Host-mediated ubiquitination of a mycobacterial protein suppresses immunity.

Mycobacterium tuberculosis is an intracellular pathogen that uses several strategies to interfere with the signalling functions of host immune molecules. Many other bacterial pathogens exploit the host ubiquitination system to promote pathogenesis1,2, but whether this same system modulates the ubiquitination of M. tuberculosis proteins is unknown. Here we report that the host E3 ubiquitin ligase ANAPC2-a core subunit of the anaphase-promoting complex/cyclosome-interacts with the mycobacterial protein Rv0222 and promotes the attachment of lysine-11-linked ubiquitin chains to lysine 76 of Rv0222 in order to suppress the expression of proinflammatory cytokines. Inhibition of ANAPC2 by specific short hairpin RNA abolishes the inhibitory effect of Rv0222 on proinflammatory responses. Moreover, mutation of the ubiquitination site on Rv0222 impairs the inhibition of proinflammatory cytokines by Rv0222 and reduces virulence during infection in mice. Mechanistically, lysine-11-linked ubiquitination of Rv0222 by ANAPC2 facilitates the recruitment of the protein tyrosine phosphatase SHP1 to the adaptor protein TRAF6, preventing the lysine-63-linked ubiquitination and activation of TRAF6. Our findings identify a previously unrecognized mechanism that M. tuberculosis uses to suppress host immunity, and provide insights relevant to the development of effective immunomodulators that target M. tuberculosis.

Nuclear cGAS suppresses DNA repair and promotes tumorigenesis.

Accurate repair of DNA double-stranded breaks by homologous recombination preserves genome integrity and inhibits tumorigenesis. Cyclic GMP-AMP synthase (cGAS) is a cytosolic DNA sensor that activates innate immunity by initiating the STING-IRF3-type I IFN signalling cascade1,2. Recognition of ruptured micronuclei by cGAS links genome instability to the innate immune response3,4, but the potential involvement of cGAS in DNA repair remains unknown. Here we demonstrate that cGAS inhibits homologous recombination in mouse and human models. DNA damage induces nuclear translocation of cGAS in a manner that is dependent on importin-α, and the phosphorylation of cGAS at tyrosine 215-mediated by B-lymphoid tyrosine kinase-facilitates the cytosolic retention of cGAS. In the nucleus, cGAS is recruited to double-stranded breaks and interacts with PARP1 via poly(ADP-ribose). The cGAS-PARP1 interaction impedes the formation of the PARP1-Timeless complex, and thereby suppresses homologous recombination. We show that knockdown of cGAS suppresses DNA damage and inhibits tumour growth both in vitro and in vivo. We conclude that nuclear cGAS suppresses homologous-recombination-mediated repair and promotes tumour growth, and that cGAS therefore represents a potential target for cancer prevention and therapy.

Isolation of Non-Tuberculous Mycobacteria (NTM) from Household Water of Patients with NTM Pulmonary Disease.

Objective: This study aims to investigate the prevalence of NTM in household water in China and assess its potential role as a source of infection for NTM pulmonary disease, a crucial step for understanding and controlling the spread of this increasingly prevalent disease. Methods: To examine the prevalence of mycobacteria in household water, 500 mL water samples and swabs were collected from all taps of 19 patients' homes. The amplification of mycobacterial 16SrRNA with bacteriological identification was as a protocol to discriminate mycobacterial isolations from non- mycobacterial isolations. The 570bp 16SrRNA amplicon was sequenced and used to define mycobacterial species. Results: The mycobacteria isolated from clinical samples from 19 patients included M. intracellulare, M. avium, M. abscessus, and M. kansasii. NTM isolated from household water of patients included M. avium (1 case), M. abscessus (2 cases), M. kansasii (8 cases), M. gordonae (1 case), M. gilvum (1 case), M. fortuitum (1 case), M. porcinum (1 case). M. abscessus, M. kansasii, and M. avium causing human disease were isolated from household water. Though M. intracellulare was the predominant species isolated from patients with NTM pulmonary disease, it was not found in household water. In addition, our results revealed that NTM preferentially colonize in biofilm/sediment (75% of positive growths were from tap swab samples), indicating the significance of finding specific NTM species in household water in relation to the patients' conditions, or the lack of correlation between M. intracellulare in patients and its absence in household water. Conclusions: The isolation of pathogenic NTM species from household water underscores the critical role of water hygiene in preventing NTM pulmonary disease and highlights the need for targeted public health strategies.

