G-site residue S67 is involved in the fungicide-degrading activity of a tau class glutathione S-transferase from Carica papaya.

Thiram is a toxic fungicide extensively used for the management of pathogens in fruits. Although it is known that thiram degrades in plant tissues, the key enzymes involved in this process remain unexplored. In this study, we report that a tau class glutathione S-transferase (GST) from Carica papaya can degrade thiram. This enzyme was easily obtained by heterologous expression in Escherichia coli, showed low promiscuity toward other thiuram disulfides, and catalyzed thiram degradation under physiological reaction conditions. Site-directed mutagenesis indicated that G-site residue S67 shows a key influence for the enzymatic activity toward thiram, while mutation of residue S13, which reduced the GSH oxidase activity, did not significantly affect the thiram-degrading activity. The formation of dimethyl dithiocarbamate, which was subsequently converted into carbon disulfide, and dimethyl dithiocarbamoylsulfenic acid as the thiram degradation products suggested that thiram undergoes an alkaline hydrolysis that involves the rupture of the disulfide bond. Application of the GST selective inhibitor 4-chloro-7-nitro-2,1,3-benzoxadiazole reduced papaya peel thiram-degrading activity by 95%, indicating that this is the main degradation route of thiram in papaya. GST from Carica papaya also catalyzed the degradation of the fungicides chlorothalonil and thiabendazole, with residue S67 showing again a key influence for the enzymatic activity. These results fill an important knowledge gap in understanding the catalytic promiscuity of plant GSTs and reveal new insights into the fate and degradation products of thiram in fruits.

A PilZ domain protein interacts with the transcriptional regulator HinK to regulate type VI secretion system in Pseudomonas aeruginosa.

Type VI secretion systems (T6SS) are bacterial macromolecular complexes that secrete effectors into target cells or the extracellular environment, leading to the demise of adjacent cells and providing a survival advantage. Although studies have shown that the T6SS in Pseudomonas aeruginosa is regulated by the Quorum Sensing system and second messenger c-di-GMP, the underlying molecular mechanism remains largely unknown. In this study, we discovered that the c-di-GMP-binding adaptor protein PA0012 has a repressive effect on the expression of the T6SS HSI-I genes in P. aeruginosa PAO1. To probe the mechanism by which PA0012 (renamed TssZ, Type Six Secretion System -associated PilZ protein) regulates the expression of HSI-I genes, we conducted yeast two-hybrid screening and identified HinK, a LasR-type transcriptional regulator, as the binding partner of TssZ. The protein-protein interaction between HinK and TssZ was confirmed through co-immunoprecipitation assays. Further analysis suggested that the HinK-TssZ interaction was weakened at high c-di-GMP concentrations, contrary to the current paradigm wherein c-di-GMP enhances the interaction between PilZ proteins and their partners. Electrophoretic mobility shift assays revealed that the non-c-di-GMP-binding mutant TssZR5A/R9A interacts directly with HinK and prevents it from binding to the promoter of the quorum-sensing regulator pqsR. The functional connection between TssZ and HinK is further supported by observations that TssZ and HinK impact the swarming motility, pyocyanin production, and T6SS-mediated bacterial killing activity of P. aeruginosa in a PqsR-dependent manner. Together, these results unveil a novel regulatory mechanism wherein TssZ functions as an inhibitor that interacts with HinK to control gene expression.

Research on multi-dimensional reconstruction mechanism of cloud native full link in the metaverse scenario.

Recently, the Microservices whole link whole life cycle optimization framework has been a research hotspot in the academic and engineering fields, especially how to encapsulate the component security mechanism and conduct high concurrency testing based on the existing active framework. The whole link delay, load, and cost have always been the key optimization goals of Microservices splitting and deployment logic. Based on this, this paper proposes a multi-objective optimization Microservices framework that takes into account the security mechanism, Define the fitness function, define the upper and lower limits, and perform multi-dimensional constraints to filter for global and local optima. At the same time, reconstruct the circuit breaker current limiting mechanism, dynamically detect protocol parameters, and perceive the logical relationship of heartbeat status in real-time. Experimental results have shown that this innovative framework can solve scenarios such as high concurrency, high reliability, and high availability Performance bottleneck, business degradation, especially in financial and securities scenarios.

Heat-killed Lactobacillus murinus confers neuroprotection against dopamine neuronal loss by targeting NLRP3 inflammasome.

The intestinal flora has become very active in studies related to Parkinson's disease (PD) in recent years. The microbe-gut-brain axis is closely related to the maintenance of brain homeostasis as well as PD pathogenesis. Alterations in gut bacteria can contribute to neuroinflammation and dopamine (DA) neurodegeneration. Lactobacillus murinus, a gram-positive bacterium, is a commensal gut bacteria present in the mammalian gut and considered as a potential probiotic due to its beneficial effects, including anti-inflammatory and antibacterial actions. In this study, the effects of live L. murinus and heat-killed L. murinus on DA neuronal damage in rats and the underlying mechanisms were investigated. Data showed that heat-killed L. murinus ameliorated 6-hydroxydopamine-induced motor dysfunctions and loss of substantia nigra DA neurons, while no protection was shown in live L. murinus treatment. At the same time, heat-killed L. murinus reduced the activation of NLRP3 inflammasome in microglia and the secretion of pro-inflammatory factors, thus inhibiting the development of neuroinflammation. Furthermore, heat-killed L. murinus failed to display its original neuroprotective properties in NLRP3 inflammasome knockout mice. Together, heat-killed L. murinus conferred neuroprotection against DA neuronal loss via the inhibition of microglial NLRP3 inflammasome activation. These findings provide a promising potential for future applications of L. murinus, and also beneficial strategy for PD treatment.

