Gga-miR-200a-3p suppresses avian reovirus-induced apoptosis and viral replication via targeting GRB2.

Avian reovirus (ARV) is a significant pathogen that causes various clinical diseases in chickens, including viral arthritis, chronic respiratory diseases, retarded growth, and malabsorption syndrome. These conditions result in substantial economic losses for the global poultry industry. MicroRNAs (miRNAs), a type of small noncoding RNAs that regulate gene expression post transcriptionally by silencing or degrading their RNA targets, play crucial roles in response to pathogenic infections. In this study, transfection of DF-1 cells with gga-miR-200a-3p, an upregulated miRNA observed in ARV-infected cells, significantly suppressed ARV-induced apoptosis by directly targeting GRB2 and impeded ARV replication. Conversely, knockdown of endogenous gga-miR-200a-3p in DF-1 cells using a specific miRNA inhibitor enhanced ARV-induced apoptosis and promoted GRB2 expression, thereby facilitating viral growth within cells. Consistently, inhibition of GRB2 activity through siRNA-mediated knockdown reduced viral titers. Therefore, gga-miR-200a-3p plays a vital antiviral role in the host response to ARV infection by suppressing apoptosis via direct targeting of GRB2 protein. This information enhances our understanding of the mechanisms by which host cells combat against ARV infection through self-encoded small RNA molecules and expands our knowledge regarding the involvement of microRNAs in the host response to pathogenic infections.

Cultivation of phosphate-accumulating biofilm: Study of the effects of acyl-homoserine lactones (AHLs) and cyclic dimeric guanosine monophosphate (c-di-GMP) on the formation of biofilm and the enhancement of phosphate metabolism capacity.

This study investigated the mechanisms of microbial growth and metabolism during biofilm cultivation in the biofilm sequencing batch reactor (BSBR) process for phosphate (P) enrichment. The results showed that the sludge discharge was key to biofilm growth, as it terminated the competition for carbon (C) source between the nascent biofilm and the activated sludge. For the tested reactor, after the sludge discharge on 18 d, P metabolism and C source utilization improved significantly, and the biofilm grew rapidly. The P concentration of the recovery liquid reached up to 157.08 mg/L, which was sufficient for further P recovery via mineralization. Meta-omics methods were used to analyze metabolic pathways and functional genes in microbial growth during biofilm cultivation. It appeared that the sludge discharge activated the key genes of P metabolism and inhibited the key genes of C metabolism, which strengthened the polyphosphate-accumulating metabolism (PAM) as a result. The sludge discharge not only changed the types of polyphosphate-accumulating organisms (PAOs) but also promoted the growth of dominant PAOs. Before the sludge discharge, the necessary metabolic abilities that were spread among different microorganisms gradually concentrated into a small number of PAOs, and after the sludge discharge, they further concentrated into Candidatus_Contendobacter (P3) and Candidatus_Accumulibacter (P17). The messenger molecule C-di-GMP, produced mostly by P3 and P17, facilitated P enrichment by regulating cellular P and C metabolism. The glycogen-accumulating organism (GAO) Candidatus_Competibacter secreted N-Acyl homoserine lactones (AHLs), which stimulated the secretion of protein in extracellular polymeric substances (EPS), thus promoting the adhesion of microorganisms to biofilm and improving P metabolism via EPS-based P adsorption. Under the combined action of the dominant GAOs and PAOs, AHLs and C-di-GMP mediated QS to promote biofilm development and P enrichment. The research provides theoretical support for the cultivation of biofilm and its wider application.

Development of a platform method for rapid detection and characterization of domain-specific post-translational modifications in bispecific antibodies.