HNRNPC suppresses tumor immune microenvironment by activating Treg cells promoting the progression of prostate cancer.

Immune microenvironment could affect the biological progress in prostate cancer (PCa) through N6 methyl adenosine (m6A) methylation. The purpose of this study was to investigate the crosstalk between m6A methylation and immune microenvironment and explore potential biomarkers to improve the immunotherapeutic response. Firstly, according to 11 differentially expressed m6A genes between normal and tumor samples, PCa patients were divided into immune microenvironment subtype 1 (IMS1) and IMS2 based on m6A gene profiles extracted from The Cancer Genome Atlas (TCGA) database. IMS2 showed an immune "cold" phenotype with worse prognoses, and HNRNPC was identified as the biomarker of IMS2 by the protein-protein interaction network. Furthermore, through bioinformatics analyses and in vitro experiments, we found that HNRNPC-high patients showed a suppressive immune-infiltrating tumor microenvironment with a higher infiltration of regulatory T (Treg) cells. Finally, we cocultured transfected PCa cells with peripheral blood mononuclear cells (PBMC) and verified that HNRNPC inhibits tumor immunity by elevating the activation of Treg cells and suppression of effector CD8 T cell. In conclusion, we identified a "cold" immune phenotype in PCa, and HNRNPC regulating the activation of Treg cells. Activation of the immune microenvironment through targeting HNRNPC may be a potential therapeutic option for advanced PCa.

Discrepant modulating effects of dietary docosahexaenoic acid on cerebral lipids, fatty acid transporter expression and soluble beta-amyloid levels in ApoE-/- and C57BL/6J mice.

AIMS: The study was designed to explore the role of apolipoprotein E (ApoE) deficiency concomitant with dietary docosahexaenoic acid (DHA) treatment on brain ß-amyloid (Aß) and lipid levels. METHOD: A 5-month dietary DHA intervention was conducted in ApoE-deficient (ApoE-/- ) mice and wild-type C57BL/6J (C57 wt) mice. The Morris water maze test was performed to assess the behaviour of the animals. The cortical contents of soluble Aß1-40 and Aß1-42 were detected by enzyme-linked immunosorbent assay (ELISA). Cortical fatty acid levels were detected by gas chromatography. Gene and protein expression of molecules associated with cerebral Aß and lipid metabolism were measured using real-time polymerase chain reaction (PCR), Western blot and histological methods. RESULTS: DHA treatment increased the content of cortical DHA and n-3 polyunsaturated fatty acids (n-3 PUFAs) but decreased the ratio of n-6/n-3 PUFAs in ApoE-/- mice; whereas the content of cortical DHA and n-3 PUFAs in C57 wt mice remained unchanged after DHA treatment. Cerebral Fabp5 and Cd36 gene expression were significantly downregulated in DHA-fed C57 wt mice; cerebral Cd36 and Scarb1 gene expression were significantly upregulated, whereas Fabp5 gene expression was downregulated in DHA-fed ApoE-/- mice. In comparison with C57 wt mice, the content of cortical soluble Aß1-42 , total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) increased, whereas the level of high-density lipoprotein cholesterol (HDL-C) decreased in ApoE-/- mice. Interestingly, these differences were significantly reversed by DHA dietary treatment. CONCLUSION: DHA intervention has discrepant impacts on cerebral lipids, fatty acid transporter expression and soluble Aß levels in ApoE-/- and C57 wt mice, suggesting the modifying role of ApoE status on the responses of cerebral lipids and Aß metabolism to DHA treatment.

Sensing of mycobacterial arabinogalactan by galectin-9 exacerbates mycobacterial infection.

