CD137 expression and signal function drive pleiotropic γδ T-cell effector functions that inhibit intracellular M. tuberculosis growth.

Co-activation signal that induces/sustains pleiotropic effector functions of antigen-specific γδ T cells remains unknown. Here, Mycobacteria tuberculosis (Mtb) tuberculin administration during tuberculosis (TB) skin test resulted in rapid expression of co-activation signal molecules CD137 and CD107a by fast-acting Vγ2Vδ2 T cells in TB-resistant subjects (Resisters), but not patients with active TB. And, anti-CD137 agonistic antibody treatment experiments showed that CD137 signaling enabled Vγ2Vδ2 T cells to produce more effector cytokines and inhibit intracellular Mtb growth in macrophages (Mɸ). Consistently, Mtb antigen (Ag) HMBPP stimulation induced sustainable high-level CD137 expression in fresh and activated Vγ2Vδ2 T cells from uninfected subjects, but not TB patients. CD137+Vγ2Vδ2 T-cell subtype predominantly displayed central memory phenotype and mounted better proliferative responses than CD137-Vγ2Vδ2 T-cells. In response to HMBPP, CD137+Vγ2Vδ2 T-cell subtype rapidly differentiated into greater numbers of pleiotropic effector cells producing anti-Mtb cytokines compared to CD137-Vγ2Vδ2 T subtype, with the non-canonical NF-κB pathway involved. CD137 expression in Vγ2Vδ2 T cells appeared to signal anti-Mtb effector functions leading to intracellular Mtb growth inhibition in Mɸ, and active TB disrupted such CD137-driven anti-Mtb effector functions. CD137+Vγ2Vδ2 T-cells subtype exhibited an epigenetic-driven high-level expression of GM-CSF and de novo production of GM-CSF critical for Vγ2Vδ2 T-cell controlling of Mtb growth in Mϕ. Concurrently, exosomes produced by CD137+Vγ2Vδ2 T cells potently inhibited intracellular mycobacterial growth. Furthermore, adoptive transfer of human CD137+Vγ2Vδ2 T cells to Mtb-infected SCID mice conferred protective immunity against Mtb infection. Thus, our data suggest that CD137 expression/signaling drives pleiotropic γδ T-cell effector functions that inhibit intracellular Mtb growth.

Clinical prognostic value of different NPM1 mutations in acute myeloid leukemia patients.

BACKGROUND: Acute myeloid leukemia (AML) patients with various nucleophosmin 1 (NPM1) mutations are controversial in the prognosis. This study aimed to investigate the prognosis of patients according to types of NPM1 mutations (NPM1mut). METHODS: Bone marrow samples of 528 patients newly diagnosed with AML, were collected for morphology, immunology, cytogenetics, and molecular biology examinations. Gene mutations were detected by next-generation sequencing (NGS) technology. RESULTS: About 25.2% of cases exhibited NPM1mut. 83.5% of cases were type A, while type B and D were respectively account for 2.3% and 3.0%. Furthermore, 15 cases of rare types were identified, of which 2 cases have not been reported. Clinical characteristics were similar between patients with A-type NPM1 mutations (NPM1A - type mut) and non-A-type NPM1 mutations (NPM1non - A-type mut). Event-free survival (EFS) was significantly different between patients with low NPM1non - A-type mut variant allele frequency (VAF) and low NPM1A - type mut VAF (median EFS = 3.9 vs. 8.5 months, P = 0.020). The median overall survival (OS) of the NPM1non - A-type mutFLT3-ITDmut group, the NPM1A - type mutFLT3-ITDmut group, the NPM1non - A-type mutFLT3-ITDwt group, and the NPM1A - type mutFLT3-ITDwt group were 3.9, 10.7, 17.3 and 18.8 months, while the median EFS of the corresponding groups was 1.4, 5.0, 7.6 and 9.2 months (P < 0.0001 and P = 0.004, respectively). CONCLUSIONS: No significant difference was observed in OS and EFS between patients with NPM1A - type mut and NPM1non - A-type mut. However, types of NPM1 mutations and the status of FLT3-ITD mutations may jointly have an impact on the prognosis of AML patients.