Charge heterogeneity is inherent to all therapeutic antibodies and arises from post-translational modifications (PTMs) and/or protein degradation events that may occur during manufacturing. Among therapeutic antibodies, the bispecific antibody (bsAb) containing two unique Fab arms directed against two different targets presents an additional layer of complexity to the charge profile. In the context of a bsAb, a single domain-specific PTM within one of the Fab domains may be sufficient to compromise target binding and could potentially impact the stability, safety, potency, and efficacy of the drug product. Therefore, characterization and routine monitoring of domain-specific modifications is critical to ensure the quality of therapeutic bispecific antibody products. We developed a Digestion-assisted imaged Capillary isoElectric focusing (DiCE) method to detect and quantitate domain-specific charge variants of therapeutic bispecific antibodies (bsAbs). The method involves enzymatic digestion using immunoglobulin G (IgG)-degrading enzyme of S. pyogenes (IdeS) to generate F(ab)2 and Fc fragments, followed by imaged capillary isoelectric focusing (icIEF) under reduced, denaturing conditions to separate the light chains (LCs) from the Fd domains. Our results suggest that DiCE is a highly sensitive method that is capable of quantitating domain-specific PTMs of a bsAb. In one case study, DiCE was used to quantitate unprocessed C-terminal lysine and site-specific glycation of Lys98 in the complementarity-determining region (CDR) of a bsAb that could not be accurately quantitated using conventional, platform-based charge variant analysis, such as intact icIEF. Quantitation of these PTMs by DiCE was comparable to results from peptide mapping, demonstrating that DiCE is a valuable orthogonal method for ensuring product quality. This method may also have potential applications for characterizing fusion proteins, antibody-drug conjugates, and co-formulated antibody cocktails.

Neuroinflammatory response on a newly combinatorial cell-cell interaction chip.

Neuroinflammation is a common feature in various neurological disorders. Understanding neuroinflammation and neuro-immune interactions is of significant importance. However, the intercellular interactions in the inflammatory model are intricate. Microfluidic chips, with their complex micrometer-scale structures and real-time observation capabilities, offer unique advantages in tackling these complexities compared to other techniques. In this study, microfluidic chip technology was used to construct a microarray physical barrier structure with 15 µm spacing, providing well-defined cell growth areas and clearly delineated interaction channels. Moreover, an innovative hydrophilic treatment process on the glass surface facilitated long-term co-culture of cells. The developed neuroinflammation model on the chip revealed that SH-SY5Y cytotoxicity was predominantly influenced by co-cultured THP-1 cells. The co-culture model fostered complex interactions that may exacerbate cytotoxicity, including irregular morphological changes of cells, cell viability reduction, THP-1 cell migration, and the release of inflammatory factors. The integration of the combinatorial cell-cell interaction chip not only offers a clear imaging detection platform but also provides diverse data on cell migration distance, migration direction, and migration angle. Furthermore, the designed ample space for cell culture, along with microscale channels with fluid characteristics, allow for the study of inflammatory factor distribution patterns on the chip, offering vital theoretical data on biological relevance that conventional experiments cannot achieve. The fabricated user-friendly, reusable, and durable co-culture chip serves as a valuable in vitro tool, providing an intuitive platform for gaining insights into the complex mechanisms underlying neuroinflammation and other interacting models.

A Novel Perspective on PCSK9 in Alzheimer's Disease: A Focus on Amyloid Beta.

The proprotein convertase subtilisin/kexin type 9 (PCSK9) belongs to a member of the proprotein convertase (PC) family, which is mainly secreted by the liver and plays a central role in lipid metabolism. Furthermore, PCSK9 plays a multifunctional role in promoting the inflammatory response, inducing cell apoptosis and pyroptosis and affecting tumor homeostasis. The brain is the organ with the richest lipid content. Incidentally, PCSK9 increased in many brain diseases, including brain injury and Alzheimer's disease (AD). Consequently, the relationship between PCSK9 and brain diseases has attracted increasing research interest. Amyloid beta (Aß) accumulation is the central and initial event in the pathogenesis of AD. This study focuses on the effects of PCSK9 on Aß accumulation in the brain via multiple modalities to explore the potential role of PCSK9 in AD, which is characterized by progressive loss of brain cells by increasing Aß accumulation. The study also explores the new mechanism by which PCSK9 is involved in the pathogenesis of AD, providing interesting and innovative guidance for the future of PCSK9-targeted therapy for AD.

[Characterization and identification of chemical constituents from Schizonepetae Spica based on UHPLC-Q-TOF-MS/MS technique].