Mycobacterial arabinogalactan (AG) is an essential cell wall component of mycobacteria and a frequent structural and bio-synthetical target for anti-tuberculosis (TB) drug development. Here, we report that mycobacterial AG is recognized by galectin-9 and exacerbates mycobacterial infection. Administration of AG-specific aptamers inhibits cellular infiltration caused by Mycobacterium tuberculosis (Mtb) or Mycobacterium bovis BCG, and moderately increases survival of Mtb-infected mice or Mycobacterium marinum-infected zebrafish. AG interacts with carbohydrate recognition domain (CRD) 2 of galectin-9 with high affinity, and galectin-9 associates with transforming growth factor ß-activated kinase 1 (TAK1) via CRD2 to trigger subsequent activation of extracellular signal-regulated kinase (ERK) as well as induction of the expression of matrix metalloproteinases (MMPs). Moreover, deletion of galectin-9 or inhibition of MMPs blocks AG-induced pathological impairments in the lung, and the AG-galectin-9 axis aggravates the process of Mtb infection in mice. These results demonstrate that AG is an important virulence factor of mycobacteria and galectin-9 is a novel receptor for Mtb and other mycobacteria, paving the way for the development of novel effective TB immune modulators.

Transcriptional dynamics of the circulating chicken primordial germ cells revealing key genes in cell adhesion and proliferation prior to gonad colonization.

Primordial germ cells (PGCs), precursors to sperms and oocytes, are responsible for the transfer of genetic information to the next generation. The PGCs arise far away from the developing gonad and thus have to migrate across the embryo to reach their site of function. The migration of PGCs from extraembryonic regions to the genital ridges is accomplished through distinct routes among different species. In particular, the birds PGCs utilized the developing circulation system to travel long distance before settling within the gonad. This study screened the transcriptome profile of chicken PGCs isolated from the bloodstream and the genital ridges to identify the cell intrinsic signals that could guide the unique migration path through circulation. We found cell adhesion and extracellular matrix (ECM) associated pathways were highly enriched in the PGCs from blood but not gonads. The platelet-derived growth factor receptors (PDGFRA and PDGFRB) were downregulated during gonad colonization and knockdown of either PDGFRA or PDGFRB inhibit the proliferation of blood PGCs. Furthermore, the migration of blood PGCs was impaired by the suppression of PDGFRA but not PDGFRB. Hence, the chicken PGCs show dynamic transcriptional remodeling during the blood-to-gonad migration and colonization. The free-floating PGCs in the circulation already express genes associated with cell-cell and cell-ECM interactions and therefore prepare for gonadal colonization.

Retro-protein XXA is a remarkable solubilizing fusion tag for inclusion bodies.

BACKGROUND: Producing large amounts of soluble proteins from bacteria remains a challenge, despite the help of current various solubilizing fusion tags. Thus, developing novel tags is necessary. Antifreeze protein (AFP) has excellent solubility and hydrophilicity, but there are no current reports on its use as a solubilizing fusion tag. Additionally, there is no precedent for using retro-proteins (reverse sequence) as solubilizing fusion tags. Therefore, we selected the antifreeze protein AXX and obtained its retro-protein XXA by synthesizing the XXA gene for the development of a new solubilizing fusion tag. RESULTS: XXA exhibits better stability and ease of expression than AXX; hence, we focused the development of the solubilizing fusion tag on XXA. XXA fused with the tested inclusion bodies, significantly increasing the soluble expression compared with commonly used solubilizing fusion tags such as GST, Trx, Sumo, MBP, and NusA. The tested proteins became soluble after fusion with the XXA tag, and they could be purified. They maintained a soluble form after XXA tag removal. Finally, we used enzymatic digestion reaction and western blot experiments to verify that bdNEDP1 and NbALFA, which were soluble expressed by fusion with XXA, were active. CONCLUSION: We developed the novel solubilizing fusion tag XXA, which could more effectively facilitate the soluble expression of inclusion bodies compared with current commonly used tags. XXA could function at both low and high temperatures, and its moderate molecular weight has a limited impact on the output. These properties make XXA an ideal fusion tag for future research and industrial production. Moreover, for the first time, we highlighted the broad potential of antifreeze protein as a solubilizing fusion tag, bringing retro-protein into practical application.