Efficacy of short-term Peg-IFN α-2b treatment in chronic hepatitis B patients with ultra-low HBsAg levels: a retrospective cohort study.

PURPOSE: Peginterferon alfa-2b (Peg-IFN α-2b) has demonstrated superior efficacy over nucleos(t)ide analogs (NAs) in the treatment of chronic hepatitis B (CHB), particularly among patients with low levels of hepatitis B surface antigen (HBsAg). This study aims to determine whether patients with ultra-low HBsAg levels (< 200 IU/mL) can achieve significantly higher clinical cure rates with abbreviated courses of Peg-IFN α-2b therapy. METHODS: In this retrospective analysis, CHB patients with HBsAg levels below 200 IU/mL were categorized into a Peg-IFN α-2b group and a control group. The Peg-IFN α-2b group received Peg-IFN α-2b for a minimum of 24 weeks, with the possibility of early discontinuation upon achieving HBsAg clearance, and were followed through week 48. The control group remained untreated for hepatitis B virus (HBV), and was observed for 24 weeks. HBsAg clearance rates were compared between groups. Univariate and multivariate logistic regression analyses were employed to identify factors associated with HBsAg clearance . RESULTS: By week 24, the HBsAg clearance rate in the Peg-IFN α-2b group was notably 52.1% (38/73), contrasting sharply with the mere 1.3% (1/77) observed in the control group. Within the Peg-IFN α-2b group, a substantial 97.3% (71/73) of patients noted a reduction in HBsAg levels. Besides, the decision to continue or discontinue treatment after the 24-week mark had no significant impact on the HBsAg clearance rate at week 48. Multivariable analysis pinpointed baseline HBsAg levels (OR = 0.984, p = 0.001) and the presence of fatty liver (OR = 5.960, p = 0.033) as independent predictors of HBsAg clearance. CONCLUSION: Our findings confirm that a 24-week course of Peg-IFN α-2b yields robust efficacy in CHB patients with ultra-low HBsAg levels. Prolonging treatment beyond the 24-week threshold is deemed unnecessary. Both baseline HBsAg level and the presence of fatty liver emerged as significant predictors for HBsAg clearance.

Vitrification cryo-foil method for shoot tip cryopreservation and virus eradication in apple.

Establishment of a new method for improved shoot tip cryopreservation is crucial to facilitate the long-term preservation of plant germplasm as well as the use of cryotherapy for pathogen eradication. The present study reported a vitrification (V) cryo-foil method for shoot tip cryopreservation and virus eradication in apple. Shoot tip regrowth levels after cryopreservation were comparable among V cryo-foil (53 %), V cryo-plate (46 %) and conventional droplet vitrification (Dr-vi, 48 %). The V cryo-foil is more efficient to perform than Dr-vi as more shoot tips can be cryopreserved by one person. In the histological study applying an image-overlaying strategy, shoot tips cryopreserved by V cryo-foil showed a higher survival chance in the youngest leaf primordia than in the apical dome. When V cryo-foil was tested for virus eradication, fifty-five percent (55 %) of cryo-derived shoots were free of the apple stem pitting virus (ASPV), while none and less than 10 % were free of the apple stem grooving virus (ASGV) and the apple chlorotic leaf spot virus (ACLSV), respectively. Thus, these two viruses were efficiently preserved by V cryo-foil cryopreservation. Noticeably, although the shoot regrowth level was reduced to 27 %, a higher frequency (81 %) of ASPV eradication was achieved when a reduced duration of cryoprotectant exposure was applied in V cryo-foil, supporting the use of insufficient cryoprotection for improved virus eradication.

Increased functional connectivity within the salience network in patients with insomnia.