The chemical constituents of Schizonepetae Spica were qualitatively analyzed by UHPLC-Q-TOF-MS/MS. An Agilent poroshell 120 SB-C_(18) column(3.0 mm×100 mm, 2.7 µm) was used for gradient elution with 0.1% formic acid water(A)-acetonitrile(B) solution as mobile phase at the flow rate of 0.4 mL·min~(-1) and column temperature of 45 ℃. The data were collected by scanning in positive and negative ion modes, and the compounds were identified by comparison of reference materials and PeakView software. Ninety-seven compounds were identified from Schizonepetae Spica, including 28 flavonoids, 23 phenolic acids, 23 fatty acids, 15 terpenoids, and 8 other compounds. The UHPLC-Q-TOF-MS/MS method established in this study can identify the chemical components of Schizonepetae Spica rapidly, accurately, and comprehensively, and provide a basis for the basic study of pharmacodynamic substances of Schizonepetae Spica.

Piperine improves levodopa availability in the 6-OHDA-lesioned rat model of Parkinson's disease by suppressing gut bacterial tyrosine decarboxylase.

AIM: Tyrosine decarboxylase (TDC) presented in the gut-associated strain Enterococcus faecalis can convert levodopa (L-dopa) into dopamine (DA), and its increased abundance would potentially minimize the availability and efficacy of L-dopa. However, the known human decarboxylase inhibitors are ineffective in this bacteria-mediated conversion. This study aims to investigate the inhibition of piperine (PIP) on L-dopa bacterial metabolism and evaluates the synergistic effect of PIP combined with L-dopa on Parkinson's disease (PD). METHODS: Metagenomics sequencing was adopted to determine the regulation of PIP on rat intestinal microbiota structure, especially on the relative abundance of E. faecalis. Then, the inhibitory effects of PIP on L-dopa conversion and TDC expression of E. faecalis were tested in vitro. We examined the synergetic effect of the combination of L-dopa and PIP on 6-hydroxydopamine (6-OHDA)-lesioned rats and tested the regulations of L-dopa bioavailability and brain DA level by pharmacokinetics study and MALDI-MS imaging. Finally, we evaluated the microbiota-dependent improvement effect of PIP on L-dopa availability using pseudo-germ-free and E. faecalis-transplanted rats. RESULTS: We found that PIP combined with L-dopa could better ameliorate the move disorders of 6-OHDA-lesioned rats by remarkably improving L-dopa availability and brain DA level than L-dopa alone, which was associated with the effect of PIP on suppressing the bacterial decarboxylation of L-dopa via effectively downregulating the abnormal high abundances of E. faecalis and TDC in 6-OHDA-lesioned rats. CONCLUSION: Oral administration of L-dopa combined with PIP can improve L-dopa availability and brain DA level in 6-OHDA-lesioned rats by suppressing intestinal bacterial TDC.

Piperine promotes PI3K/AKT/mTOR-mediated gut-brain autophagy to degrade α-Synuclein in Parkinson's disease rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Piper longum L., a medicinal and food homologous herb, has a traditional history of use in treating gastrointestinal and neurological disorders. Piperine (PIP) the main alkaloid of P. longum, exists neuroprotective effects on various animal models of Parkinson's disease (PD). Nevertheless, the underlying mechanism, particularly the role of PIP in promoting gut-brain autophagy for α-Synuclein (α-Syn) degradation in PD, remains incompletely understood. AIM OF THE STUDY: To explore the role of PIP in regulating the gut-brain autophagy signaling pathway to reduce α-Syn levels in both the colon and substantia nigra (SN) of PD model rats. MATERIALS AND METHODS: Behavioral experiments were conducted to assess the impact of PIP on 6-hydroxydopamine (6-OHDA)-induced PD rats. The intestinal microbiome composition and intestinal metabolites were analyzed by metagenomics and GC-MS/MS. The auto-phagosomes were visualized by transmission electron microscopy. Immunohistochemistry, immunofluorescence, and western blotting were performed to assess the levels of tyrosine hydroxylase (TH), α-Syn, LC3II/LC3I, p62, and the PI3K/AKT/mTOR pathway in both the SN and colon of the rats. The pathway-related inhibitor and agonist were used to verify the autophagy mechanism in the SH-SY5Y cells overexpressing A53T mutant α-Syn (A53T-α-Syn). RESULTS: PIP improved autonomic movement and gastrointestinal dysfunctions, reduced α-Syn aggregation and attenuated the loss of dopaminergic neurons in 6-OHDA-induced PD rats. After oral administration of PIP, the radio of LC3II/LC3I increased and the expression of p62 was degraded, as well as the phosphorylation levels of PI3K, AKT and mTOR decreased in the SN and colon of rats. The effect of PIP on reducing A53T-α-Syn through the activation of the PI3K/AKT/mTOR-mediated autophagy pathway was further confirmed in A53T-α-Syn transgenic SH-SY5Y cells. This effect could be inhibited by the autophagy inhibitor bafilomycin A1 and the PI3K agonist 740 Y-P. CONCLUSIONS: Our findings suggested that PIP could protect neurons by activating autophagy to degrade α-Syn in the SN and colon, which were related to the suppression of PIP on the activation of PI3K/AKT/mTOR signaling pathway.