Effectiveness of single loading dose of dexmedetomidine combined with propofol for deep sedation of endoscopic retrograde cholangiopancreatography (ERCP) in elderly patients: a prospective randomized study.

BACKGROUND: Endoscopic retrograde cholangiopancreatography (ERCP) is an advanced endoscopic procedure and requires deep sedation. Deep sedation with dexmedetomidine for the respiratory drive preserved has become popular in recent years. However, the use of dexmedetomidine in elderly patients is controversial because its adverse events are more common. The objective of this study was to investigate the effectiveness of a single loading dose of dexmedetomidine combined with propofol for deep sedation of ERCP in elderly patients. METHODS: In this prospective randomized trial, 49 elderly patients undergoing ERCP were randomly allocated to the dexmedetomidine (DEX) or propofol (PRO) groups. The single loading dose of dexmedetomidine was set at 0.5 µg/kg at the start of anesthesia induction and loading for 10 min. The primary outcome was the cumulative dose of propofol. Secondary outcomes included time to awake, the frequency of airway interventions, and hemodynamics. RESULTS: The intraoperative cumulative dose of propofol was lower in the DEX group (111.0 ± 12.6 µg/kg/min) than the PRO group (143.7 ± 23.4 µg/kg/min) (P < 0.001). There was no statistically significant difference in the time to awake between the two groups. The incidence of artificial airway interventions and hypotension in the PRO group (36%, 60%) were significantly higher than those in the DEX group (4.2%, 16.7%) (P = 0.011, P = 0.003, respectively). In addition, the occurrence of bradycardia increased significantly in the DEX group (58.3%) compared with the PRO group (12%) (P < 0.001). CONCLUSIONS: The single loading dose of dexmedetomidine combined with propofol can reduce propofol consumption and artificial airway intervention and provide better hemodynamic stability than propofol for deep sedation in elderly patients during ERCP. TRIAL REGISTRATION: www.chictr.org.cn (Registration number ChiCTR1900028069, Registration date 10/12/2019).

Determination of Instinct Components of Biomass on the Generation of Persistent Free Radicals (PFRs) as Critical Redox Sites in Pyrogenic Chars for Persulfate Activation.

Persulfate (PS) activation on biochar (BC) is a promising technology for degrading the aqueous organic contaminants. However, the complexity of activation mechanisms and components in biomass that used to produce BC makes it difficult to predict the performance of PS activation. In this study, we employed eight sludges as the representative biomass that contained absolutely different organic or inorganic components. Results showed that the elemental composition, surface properties, and structures of the sludge-derived BCs (SBCs) clearly depended on the inherent components in the sludges. The intensities of persistent free radicals (PFRs) in the electron paramagnetic resonance (EPR) correlated positively with N-containing content of sludges as electron shuttle, but negatively with the metal content as electron acceptor. Linking with PFRs as crucial sites of triggering a radical reaction, a poly-parameter relationship of predicting PS activation for organic degradation using the sludge components was established (kobs,PN = 0.004 × Cprotein + 0.16 × CM-0.895 -0.118). However, for the PS activation on those SBCs without PFRs, this redox process only relied on the sorption or conductivity-related characteristics, not correlating with the content of intrinsic components in biomass but with pyrolysis temperatures. This study provided insightful information of predicting the remediation efficiency of PS activation on BCs and further understanding the fate of contaminants and stoichiometric efficiency of oxidants in a field application.

The Alterations in Methylene Blue/Light-Treated Frozen Plasma Proteins Revealed by Proteomics.