BACKGROUND: Insomnia is a common sleep disorder with significant negative impacts on emotional states; however, the underlying mechanism of insomnia with comorbid emotional dysregulation remains largely unknown. The salience network (SN) plays an important role in both sleep and emotional regulation. The study aimed to explore the specific alterations in functional connectivity (FC) within the SN in insomnia patients. METHODS: A total of 30 eligible patients with insomnia disorder (ID group) and 30 healthy controls (HC group) underwent resting-state functional magnetic resonance imaging (fMRI) scanning and psychometric assessments. Differences in FC within the SN were examined using seed-based region-to-region connectivity analysis. RESULTS: Compared with healthy controls, patients with insomnia showed increased FC within the SN, mainly between the anterior cingulate cortex (ACC) and right superior frontal gyrus (SFG), the right SFG and right supramarginal gyrus (SMG), and between the right insular (INS) and left SMG (P<0.05). Additionally, significant correlations were observed between increased FC and the Hamilton Depression Rating Scale (HAMD), Pittsburgh Sleep Quality Index (PSQI), and Hamilton Anxiety Rating Scale (HAMA) scores (P<0.05, after Bonferroni correction). CONCLUSIONS: These results suggest that increased FC within the SN may be related to poor sleep quality and negative emotions, highlighting the importance of the SN in the pathophysiological mechanisms of insomnia with comorbid emotional dysregulation.

Dosimetric comparison of HyperArc and InCise MLC-based CyberKnife plans in treating single and multiple brain metastases.

BACKGROUND AND PURPOSE: This study aimed to compare the dosimetric attributes of two multi-leaf collimator based techniques, HyperArc and Incise CyberKnife, in the treatment of brain metastases. MATERIAL AND METHODS: 17 cases of brain metastases were selected including 6 patients of single lesion and 11 patients of multiple lesions. Treatment plans of HyperArc and CyberKnife were designed in Eclipse 15.5 and Precision 1.0, respectively, and transferred to Velocity 3.2 for comparison. RESULTS: HyperArc plans provided superior Conformity Index (0.91 ± 0.06 vs. 0.77 ± 0.07, p < 0.01) with reduced dose distribution in organs at risk (Dmax, p < 0.05) and lower normal tissue exposure (V4Gy-V20Gy, p < 0.05) in contrast to CyberKnife plans, although the Gradient Indexes were similar. CyberKnife plans showed higher Homogeneity Index (1.54 ± 0.17 vs. 1.39 ± 0.09, p < 0.05) and increased D2% and D50% in the target (p < 0.05). Additionally, HyperArc plans had significantly fewer Monitor Units (MUs) and beam-on time (p < 0.01). CONCLUSION: HyperArc plans demonstrated superior performance compared with MLC-based CyberKnife plans in terms of conformity and the sparing of critical organs and normal tissues, although no significant difference in GI outcomes was noted. Conversely, CyberKnife plans achieved a higher target dose and HI. The study suggests that HyperArc is more efficient and particularly suitable for treating larger lesions in brain metastases.

Antibiotic Alternatives: Multifunctional Ultra-Small Metal Nanoclusters for Bacterial Infectious Therapy Application.

The prevalence of major bacterial infections has emerged as a significant menace to human health and life. Conventional treatment methods primarily rely on antibiotic therapy, but the overuse of these drugs has led to a decline in their efficacy. Moreover, bacteria have developed resistance towards antibiotics, giving rise to the emergence of superbugs. Consequently, there is an urgent need for novel antibacterial agents or alternative strategies to combat bacterial infections. Nanoantibiotics encompass a class of nano-antibacterial materials that possess inherent antimicrobial activity or can serve as carriers to enhance drug delivery efficiency and safety. In recent years, metal nanoclusters (M NCs) have gained prominence in the field of nanoantibiotics due to their ultra-small size (less than 3 nm) and distinctive electronic and optical properties, as well as their biosafety features. In this review, we discuss the recent progress of M NCs as a new generation of antibacterial agents. First, the main synthesis methods and characteristics of M NCs are presented. Then, we focus on reviewing various strategies for detecting and treating pathogenic bacterial infections using M NCs, summarizing the antibacterial effects of these nanoantibiotics on wound infections, biofilms, and oral infections. Finally, we propose a perspective on the remaining challenges and future developments of M NCs for bacterial infectious therapy.

