tolerance. To identify the genetic basis and unique mechanisms for drought tolerance in Tibetan wild barley. Whole genome resequencing and comparative RNA-seq approaches were performed to identify candidate genes associated with drought tolerance via investigating the genetic diversity and transcriptional variation between cultivated and Tibetan wild barley. Bioinformatics, population genetics, and gene silencing were conducted to obtain insights into ecological adaptation in barley and functions of key genes. Over 20 million genetic variants and a total of 15,361 significantly affected genes were identified in our dataset. Combined genomic, transcriptomic, evolutionary, and experimental analyses revealed 26 water deficit resilience-associated genes in the drought-tolerant wild barley XZ5 with unique genetic variants and expression patterns. Functional prediction revealed Tibetan wild barley employs effective regulators to activate various responsive pathways with novel genes, such as Zinc-Induced Facilitator-Like 2 (HvZIFL2) and Peroxidase 11 (HvPOD11), to adapt to water deficit conditions. Gene silencing and drought tolerance evaluation in a natural barley population demonstrated that HvZIFL2 and HvPOD11 positively regulate drought tolerance in barley. Our findings reveal functional genes that have been selected across barley's complex history of domestication to thrive in water deficit environments. This will be useful for molecular breeding and provide new insights into drought-tolerance mechanisms in wild relatives of major cereal crops. [ABSTRACT FROM AUTHOR]

[Display omitted] • microRNA expression compared in Cd-tolerant/sensitive barley Zhenong8 vs W6nk2. • First report a novel gene HvNAT2 targeted by miR156g-3p_3 vital in barley Cd tolerance. • HvNAT2 evolved from the Zygnematales in Streptophyte algae and positively regulates Cd tolerance. • Overexpression of HvNAT2 enhances the expression of genes for ROS scavenging and antoxidative enzyme activity. Cadmium (Cd) is one of the most detrimental heavy metal pollutants, seriously affecting crop production and human health. Nucleobase-ascorbic acid transporters (NAT) are widely present in many living organisms including plants, animals and microbes; however, the role of NAT in plant Cd tolerance remains unknown. To identify Cd-induced miRNAs that target HvNAT2 and to determine the role of this gene and its product in Cd tolerance. High-throughput-sequencing was used to identify the miRNA expression profile of barley roots in response to Cd stress. Overexpression (OX) and RNAi lines were then constructed for HvNAT2 and comparative transcriptomic analysis was performed to determine the function of this transporter examining its effects on traits such as Cd uptake/flux and translocation, morphology and antioxidant capacity in relation to Cd tolerance. In addition, phylogenetic analysis was performed to obtain insights into the evolution of HvNAT2. Cd stress-induced genome-wide expression profiles of miRNAs identified a Cd-induced miRNA, miR156g-3p_3, that had HvNAT2 as its target. HvNAT2 was negatively regulated in the high-Cd-accumulating and Cd-tolerant genotype Zhenong8. Evolutionary analysis indicated that orthologues of the plasma membrane localized, HvNAT2 , can be traced back to the sister group of land plants, the streptophyte algae. Overexpression of HvNAT2 increases Cd tolerance with higher tissue Cd accumulation but less oxidative damage in transgenic barley plants. RNAi of HvNAT2 leads to a significant reduction of Cd tolerance. The higher Cd accumulation in roots of the OX3 line was also demonstrated by confocal microscopy and electrophysiology. Transcriptome analysis showed that the enhancement of antioxidant capacity by HvNAT2 was related to stress signaling pathways. Furthermore, oxidative stress tolerance in HvNAT2 -OX plants was regulated by the synthesis of phytochelatins and the glutathione metabolism cycle. Our study reveals a key molecular mechanism of NAT in Cd tolerance in plants that is useful for sustainable agricultural production and management of hazardous this heavy metal for better environment management and ecosystem function. [ABSTRACT FROM AUTHOR]

Summary: Noncanonical Wnt signals control morphogenetic movements during vertebrate gastrulation. Casein kinase I epsilon (CKIɛ) is a Wnt-regulated kinase that regulates Wnt/β-catenin signaling and has a β-catenin-independent role(s) in morphogenesis that is poorly understood. Here we report the identification of a CKIɛ binding partner, SIPA1L1/E6TP1, a GAP (GTPase activating protein) of the Rap small GTPase family. We show that CKIɛ phosphorylates SIPA1L1 to reduce its stability and thereby increase Rap1 activation. Wnt-8, which activates CKIɛ, enhances the CKIɛ-dependent phosphorylation and degradation of SIPA1L1. In early Xenopus or zebrafish development, inactivation of the Rap1 pathway results in abnormal gastrulation and a shortened anterior-posterior axis. Although CKIɛ also transduces Wnt/β-catenin signaling, inhibition of Rap1 does not alter β-catenin-regulated gene expression. Our data demonstrate a role for CKIɛ in noncanonical Wnt signaling and indicate that Wnt regulates morphogenesis in part through CKIɛ-mediated control of Rap1 signaling. [Copyright &y& Elsevier]