INTRODUCTION: The aim of this study was to investigate the modified proteins in methylene blue/light-treated frozen plasma (MB-FP) compared with fresh frozen plasma (FFP) in order to gain a better application of MB/light-treated plasma in clinic transfusion. METHODS: MB-FP and FFP were collected from Changchun central blood station, and a trichloroacetic acid/acetone precipitation method was used to remove albumin for the enrichment of lower abundance proteins. The plasma protein in MB-FP and FFP were separated using two-dimensional gel electrophoresis (2-DE) and the differentially expressed protein spots were analyzed using mass spectrometry. Finally, the differentially expressed proteins were tested using Western blot and enzyme-linked immunosorbent assay (ELISA). RESULTS: Approximately 14 differentially expressed protein spots were detected in the MB-FP, and FFP was chosen as the control. After 2-DE comparison analysis and mass spectrometry, 8 significantly differentially expressed protein spots were identified, corresponding to 6 different proteins, including complement C1r subcomponent (C1R), inter-alpha-trypsin inhibitor heavy chain H4 (ITI-H4), keratin, type II cytoskeletal 1 (KRT1), hemopexin (HPX), fibrinogen gamma chain (FGG), and transthyretin (TTR). Western blot showed no significant difference in the expression level of KRT1 between MB-FP and FFP (p > 0.05). Both Western blot and ELISA indicated that the level of HPX was significantly higher in FFP than in MB-FP (p < 0.05). CONCLUSION: This comparative proteomics study revealed that some significantly modified proteins occur in MB-FP, such as C1R, ITI-H4, KRT1, HPX, FGG, and TTR. Our findings provide more theoretical data for using MB-FP in transfusion medicine. However, the relevance of the data for the transfusion of methylene blue/light-treated plasma remains unclear. The exact modification of these proteins and the effects of these modified proteins on their functions and their effects in clinical plasma infusion need to be further studied.

Use of thiazide diuretics for the prevention of recurrent kidney calculi: a systematic review and meta-analysis.

BACKGROUND: Thiazide diuretics reduce the risk of recurrent kidney calculi in patients with kidney calculi or hypercalciuria. However, whether thiazide diuretics can definitely prevent recurrent kidney calculi remains unclear. We aimed to evaluate the effect and safety of thiazide diuretics on recurrent kidney calculi. METHODS: The PubMed, Cochrane Library, and EMBASE databases were systematically searched using the keywords thiazide diuretics and kidney calculi to identify randomized controlled trials (RCTs). The primary outcome was the incidence of recurrent kidney calculi, and the secondary outcome was the 24-h urinary calcium level. The pooled risk ratio (RR), risk difference (RD), standardized mean difference (SMD), and 95% confidence interval (CI) were calculated. The evidence quality was graded using the GRADE criteria, and recommendations for recurrent kidney calculus prevention using thiazide diuretics were reassessed. RESULTS: Eight RCTs involving 571 patients were included. The pooled RR for the incidence of kidney calculi in the thiazide diuretic groups was 0.44 (95% CI 0.33-0.58, P < 0.0001) compared to that in the placebo and untreated groups; the pooled RD was - 0.23 (95% CI - 0.30 to - 0.16, P < 0.0001). The pooled SMD for the 24-h urinary calcium level was - 18.59 (95% CI - 25.11 to - 12.08, P < 0.0001). The thiazide diuretic groups had a high incidence of adverse reactions and low tolerance. The evidence quality for decrease in kidney calculus incidence using thiazide diuretics was low, while that for the 24-h urinary calcium level decrease among those with recurrent kidney calculi was moderate, and that for the decrease in kidney calculus incidence using short-acting and long-acting thiazide diuretics was low. The overall strength of recommendation for prevention of recurrent renal calculi using thiazide diuretics was not recommended. The subgroup and sensitivity analysis findings were robust. CONCLUSIONS: Long-term use of thiazide diuretics reduces the incidence of recurrent renal calculi and 24-h urinary calcium level. However, the benefits are insufficient, and the evidence quality is low. Considering the adverse effects, poor patient compliance, and economic burden of long-term medication, their use in preventing recurrent kidney calculi is not recommended.

Temporal patterns of diversification in Brassicaceae demonstrate decoupling of rate shifts and mesopolyploidization events.