Chemical characteristics and water stability evaluation of groundwater in the CKDu Zone of Sri Lanka.

Groundwater is the main source of drinking water for the rural population in the chronic kidney disease of unknown etiology (CKDu) zone of the North Central Province (NCP) in Sri Lanka. In this study, a total of 334 groundwater samples (311 dug wells, 21 tube wells and 2 springs) during the wet season from two aquifers in the NCP were collected, and investigated their chemical characteristics and evaluate their water quality, including groundwater chemistry, main ion sources, the corrosion and scaling potential of groundwater. The results showed that the two hydrochemical types of groundwater in the NCP were mainly of the Ca-HCO3, Na·Ca-HCO3 types, with the main HCO3-, Na+ and Ca2+ ions in both types of groundwater originating from silicate and evaporite salt dissolution and influenced by alternating cation adsorption, while the presence of NO3- was mainly anthropogenic. Evaluation of water stability using namely Langelier saturation index (LSI), Ryznar stability index (RSI), Puckorius scaling index (PSI) and Larson-Skold index (LS), indicated that most groundwater presents corrosion potential and has corrosion behavior tendency of metals to some degrees. The water quality of Polonnaruwa was better than that of Anuradhapura in the NCP, and when the groundwater was worse than the "good" grade, which must be properly treated before it is used as drinking water.

Abnormal alterations of regional spontaneous neuronal activity and functional connectivity in insomnia patients with difficulty falling asleep: a resting-state fMRI study.

BACKGROUND: Insomnia disorder (ID) seriously affects people's daily life. Difficulty falling asleep is the most commonly reported complaint in patients with ID. However, the mechanism of prolonged sleep latency (SL) is still obscure. The aim of our present study was to investigate the relationship between prolonged SL and alterations in spontaneous neural activity and brain functional connectivity (FC) in ID patients using functional magnetic resonance imaging (fMRI). METHODS: A total of 52 insomniacs with difficulty falling asleep and 30 matched healthy controls (HCs) underwent resting-state fMRI. The amplitude of low-frequency fluctuation (ALFF) was measured and group differences were compared. The peak areas with significantly different ALFF values were identified as the seed regions to calculate FC to the whole brain. SL was assessed by a wrist actigraphy device in ID patients. The Pittsburgh Sleep Quality Index (PSQI), Hamilton Anxiety Rating Scale (HAMA), and Hyperarousal Scale (HAS) were evaluated in both ID patients and HCs. Finally, correlation analyses were performed between the clinical features and FC/ALFF values. RESULTS: ID patients showed higher PSQI, HAMA, HAS scores than HCs. The functional MRI results indicated increased ALFF value in the left insula and right amygdala and decreased ALFF value in the right superior parietal lobe (SPL) in ID patients. The seed-based FC analysis demonstrated increased FC between the left insula and the bilateral precentral gyrus and FC between the right amygdala and the left posterior cingulate cortex (PCC) in patients with ID. Correlation analysis indicated that the increased FC value of the right amygdala-left PCC was positively correlated with SL measured by actigraphy. CONCLUSION: This study revealed abnormal regional spontaneous fluctuations in the right amygdala, left insula, and right SPL, as well as increased FC in the left insula-precentral and right amygdala-left PCC. Moreover, the prolonged SL was positively correlated with the abnormal FC in the right amygdala-left PCC in ID patients. The current study showed the correlation between prolonged SL and the abnormal function of emotion-related brain regions in ID patients, which may contribute to a better understanding of the neural mechanisms underlying difficulty falling asleep in patients with ID. CLINICAL TRIAL REGISTRATION: http://www.chictr.org.cn ., ChiCTR1800015282. Registered on 20th March 2018.

High expression of IL4I1 is correlated with poor prognosis and immune infiltration in thyroid cancer.