Using Artificial Intelligence to Gauge Competency on a Novel Laparoscopic Training System.

OBJECTIVE: Laparoscopic surgical skill assessment and machine learning are often inaccessible to low-and-middle-income countries (LMIC). Our team developed a low-cost laparoscopic training system to teach and assess psychomotor skills required in laparoscopic salpingostomy in LMICs. We performed video review using AI to assess global surgical techniques. The objective of this study was to assess the validity of artificial intelligence (AI) generated scoring measures of laparoscopic simulation videos by comparing the accuracy of AI results to human-generated scores. DESIGN: Seventy-four surgical simulation videos were collected and graded by human participants using a modified OSATS (Objective Structured Assessment of Technical Skills). The videos were then analyzed via AI using 3 different time and distance-based calculations of the laparoscopic instruments including path length, dimensionless jerk, and standard deviation of tool position. Predicted scores were generated using 5-fold cross validation and K-Nearest-Neighbors to train classifiers. SETTING: Surgical novices and experts from a variety of hospitals in Ethiopia, Cameroon, Kenya, and the United States contributed 74 laparoscopic salpingostomy simulation videos. RESULTS: Complete accuracy of AI compared to human assessment ranged from 65-77%. There were no statistical differences in rank mean scores for 3 domains, Flow of Operation, Respect for Tissue, and Economy of Motion, while there were significant differences in ratings for Instrument Handling, Overall Performance, and the total summed score of all 5 domains (Summed). Estimated effect sizes were all less than 0.11, indicating very small practical effect. Estimated intraclass correlation coefficient (ICC) of Summed was 0.72 indicating moderate correlation between AI and Human scores. CONCLUSIONS: Video review using AI technology of global characteristics was similar to that of human review in our laparoscopic training system. Machine learning may help fill an educational gap in LMICs where direct apprenticeship may not be feasible.

Structural insights into the allosteric inhibition of P2X4 receptors.

P2X receptors are ATP-activated cation channels, and the P2X4 subtype plays important roles in the immune system and the central nervous system, particularly in neuropathic pain. Therefore, P2X4 receptors are of increasing interest as potential drug targets. Here, we report the cryo-EM structures of the zebrafish P2X4 receptor in complex with two P2X4 subtype-specific antagonists, BX430 and BAY-1797. Both antagonists bind to the same allosteric site located at the subunit interface at the top of the extracellular domain. Structure-based mutational analysis by electrophysiology identified the important residues for the allosteric inhibition of both zebrafish and human P2X4 receptors. Structural comparison revealed the ligand-dependent structural rearrangement of the binding pocket to stabilize the binding of allosteric modulators, which in turn would prevent the structural changes of the extracellular domain associated with channel activation. Furthermore, comparison with the previously reported P2X structures of other subtypes provided mechanistic insights into subtype-specific allosteric inhibition.

[Chemical constituents from stems and leaves of Cratoxylum cochinchinense and their inhibitory effects on proliferation of synoviocytes in vitro].