BACKGROUND AND AIMS: Whole-genome duplication (WGD) events are considered important driving forces of diversification. At least 11 out of 52 Brassicaceae tribes had independent mesopolyploid WGDs followed by diploidization processes. However, the association between mesopolyploidy and subsequent diversification is equivocal. Herein we show the results from a family-wide diversification analysis on Brassicaceae, and elaborate on the hypothesis that polyploidization per se is a fundamental driver in Brassicaceae evolution. METHODS: We established a time-calibrated chronogram based on whole plastid genomes comprising representative Brassicaceae taxa and published data spanning the entire Rosidae clade. This allowed us to set multiple calibration points and anchored various Brassicaceae taxa for subsequent downstream analyses. All major splits among Brassicaceae lineages were used in BEAST analyses of 48 individually analysed tribes comprising 2101 taxa in total using the internal transcribed spacers of nuclear ribosomal DNA. Diversification patterns were investigated on these tribe-wide chronograms using BAMM and were compared with family-wide data on genome size variation and species richness. KEY RESULTS: Brassicaceae diverged 29.9 million years ago (Mya) during the Oligocene, and the majority of tribes started diversification in the Miocene with an average crown group age of about 12.5 Mya. This matches the cooling phase right after the Mid Miocene climatic optimum. Significant rate shifts were detected in 12 out of 52 tribes during the Mio- and Pliocene, decoupled from preceding mesopolyploid WGDs. Among the various factors analysed, the combined effect of tribal crown group age and net diversification rate (speciation minus extinction) is likely to explain sufficiently species richness across Brassicaceae tribes. CONCLUSIONS: The onset of the evolutionary splits among tribes took place under cooler and drier conditions. Pleistocene glacial cycles may have contributed to the maintenance of high diversification rates. Rate shifts are not consistently associated with mesopolyploid WGD. We propose, therefore, that WGDs in general serve as a constant 'pump' for continuous and high species diversification.

Enhanced Transformation of Cr(VI) by Heterocyclic-N within Nitrogen-Doped Biochar: Impact of Surface Modulatory Persistent Free Radicals (PFRs).

Redox processes mediated by biochar(BC) enhanced the transformation of Cr(VI), which is largely dependent on the presence of PFRs as electron donors. Natural or artificial dopants in BC's could regulate inherent carbon configuration and PFRs. Until recently, the modulation of PFRs and transformation of Cr(VI) in BC by nonmetal-heterocyclic dopants was barely studied. In this study, changes in PFRs introduced by various nitrogen-dopants within BC are presented and the capacity for Cr(VI) transformation without light was investigated. It was found N-dopants were effectively embedded in carbon lattices through activated-Maillard reaction thus altering their charge and PFRs. Transformation of Cr(VI) in N doped biochar relied on mediated direct reduction by surface modulatory PFRs. The kinetic rate of transformation of Cr(VI) was increased 1.4-5 fold in N-BCs compared to nondoped BCs. Theortical calculation suggested a deficiency in surface electrons induced Lewis acid-base bonding which could acted as a bridge for electron transfer. Results of PCA and orbital energy indicated a colinear relationship between PFRs and pyrrolic N, as well as its dual-mode transformation of Cr(VI). This study provides an improved understanding of how N-doped BC contributes to the evolution of PFRs and their corresponding impacts on the transformation of Cr(VI) in environments.

Preparation and application of a novel magnetic molecularly imprinted polymer for simultaneous and rapid determination of three trace endocrine disrupting chemicals in lake water and milk samples.

Exposure to endocrine disruptor substances will alter the function of the endocrine system and then cause adverse effects on human health. Among these endocrine disrupting chemicals, hexestrol, nonylphenol, and bisphenol A are most commonly used worldwide. In this study, we aim to develop a simple, rapid, and efficient analytical method for the simultaneous determination of trace hexestrol, nonylphenol, and bisphenol A in lake water and milk samples. A magnetic molecularly imprinted polymer-assisted magnetic solid-phase extraction technique was applied. The magnetic molecularly imprinted polymer was prepared and characterized by electron scanning microscopy and Fourier transform infrared spectroscopy. Subsequently, different experiments were conducted to optimize the magnetic solid-phase extraction conditions. High-performance liquid chromatography with UV detection was employed to determine hexestrol, nonylphenol, and bisphenol A. Limits of detection of the developed method were from 0.1 to 0.3 µg L-1 and spiked recoveries ranged from 89.9 to 102.5%, with a relative standard deviation of < 2.5% (intraday). Results obtained from this study showed that the proposed magnetic solid-phase extraction method was a simple, rapid, and sensitive sample pre-treatment method for the determination of trace hexestrol, nonylphenol, and bisphenol in different aqueous samples.