BACKGROUND: Thyroid cancer-related deaths mostly result from metastasis. It was reported that the immunometabolism associated enzyme interleukin-4-induced-1 (IL4I1) was related to tumor metastasis. The present study was intended to investigate the effects of IL4I1 on thyroid cancer metastasis and its relationship with the prognosis. METHODS: Data from Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) were analyzed to find out the different mRNA expressions of IL4I1 between thyroid cancer and normal tissues. And Human Protein Atlas (HPA) was used to assess IL4I1 protein expression. To further differentiate thyroid cancer from normal tissues and estimate the impact of IL4I1 on the prognosis, the receiver operating characteristic curve (ROC) and Kaplan-Meier (KM) method was performed. The protein-protein interaction (PPI) network was established using STRING, and functional enrichment analyses were conducted by "clusterProfiler" package. Then, we assayed the correlation between IL4I1 and some related molecules. The relationship between IL4I1 and immune infiltration was performed using "Gene Set Variation Analysis (GSVA)" package in TCGA and tumor-immune system interaction database (TISIDB). Finally, we did in vitro experiments in order to further prove the bioeffects of IL4I1 on metastasis. RESULTS: The expression of IL4I1 mRNA and IL4I1 protein was significantly upregulated in thyroid cancer tissues. The increment of IL4I1 mRNA expression was related to high-grade malignancy, lymph node metastases and extrathyroidal extension. The ROC curve displayed the cutoff value of 0.782, with the sensitivity of 77.5% and the specificity of 77.8%. KM survival analysis showed that there was a worse PFS in patients with high IL4I1 expression than those with low IL4I1 expression (p = 0.013). Further study indicated that IL4I1 was associated with lactate, body fluid secretion, positive regulation of T cell differentiation, and cellular response to nutrients in Gene Ontology (GO) analysis. Moreover, IL4I1 was found correlated with immune infiltration. Finally, the in vitro experiments revealed the promotion of IL4I1 on cancer cell proliferation, migration and invasion. CONCLUSIONS: The increased IL4I1 expression is markedly correlated with the immune imbalance in the tumor microenvironment (TME) and predicts poor survival in thyroid cancer. This study reveals the potential clinical biomarker of poor prognosis and the target of immune therapy in thyroid cancer.

Potential for efficient microbial remediation of Cr(VI) in wastewater using Deinococcus wulumuqiensis R12.

AIMS: This study aimed to investigate the potential of Deinococcus wulumuqiensis R12 (D. wulumuqiensis R12) as a bioadsorbent for Cr(VI) removal from aqueous solutions. METHODS AND RESULTS: Effects of various factors, including initial concentration of Cr(Ⅵ), pH, adsorbent dosage, and time were explored. The maximal Cr removal efficiency was achieved by adding D. wulumuqiensis R12 to the solution at pH 7.0 for 24 h, with an initial Cr concentration of 7 mg l-1. Characterization of bacterial cells showed that Cr was adsorbed to the surface of D. wulumuqiensis R12 by combining with functional groups, such as carboxyl and amino groups on the surface. Furthermore, D. wulumuqiensis R12 was able to keep its bioactivity in the presence of Cr and tolerate Cr concentrations as high as 60 mg l-1. CONCLUSIONS: Deinococcus wulumuqiensis R12 demonstrates a comparatively high adsorption capacity for Cr(VI). Under the optimized conditions, the removal ratio reached 96.4% with 7 mg l-1 Cr(VI), and the maximal biosorption capacity was 2.65 mg g-1. More importantly, it was found that D. wulumuqiensis R12 still had strong metabolic activity and maintained its viability after adsorbing Cr(VI), which is beneficial for biosorbent stability and reuse.

Low expression of the intestinal metabolite butyric acid and the corresponding memory pattern regulate HDAC4 to promote apoptosis in rat hippocampal neurons.