The chemical constituents from the stems and leaves of Cratoxylum cochinchinense were isolated and purified using silica gel, ODS gel, and Sephadex LH-20 gel column chromatography, as well as preparative HPLC. The chemical structures of all isolated compounds were identified on the basis of their physicochemical properties, spectroscopic analyses, and the comparison of their physicochemical and spectroscopic data with the reported data in literature. As a result, 21 compounds were isolated from the 90% ethanol extract of the stems and leaves of C. cochinchinense, which were identified as cratocochine(1), 1-hydroxy-3,7-dimethoxyxanthone(2), 1-hydroxy-5,6,7-trimethoxyxanthone(3), ferrxanthone(4), 3,6-dihydroxy-1,5-dimethoxyxanthone(5), 3,6-dihydroxy-1,7-dimethoxyxanthone(6), 1,2,5-trihydroxy-6,8-dimethoxyxanthone(7), securixanthone G(8), gentisein(9), 3,7-dihydroxy-1-methoxyxanthone(10), pancixanthone B(11), garcimangosxanthone A(12), pruniflorone L(13), 9-hydroxy alabaxanthone(14), cochinchinone A(15), luteolin(16), 3,5'-dimethoxy-4',7-epoxy-8,3'-neolignane-5,9,9'-triol(17), N-benzyl-9-oxo-10E,12E-octadecadienamide(18), 15-hydroxy-7,13E-labdadiene(19), stigmasta-4,22-dien-3-one(20), and stigmast-5-en-3ß-ol(21). Among these isolates, compound 1 was a new xanthone, compounds 2-5, 7, 8, 12, and 16-21 were isolated from the Cratoxylum plant for the first time, and compounds 11 and 13 were obtained from C. cochinchinense for the first time. Furthermore, all isolated compounds 1-21 were appraised for their anti-rheumatoid arthritis activities by MTS method through measuring their anti-proliferative effect on synoviocytes in vitro. As a result, xanthones 1-15 displayed notable anti-rheumatoid arthritis activities, which showed inhibitory effects on the proliferation of MH7A synoviocytes with the IC_(50) values ranging from(8.98±0.12) to(228.68±0.32) µmol·L~(-1).

Psychological interventions for the prevention of depression relapse: systematic review and network meta-analysis.

Depression is highly prevalent and easily relapses. Psychological interventions are effective for the prevention of depression relapse. This systematic review and network meta-analysis aimed to compare the efficacy at the same follow-up time points of psychological interventions in depression. We searched PubMed, Embase, and PsycINFO via OVID, and the Cochrane Library published up to December 12, 2021, and PubMed up to July 1, 2022. The primary outcome was depression relapse, considering the same time points that were extracted on survival curves or relapse curves. The study protocol was registered with PROSPERO, CRD42022343327. A total of 2,871 patients were included from 25 RCTs. Mindfulness-based cognitive therapy (MBCT) was significantly better than placebo at the 3 months, the 6 months, and the 9 months at follow-up. Cognitive behavioral therapy (CBT) was significantly better than treatment as usual at the 3 months, the 9 months, the 12 months, and the 15 months at follow-up. CBT was significantly better than placebo at the 21 months and the 24 months at follow-up. Behavioral activation therapy was significantly better than placebo at the 21 months and the 24 months at follow-up. Interpersonal psychotherapy was significantly better than placebo at the 24-month follow-up. All psychological interventions included in the study were significantly better than supportive counseling most of the time. The results were robust in various sensitivity and subgroup analyses. In conclusion, MBCT had a continuous effect in preventing relapse of depression. CBT had the longest but not continuous effect in preventing relapse of depression. The effects of behavioral activation therapy and interpersonal therapy for the prevention of depression appeared late. All psychological interventions included in the study were more effective than supportive counseling. More evidence is needed from large comparative trials that provide long-term follow-up data.

Xanthones with Potential Anti-Inflammatory and Anti-HIV Effects from the Stems and Leaves of Cratoxylum cochinchinense.

Four new xanthones, cratocochinones A-D (1-4), together with eight known analogues (5-12), were isolated from the stems and leaves of Cratoxylum cochinchinense. The chemical structures of cratocochinones A-D (1-4) were elucidated by comprehensive spectroscopic analyses and the known compounds were identified by comparisons with the spectral data reported in the literature. All isolated compounds 1-12 were evaluated for their anti-inflammatory activities and anti-HIV-1 activities. Compounds 1-12 showed remarkable inhibitory activities on nitric oxide (NO) production induced by lipopolysaccharide in mouse macrophage RAW 264.7 cells in vitro, with IC50 values in the range of 0.86 ± 0.05 to 18.36 ± 0.21 µM. Meanwhile, compounds 1-12 exhibited significant anti-HIV-1 activities with EC50 which ranged from 0.22 to 11.23 µM. These findings indicate that the discoveries of these xanthones, isolated from the stems and leaves of C. cochinchinense, showing significant anti-inflammatory and anti-HIV-1 effects could be of great importance to the research and development of new natural anti-inflammatory and anti-HIV agents.