Trop2 Promotes Multidrug Resistance by Regulating Notch1 Signaling Pathway in Gastric Cancer Cells.

BACKGROUND Chemotherapy is widely used in gastric cancer treatment, but multidrug resistance remains a leading cause of chemotherapy failure. Trop2 is highly expressed in gastric tumor tissues and greatly influences cancer progression. However, little is known about the relationship between Trop2 and drug resistance in gastric cancer. MATERIAL AND METHODS In the present study, Trop2 was knocked down in BGC823 cells and overexpressed in HGC27. CCK-8 assay was performed to explore the relationship of Trop2 expression and cell proliferation treated with anticancer drugs. Flow cytometry was performed to assess the relationship between Trop2 and cell apoptosis after chemotherapy. Subcutaneous xenograft models were generated to explore the curative effect of DDP to GC in vivo. MRP1 and Notch1 expressions were assessed by Western blot. RESULTS Trop2 decreased cell proliferation inhibition and apoptosis after chemotherapeutic treatments. DDP showed stronger therapeutic effects on Trop2-knockdown tumor than control in vivo. MRP1 and Notch1 signaling pathway were confirmed to participate in Trop2-induced drug resistance. CONCLUSIONS Our findings suggest that Trop2 promotes the resistance of gastric cancer to chemotherapy by activating the Notch1 pathway.

A deletion in the RD105 region confers resistance to multiple drugs in Mycobacterium tuberculosis.

BACKGROUND: The emergence of drug-resistant strains of Mycobacterium tuberculosis (Mtb), especially those that are multidrug resistant poses a serious threat to global tuberculosis control. However, the mechanism underlying the occurrence of drug resistance against more than one drug is poorly understood. Given that the Beijing/W strains are associated with outbreaks and multidrug resistance, they may harbor a genetic advantage and provide useful insight into the disease. One marker found in all Beijing/W Mtb strains is a deletion of RD105 region that results in a gene fusion, Rv0071/74, with a variable number (3-9 m) of VDP (V: Val, D: Asp; P: Pro) repeats (coded by gtggacccg repeat sequences) at the N-terminal. Here, we report that this variable number of VDP repeats in Rv0071/74 regulates the development of multidrug resistance. RESULTS: We collected and analyzed 1255 Beijing/W clinical strains. The results showed that the number of VDP repeats in Rv0071/74 was related to the development of multidrug resistance, and the deletion of Rv0071/74-9 m from Beijing/W clinical strain restored drug susceptibility. Rv0071/74-9 m also increased resistance to multiple drugs when transferred to different mycobacterial strains. Cell-free assays indicate that the domain carrying 4-9 VDP repeats (4-9 m) showed a variable binding affinity with peptidoglycan and Rv0071/74 cleaves peptidoglycan. Furthermore, Rv0071/74-9 m increased cell wall thickness and reduced the intracellular concentration of antibiotics. CONCLUSIONS: These findings not only identify Rv0071/74 with VDP repeats as a newly identified multidrug resistance gene but also provide a new model for the development of multiple drug resistance.

A novel gene arrangement among the Stylommatophora by the complete mitochondrial genome of the terrestrial slug Meghimatium bilineatum (Gastropoda, Arionoidea).