After intensive research on the gut-brain axis, intestinal dysbiosis is considered to be one of the important pathways of cognitive decline. Microbiota transplantation has long been thought to reverse the behavioral changes in the brain caused by colony dysregulation, but in our study, microbiota transplantation seemed to improve only behavioral brain function, and there was no reasonable explanation for the high level of hippocampal neuron apoptosis that remained. Butyric acid is one of the short-chain fatty acids of intestinal metabolites and is mainly used as an edible flavoring. It is commonly used in butter, cheese and fruit flavorings, and is a natural product of bacterial fermentation of dietary fiber and resistant starch in the colon, acting similarly to the small-molecule HDAC inhibitor TSA. The effect of butyric acid on HDAC levels in hippocampal neurons in the brain remains unclear. Therefore, this study used rats with low bacterial abundance, conditional knockout mice, microbiota transplantation, 16S rDNA amplicon sequencing, and behavioral assays to demonstrate the regulatory mechanism of short-chain fatty acids on the acetylation of hippocampal histones. The results showed that disturbance of short-chain fatty acid metabolism led to high HDAC4 expression in the hippocampus and regulated H4K8ac, H4K12ac, and H4K16ac to promote increased neuronal apoptosis. However, microbiota transplantation did not change the pattern of low butyric acid expression, resulting in maintained high HDAC4 expression in hippocampal neurons with continued neuronal apoptosis. Overall, our study shows that low levels of butyric acid in vivo can promote HDAC4 expression through the gut-brain axis pathway, leading to hippocampal neuronal apoptosis, and demonstrates that butyric acid has great potential value for neuroprotection in the brain. In this regard, we suggest that patients with chronic dysbiosis should pay attention to changes in the levels of SCFAs in their bodies, and if deficiencies occur, they should be promptly supplemented through diet and other means to avoid affecting brain health.

Dual-Enzyme Cascade Composed of Chitosan Coated FeS2 Nanozyme and Glucose Oxidase for Sensitive Glucose Detection.

Immobilizing enzymes with nanozymes to catalyze cascade reactions overcomes many of the shortcomings of biological enzymes in industrial manufacturing. In the study, glucose oxidases were covalently bound to FeS2 nanozymes as immobilization carriers while chitosan encapsulation increased the activity and stability of the immobilized enzymes. The immobilized enzymes exhibited a 10% greater increase in catalytic efficiency than the free enzymes while also being more stable and catalytically active in environments with an alkaline pH of 9.0 and a high temperature of 100 °C. Additionally, the FeS2 nanozyme-driven double-enzyme cascade reaction showed high glucose selectivity, even in the presence of lactose, dopamine, and uric acid, with a limit of detection (LOD) (S/N = 3) as low as 1.9 × 10-6 M. This research demonstrates that nanozymes may be employed as ideal carriers for biological enzymes and that the nanozymes can catalyze cascade reactions together with natural enzymes, offering new insights into interactions between natural and synthetic biosystems.

Whole-exome sequencing identified novel variants in CPLANE1 that causes oral-facial-digital syndrome â ¥ by inducing primary cilia abnormality.

Oral-facial-digital syndrome (OFDS) is a multisystemic ciliopathic disorder with an autosomal recessive mode of inheritance. OFDS usually manifests with typical craniofacial anomalies and variable occurrence of polydactyly. Germline variants in CPLANE1 cause OFDS VI. In this study, we investigated a 26-year-old Chinese female patient who was 23+1  weeks pregnant. She had a history of adverse pregnancy outcomes with multiple foetal malformations. We performed ultrasonography and identified the foetus as having a posterior fossa Blake cyst and postaxial polydactyly. The patient decided to terminate her pregnancy, and further genetic molecular analysis was performed. We identified the aborted foetus as having postaxial polydactyly. Whole-exome sequencing identified a missense variant (c.3599C>T, p.A1200V) in exon 20 and a c.834+1G>T variant in exon 7 of CPLANE1 (NM_023073.3) in the foetus. Sanger sequencing confirmed that these variants came from the parents of the foetus. In this study, we investigated a family with OFDS VI through genetic testing and bioinformatics analysis, which provided powerful help for prenatal diagnosis. Then, we demonstrated that the cell migration rate and the number of cilia were decreased after interference with CPLANE1 expression in NIH/3T3 cells. After CPLANE1 knockdown, the Hh signalling pathway was inhibited, and the Hh pathway activator SAG reversed the inhibitory effect. This is the first report of a family with OFDS VI in the Chinese population.