• CsGABA-T might indirectly contribute to the synthesis of theanine through transamination. • The high genetic diversity of tea resources may contribute to the different seasonal accumulation patterns of catechins in different varieties. • Accumulation of catechins in summer is primarily due to its biosynthesis. The tea plant (Camellia sinensis L.) is an important commercial crop. Theanine and catechins, abundant in the tea plant and represented as secondary metabolites of the tea plant, contribute to the refreshing and astringent taste of tea, respectively. In this study, three C. sinensis cultivars, ' Baojing Huangjincha 1 ′ (HJC1), ' Huangjincha 2 ′ (HJC2), and ' Huangjincha 168 ′ (HJC168) harvested in spring and summer were used to investigate differences in theanine and catechin concentrations by high-performance liquid chromatography. The expression levels of genes associated with theanine and catechin biosynthesis were investigated using a real-time quantitative polymerase chain reaction. Results indicated that theanine, aspartic acid, serine, glutamate and alanine contents were higher in spring in three cultivars, while CsGDH1 showed higher expression in summer. As for catechins, HJC1 and HJC2 accumulated more epigallocatechin gallate (EGCG) and epicatechin gallate (ECG) contents in summer. On the contrary, HJC168 showed higher EGCG and ECG contents in spring. CsF3H showed higher expression in spring than in summer in three cultivars. Correlation analysis indicated a positive correlation between the CsGABA-T expression and theanine contents in spring and summer (p <0.05), with the activity of GABA-T in theanine synthesis, implying that CsGABA-T might be a crucial positive regulator in theanine accumulation. Besides, most catechin compounds were positively correlated with related genes expression during summer. It was presumed that catechin synthesis contributes to catechin accumulation in summer. In summary, this study provides insights into the potential regulatory mechanisms of theanine and catechins in spring and summer. [ABSTRACT FROM AUTHOR]

Background & Aims: HBV immune escape represents a challenge to prevention, diagnosis, and treatment of hepatitis B. Here, we analyzed the molecular and clinical characteristics of HBV immune escape mutants in a Chinese cohort of chronically infected patients. Methods: Two hundred sixteen patients with HBsAg and anti-HBs were studied, with one hundred eighty-two HBV carriers without anti-HBs as a control group. Recombinant HBsAg bearing the most frequent N-glycosylation mutations were expressed in CHO and HuH7 cells. After confirming N-glycosylation at the most frequent sites (129 and 131), together with inserted mutations, functional analysis were performed to study antigenicity and secretion capacity. Results: One hundred twenty-three patients had the wild-type HBs gene sequence, 93 patients (43%) had mutants in the major hydrophilic region (MHR), and 47 of the 93 patients had additional N-glycosylation mutations, which were transmitted horizontally to at least 2 patients, one of whom was efficiently vaccinated. The frequency of N-glycosylation mutation in the case group was much higher than that of the control group (47/216 vs. 1/182). Compared with wild-type HBsAg, HBsAg mutants reacted weakly with anti-HBs using a chemiluminescent microparticle enzyme immunoassay. Native gel analysis of secreted virion in supernatants of transfected HuH7 cells indicated that mutants had better virion enveloping and secretion capacity than wild-type HBV. Conclusions: Our results suggest that specific HBsAg MHR N-glycosylation mutations are implicated in HBV immune escape in a high endemic area. [Copyright &y& Elsevier]

CENPU, encoding an important factor involved in kinetochore assembly during mitosis, is associated with shorter survival rates in lung adenocarcinoma (LUAD) patients. CENPU promotes growth rates and invasive behavior of LUAD cells; however, its mechanism of action in LUAD progression remains to be elucidated. CENPU mRNA and protein expression were elevated in LUAD tumors, and high CENPU gene expression was associated with inferior survival prognosis in LUAD patients. CENPU knockdown inhibited LUAD cell proliferation, clone formation, migration, invasion, and epithelial-mesenchymal transition (EMT) in addition to inducing cell cycle arrest and apoptosis in vitro and reduced LUAD xenograft tumor growth in vivo. Furthermore, we identified CENPU-regulated genes significantly enriched for proliferation and apoptosis pathways, and identified HSP Family Member C10 (DNAJC10) as putative effector of CENPU. CENPU knockdown produced DNAJC10 protein downregulation, and DNAJC10 overexpression partially rescued the phenotypic effects of CENPU knockdown in LUAD cells. Moreover, CENPU's coiled-coil domain was essential for CENPU's phenotypic effects in LUAD cells. In conclusion, the kinetochore component CENPU plays a critical role in LUAD cell proliferation and invasiveness. Targeting CENPU-DNAJC10 axis may inhibit LUAD tumor cell proliferation and metastasis. [ABSTRACT FROM AUTHOR]