Investigation of High Molecular Weight Size Variant Formation in Antibody-Drug Conjugates: Microbial Transglutaminase-Mediated Crosslinking.

Microbial transglutaminase (mTG) has become a powerful tool for manufacturing antibody-drug conjugates (ADCs). It enables site-specific conjugation by catalyzing formation of stable isopeptide bond between glutamine (Q) side chain and primary amine. However, the downstream impact of mTG-mediated conjugation on ADC product quality, especially on high molecular weight (HMW) size variant formation has not been studied in a systematic manner. This study investigates the mechanisms underlying the formation of HMW size variants in mTG-mediated ADCs using size exclusion chromatography (SEC) and liquid chromatography-mass spectrometry (LC-MS). Our findings revealed that the mTG-mediated glutamine and lysine (K) crosslinking is the primary source of the increased level of HMW size variants in the ADCs. In the study, two monoclonal antibodies (mAbs) with glutamine engineered for site-specific conjugation were used as model systems. Based on the LC-MS analysis, a single lysine (K56) in the heavy chain (HC) was identified as the major Q-K crosslinking site in one of the two mAbs. The HC C-terminal K was observed to crosslink to the target Q in both mAbs. Quantitative correlation was established between the percentage of HMW size variants determined by SEC and the percentage of crosslinked peptides quantified by MS peptide mapping. Importantly, it was demonstrated that the level of HMW size variants in the second ADC was substantially reduced by the complete removal of HC C-terminal K before conjugation. The current work demonstrates that crosslinking and other side reactions during mTG-mediated conjugation needs to be carefully monitored and controlled to ensure process consistency and high product quality of the final ADC drug product.

Non-oxidative modified low-density lipoproteins: The underappreciated risk factors for atherosclerosis.

Atherosclerosis, the pathological basis of most cardiovascular diseases, is a main risk factor causing about 20 million deaths each year worldwide. Oxidized low-density lipoprotein is recognized as the most important and independent risk factor in initiating and promoting atherosclerosis. Numerous antioxidants are extensively used in clinical practice, but they have no significant effect on reducing the morbidity and mortality of cardiovascular diseases. This finding suggests that researchers should pay more attention to the important role of non-oxidative modified low-density lipoprotein in atherosclerosis with a focus on oxidized low-density lipoprotein. This review briefly summarizes several important non-oxidative modified low-density lipoproteins associated with atherosclerosis, introduces the pathways through which these non-oxidative modified low-density lipoproteins induce the development of atherosclerosis in vivo, and discusses the mechanism of atherogenesis induced by these non-oxidative modified low-density lipoproteins. New therapeutic strategies and potential drug targets are provided for the prevention and treatment of atherosclerotic cardiovascular diseases.

Tektin makes a microtubule a "micropillar".

Inside sperm flagella, there are nine doublet microtubules composed of A and B tubules. In this issue of Cell, Leung et al. and Zhou et al. present high-resolution cryo-EM structures of doublet microtubules from mammalian sperms and show unprecedented structures of the A tubules, which are almost entirely occupied with tektin bundles.

A new insight in enhancing phosphate enrichment in biofilm process: Comparison of the key metabolic pathways in highly-efficient and dominant PAOs based on metagenomics.

The formation of dominant phosphate-accumulating organisms (PAOs) is essential for the high enrichment of phosphate in biofilm sequencing batch reactors (BSBR) for phosphorus recovery. The dominant PAOs in the biofilm process have not been isolated and purified, and the key metabolic pathways that promote the formation of dominant PAOs are still unclear. In this study, four strains of highly-efficient PAOs were obtained by an innovative isolation procedure. The relationship between the abundance of highly-efficient and dominant PAOs and the phosphate removal ability was compared. We found that the abundance of PAOs was positively correlated with the phosphate removal efficiency in vitro pure culture and complex biofilm process. Metagenomics analysis revealed that compared with highly-efficient PAOs cultured in vitro, dominant PAOs in biofilms had unique key metabolic pathways, F-ATPases and N-Acyl homoserine lactones (AHLs). F-ATPases are important for maintaining the proton motive force (PMF) required for the uptake of carbon sources by PAOs, and AHLs are participating in phosphate metabolism through quorum sensing (QS) mediated secretion of extracellular polymeric substance (EPS). The formation of dominant PAOs was promoted by optimizing carbon source uptake and phosphate metabolism. This study revealed that the difficult isolation of dominant PAOs was due to the AHLs-mediated QS, and we identified the key pathways regulating the formation of dominant PAOs in biofilms through genomics analysis. Our findings provide insights in enhancing phosphate enrichment in BSBR by modulating the components of microbial community under the low concentration of carbon source consumption.