TRPM2 regulates autophagy to participate in hepatitis B virus replication.

Hepatitis B virus (HBV) affects over 300 million people across the world and is further associated with the self-digesting process of autophagy. Accordingly, the current study set out to explore the role of transient receptor potential cation channel subfamily M member 2 (TRPM2) in HBV replication. Firstly, Huh-7 cells were transfected with the pHBV1.3 plasmid to detect the expression patterns of TRPM2 and neutrophil cytosolic factor 1 (p47 phox), followed by evaluating the role of TRPM2 in autophagy and HBV replication and exploring the interaction between TRPM2 and p47 phox. Collaborative experiments were further designed to explore the role of p47 phox and autophagy in TRPM2 regulation of HBV replication, in addition to animal experimentation to validate the role of TRPM2/p47 phox axis in vivo. It was found that TRPM2 up-regulation was associated with HBV replication. On the other hand, silencing of TRPM2 inhibited HBV replication and autophagy in vitro and in vivo, as evidenced by reduced HBV DNA load, HBV mRNA, HBeAg and HBsAg, and diminished autophagic spot number, LC3 II/I ratio, Beclin-1 expressions and increased p62 expressions. Mechanistic experimentation illustrated that TRPM2 interacted with p47 phox and positively regulated p47 phox, such that p47 phox up-regulation or use of Rapamycin (autophagy activator) weakened the inhibitory role of silencing TRPM2. Collectively, our findings indicated that HBV infection promotes TRPM2 expression, and TRPM2 interacts with p47 phox to induce autophagy and facilitate HVB replication.

Tailoring the Tag/Catcher System by Integrating Covalent Bonds and Noncovalent Interactions for Highly Efficient Protein Self-Assembly.

Covalent bonds and noncovalent interactions play crucial roles in enzyme self-assembly. Here, we designed a Tag/Catcher system named NGTag/NGCatcher in which the Catcher is a highly charged protein that can bind proteins with positively charged tails and rapidly form a stable isopeptide bond with NGTag. In this study, we present a multienzyme strategy based on covalent bonds and noncovalent interactions. In vitro, mCherry, YFP, and GFP can form protein-rich three-dimensional networks based on NGCatcher, NGTag, and RK (Arginine/Lysine) tails, respectively. Furthermore, this technology was applied to improve lycopene production in Escherichia coli. Three key enzymes were involved in lycopene production variants from Deinococcus wulumuqiensis R12 of NGCatcher_CrtE, NGTag_Idi, and RKIspARK, where the multienzyme complexes were clearly observed in vivo and in vitro, and the lycopene production in vivo was 17.8-fold higher than that in the control group. The NGTag/NGCatcher system will provide new opportunities for in vivo and in vitro multienzyme catalysis.

Sequence, structure, and function of the Dps DNA-binding protein from Deinococcus wulumuqiensis R12.

Deinococcus wulumuqiensis R12, which was isolated from arid irradiated soil in Xinjiang province of China, belongs to a genus that is well-known for its extreme resistance to ionizing radiation and oxidative stress. The DNA-binding protein Dps has been studied for its great contribution to oxidative resistance. To explore the role of Dps in D. wulumuqiensis R12, the Dps sequence and homology-modeled structure were analyzed. In addition, the dps gene was knocked out and proteomics was used to verify the functions of Dps in D. wulumuqiensis R12. Docking data and DNA binding experiments in vitro showed that the R12 Dps protein has a better DNA binding ability than the Dps1 protein from D. radiodurans R1. When the dps gene was deleted in D. wulumuqiensis R12, its resistance to H2O2 and UV rays was greatly reduced, and the cell envelope was destroyed by H2O2 treatment. Additionally, the qRT-PCR and proteomics data suggested that when the dps gene was deleted, the catalase gene was significantly down-regulated. The proteomics data indicated that the metabolism, transport and oxidation-reduction processes of D. wulumuqiensis R12 were down-regulated after the deletion of the dps gene. Overall, the data conformed that Dps protein plays an important role in D. wulumuqiensis R12.