Stylommatophora is a main clade of Gastropoda that encompasses approximately 112 gastropod families and may exceed a total of 30,000 species. Twenty-four complete stylommatophoran mitogenomes have been sequenced to date, yet our understanding of mitochondrial evolution in stylommatophorans is still in its infancy. To further expand the set of available mitogenomes, we sequenced the mitogenome of Meghimatium bilineatum (Arionoidea: Philomycidae), a widespread land slug in East Asia. This is the first report on a mitogenome of the superfamily Arionoidea, and indeed on a terrestrial slug. The mitogenome of Meghimatium bilineatum comprises 13,972 bp and exhibits a novel, highly distinctive gene arrangement among the Stylommatophora. Phylogenetic reconstructions based on the sequences of all protein-coding genes consistently recovered Meghimatium bilineatum as sister-group of the Succineidae. A phylogenetic reconstruction based on gene order, however, suggested a highly divergent tree topology, which is less credible when taking into account prior knowledge of stylommatophoran relationships. Our CREx (Common interval Rearrangement Explorer) analysis suggested that three successive events of tandem duplication random loss (TDRL) best explain the evolutionary process of gene order rearrangement in Meghimatium bilineatum from an ancestral stylommatophoran mitogenome. The present example offers new insights into the mechanisms of mitogenome rearrangements in gastropods at large and into the usefulness of mitogenomic gene order as a phylogenetic marker.

Towards a global phylogeny of freshwater mussels (Bivalvia: Unionida): Species delimitation of Chinese taxa, mitochondrial phylogenomics, and diversification patterns.

The Yangtze River Basin in China is one of the global hotspots of freshwater mussel (order Unionida) diversity with 68 nominal species. Few studies have tested the validity of these nominal species. Some taxa from the Yangtze unionid fauna have not been adequately examined using molecular data and well-positioned phylogenetically with respect to the global Unionida. We evaluated species boundaries of Chinese freshwater mussels, and disentangled their phylogenetic relationships within the context of the global freshwater mussels based on the multi-locus data and complete mitochondrial genomes. Moreover, we produced the time-calibrated phylogeny of Unionida and explored patterns of diversification. COI barcode data suggested the existence of 41 phylogenetic distinct species from our sampled 40 nominal taxa inhabiting the middle and lower reaches of the Yangtze River. Maximum likelihood and Bayesian inference analyses on three loci (COI, 16S, and 28S) and complete mitochondrial genomes showed that the subfamily Unioninae sensu stricto was paraphyletic, and the subfamily Anodontinae should be subsumed under Unioninae. In addition, we described two new tribes (Aculamprotulini tribe nov. and Lepidodesmini tribe nov.) in the subfamily Unioninae and one new genus (Parvasolenaiagen. nov.) in the subfamily Gonideinae. Molecular dating analysis suggested freshwater mussels diversified at 346.1 Mya (HPD = 286.6-409.9). The global diversification rate for Unionida was estimated to be 0.025 species/Myr. Our study found only a single well-supported rate shift in Unionida diversification, occurring at the base of the subfamily Ambleminae. The evolution of active host-attraction may have triggered the burst of speciation in Ambleminae, and the environment and geography of the Mississippi River Basin likely sustained this radiation.

Rapid determination of antiviral medication ribavirin in different feedstuffs using a novel magnetic molecularly imprinted polymer coupled with high-performance liquid chromatography.

A novel magnetic molecularly imprinted polymer adsorbing material was successfully synthesized to detect ribavirin in animal feedstuff. Molecularly imprinted polymer was prepared through surface polymerization by using ribavirin as template molecule, methyl methacrylate, and γ-methacryloxypropyl trimethoxy silane functionalized magnetic mesoporous silica as bifunctional monomers, and ethylene diglycidyl ether as crosslinking agent. The prepared magnetic molecularly imprinted polymer was characterized by scanning electron microscopy and infrared spectroscopy. Static and dynamic adsorption experiments and selective adsorption analysis were performed to evaluate the adsorption and selectivity of magnetic molecularly imprinted polymer. Different experiments were conducted to optimize the magnetic solid-phase extraction conditions. Under optimal experimental conditions, a magnetic molecularly imprinted solid-phase extraction coupled with high-performance liquid chromatography method was successfully developed for ribavirin detection. The established method achieved a satisfactory linear range of 0.20-50 mg/L (R2  > 0.99) and a low detection limit (0.081 mg/kg). An average recovery of 92-105% with relative standard deviation of <6.5% was obtained upon the application of the developed method to detect ribavirin in real feedstuff samples. Thus, established method can be used for the rapid and simple separation and detection of added ribavirin in feedstuff.