Antithetic effects of agonists and antagonists on the structural fluctuations of TRPV1 channel.

Transient receptor potential vanilloid member 1 (TRPV1) is a heat and capsaicin receptor that allows cations to permeate and cause pain. As the molecular basis for temperature sensing, the heat capacity (ΔCp) model [D. E. Clapham, C. Miller, Proc. Natl. Acad. Sci. U.S.A. 108, 19492-19497 (2011).] has been proposed and experimentally supported. Theoretically, heat capacity is proportional to a variance in enthalpy, presumably related to structural fluctuation; however, the fluctuation of TRPV1 has not been directly visualized. In this study, we directly visualized single-molecule structural fluctuations of the TRPV1 channels in a lipid bilayer with the ligands resiniferatoxin (agonist, 1,000 times hotter than capsaicin) and capsazepine (antagonist) by high-speed atomic force microscopy. We observed the structural fluctuations of TRPV1 in an apo state and found that RTX binding enhances structural fluctuations, while CPZ binding suppresses fluctuations. These ligand-dependent differences in structural fluctuation would play a key role in the gating of TRPV1.

Characterization of physicochemical properties, flavor volatiles and phenolic compounds of feijoa fruit varieties.

Thirteen varieties of feijoa (Feijoa sellowiana) fruit were collected and the physical and chemical properties of feijoa peel, flesh, seed, and leaf were analyzed. Large diversities in the physicochemical characteristics and phenolic and volatile composition among various parts and between different varieties of feijoa were observed. Degrees Brix of whole fruits ranged from 10.1 (Anatoki) to 18.0 (No. 2) °Brix. Procyanidin B-type tetramer, procyanidin B-type dimer, and procyanidin C-type trimer had the highest concentrations in all parts and varieties of feijoa. Caffeoyl glucose, dihydroferulic acid 4-O-glucuronide, galloyl glucose, and lariciresinol-sesquilignan were detected in feijoa fruits and leaves. A total of 105 esters, 68 terpenes, 20 alcohols, 31 hydrocarbons, 12 aldehydes, and 11 ketones were related to aromatic attributes of fruits and leaves. Early season and mid-season varieties had larger variations in the chemical properties than late-season varieties. Anatoki, Kakariki, and No.1, have the potential to be developed for attractive flavor and functional properties.

Novel Mg 2+ binding sites in the cytoplasmic domain of the MgtE Mg 2+ channels revealed by X-ray crystal structures.

MgtE is a Mg 2+-selective channel regulated by the intracellular Mg 2+ concentration. MgtE family proteins are highly conserved in all domains of life and contribute to cellular Mg 2+ homeostasis. In humans, mutations in the SLC41 proteins, the eukaryotic counterparts of the bacterial MgtE, are known to be associated with various diseases. The first MgtE structure from a thermophilic bacterium, Thermus thermophilus, revealed that MgtE forms a homodimer consisting of transmembrane and cytoplasmic domains with a plug helix connecting the two and that the cytoplasmic domain possesses multiple Mg 2+ binding sites. Structural and electrophysiological analyses revealed that the dissociation of Mg 2+ ions from the cytoplasmic domain induces structural changes in the cytoplasmic domain, leading to channel opening. Thus, previous works showed the importance of MgtE cytoplasmic Mg 2+ binding sites. Nevertheless, due to the limited structural information on MgtE from different species, the conservation and diversity of the cytoplasmic Mg 2+ binding site in MgtE family proteins remain unclear. Here, we report crystal structures of the Mg 2+-bound MgtE cytoplasmic domains from two different bacterial species, Chryseobacterium hispalense and Clostridiales bacterium, and identify multiple Mg 2+ binding sites, including ones that were not observed in the previous MgtE structure. These structures reveal the conservation and diversity of the cytoplasmic Mg 2+ binding site in the MgtE family proteins.