Effects of valerate on intestinal barrier function in cultured Caco-2 epithelial cell monolayers.

BACKGROUND: Short-chain fatty acids (SCFAs) are a group of microbial metabolites of undigested dietary fiber, protein and unabsorbed amino acids in the colon, well-known for their gut health promoting benefits. A relatively high intestinal level of valerate was found in the healthy human subjects. However, the intestinal protection effects and the underlying mechanism of valerate are waiting to be verified and elucidated. METHODS AND RESULTS: In the present study, valerate, a SCFAs mainly converted from proteins or amino acids, was demonstrated to promote intestinal barrier function at its physiological concentrations of 0-4 mM in the Caco-2 cell monolayer model of intestinal barrier using transepithelial electrical resistance (TEER) assay and paracellular permeability assay. Valerate achieved the maximum increase in the TEER at 2 mM and reduced the paracellular permeability. Its intestinal barrier function promoting activity is similar to that of butyrate, with a broader range of effective concentrations than the later. Through western blot analysis, this activity is linked to the valerate-induced AMPK activation and tight junctions (TJs) assembly, but not to the reinforced expression of TJs related proteins. CONCLUSIONS: It provides direct experimental evidence supporting valerate's function in intestinal health, implying the once under-valued function of valerate and its amino acid precursors. The valerate's role in regulating intestine homeostasis and its possible synergetic effects with other SCFAs warranted to be further investigated.

A combined "AIE + ESIPT" fluorescent probe for detection of lipase activity.

Lipases are acyl hydrolases that play a key role in fat digestion and monitoring of acute pancreatitis by cleaving long-chain triglycerides into polar lipids. Due to an opposite polarity between the hydrophilic enzyme and their lipophilic substrates, lipase reaction occurs at the interface between the aqueous and the oil phases. It is quite challenging to develop a specific probe to detect lipase activity, especially in homogeneous systems. Herein we designed a blue fluorescent probe HBT-LDC for specific detection of lipase activity based on both aggregation induced emission (AIE) and excited-state intramolecular proton transfer (ESIPT) effect. The probe shows significant advantages, such as high selectivity and excellent sensitivity, no self-quenching at high concentration, large Stokes shift, low cytotoxicity, and good biocompatibility. The linear range for in vitro quantification of lipase is 0.2-1.3 mg/mL with a detection limit of 0.01 mg/mL. The probe has been successfully applied to the evaluation of commercial lipase and the in vivo bioimaging sensing exogenous lipase activity in HeLa cells. The results revealed that the probe could serve as a potential tool in lipase related drug discovery and disease diagnostics.

Complete Genome Sequencing Analysis of Deinococcus wulumuqiensis R12, an Extremely Radiation-Resistant Strain.

Genome sequencing was performed by the PacBio RS II platform and Illumina HiSeq 4000 platform to discover the metabolic profile of the Deinococcus wulumuqiensis R12, which was isolated from radiation-contaminated soils in Xinjiang Uygur Autonomous Region of northwest China. The genome of 3.5 Mbp comprises one circular chromosome and four circular plasmids with 3679 genes and a GC content of 66.97%. A total of 41 new transcriptional factors were identified using the DeepTFactor tool. Genomic analysis revealed the presence of genes for homologous recombination repair, which suggested high recombination efficiency in R12. Three Type I and one Type II RM systems, two CRISPR arrays, and one Cas-Type IC protein were found, allowing the development of endogenous CRISPR-Cas gene-editing tools. Additionally, we found that R12 has a broad spectrum of substrate utilization, which was validated by physiological experiments. Genes involved in the carotenoid biosynthesis pathway and the antioxidative system were also identified. Overall, the comprehensive description of the genome of R12 will facilitate the additional exploitation of this strain as a versatile cell factory for biotechnological applications.