Effects of levodopa on gut bacterial antibiotic resistance in Parkinson's disease rat.

The second most prevalent neurodegenerative ailment, Parkinson's disease (PD), is characterized by both motor and non-motor symptoms. Levodopa is the backbone of treatment for PD at the moment. However, levodopa-induced side effects, such as dyskinesia, are commonly seen in PD patients. Recently, several antibiotics were found to present neuroprotective properties against neurodegenerative and neuro-inflammatory processes, which might be developed to effective therapies against PD. In this study, we aimed to identify if levodopa treatment could influence the gut bacterial antibiotic resistance in PD rat. Fecal samples were collected from healthy rats and 6-OHDA induced PD rats treated with different doses of levodopa, metagenomic sequencing data showed that levodopa resulted in gut bacteria composition change, the biomarkers of gut bacteria analyzed by LEfSe changed as well. More interestingly, compared with levodopa (5 mg/kg)-treated or no levodopa-treated PD rats, levodopa (10 mg/kg) caused a significant decrease in the abundance of tetW and vanTG genes in intestinal bacteria, which were related to tetracycline and vancomycin resistance, while the abundance of AAC6-lb-Suzhou gene increased apparently, which was related to aminoglycosides resistance, even though the total quantity of Antibiotic Resistance Gene (ARG) and Antibiotic Resistance Ontology (ARO) among all groups did not significantly differ. Consequently, our results imply that the combination of levodopa and antibiotics, such as tetracycline and vancomycin, in the treatment of PD may decrease the amount of corresponding antibiotic resistance genes in gut bacteria, which would give a theoretical basis for treating PD with levodopa combined with tetracycline and vancomycin in the future.

Relationship Between Gut Bacteria and Levodopa Metabolism.

Parkinson's disease (PD) is one of the most common neurodegenerative diseases, characterized by the reduction of dopamine neurons in the substantia nigra. Levodopa, as a dopamine supplement, is the gold-standard therapeutic drug for PD. The metabolism of levodopa in the periphery not only decreases its bioavailability but also affects its efficacy. Thus, it is necessary to investigate how levodopa is metabolized. A growing number of studies have shown that intestinal bacteria, such as Enterococcus faecalis, Eggerthella lenta and Clostridium sporogenes, could metabolize levodopa in different ways. In addition, several pathways to reduce levodopa metabolism by gut microbiota were confirmed to improve levodopa efficacy. These pathways include aromatic amino acid decarboxylase (AADC) inhibitors, antibiotics, pH and (S)-α-fluoromethyltyrosine (AFMT). In this review, we have summarized the metabolic process of levodopa by intestinal bacteria and analyzed potential approaches to reduce the metabolism of levodopa by gut microbiota, thus improving the efficacy of levodopa.

Insights into the roles of bacterial infection and antibiotics in Parkinson's disease.

Parkinson's disease (PD) is one of the most common neurodegenerative disorders, which is accompanied with the classical motor symptoms and a range of non-motor symptoms. Bacterial infection affects the neuroinflammation associated with the pathology of PD and various antibiotics have also been confirmed to play an important role not only in bacterial infection, but also in the PD progression. This mini-review summarized the role of common bacterial infection in PD and introduced several antibiotics that had anti-PD effects.

New hope for Parkinson's disease treatment: Targeting gut microbiota.

There might be more than 10 million confirmed cases of Parkinson's disease (PD) worldwide by 2040. However, the pathogenesis of PD is still unclear. Host health is closely related to gut microbiota, which are affected by factors such as age, diet, and exercise. Recent studies have found that gut microbiota may play key roles in the progression of a wide range of diseases, including PD. Changes in the abundance of gut bacteria, such as Helicobacter pylori, Enterococcus faecalis, and Desulfovibrio, might be involved in PD pathogenesis or interfere with PD therapy. Gut microbiota and the distal brain achieve action on each other through a gut-brain axis composed of the nervous system, endocrine system, and immune system. Here, this review focused on the current understanding of the connection between Parkinson's disease and gut microbiota, to provide potential therapeutic targets for PD.

Resveratrol Alleviates Levodopa-Induced Dyskinesia in Rats.

Dyskinesia is a serious complication of Parkinson's disease during levodopa (L-DOPA) treatment. The pathophysiology of L-DOPA-induced dyskinesia (LID) is complex and not fully illuminated. At present, treatment of dyskinesia is quite limited. Recent studies demonstrated neuroinflammation plays an important role in development of LID. Thus, inhibition of neuroinflammation might open a new avenue for LID treatment. Resveratrol (RES) is the most well-known polyphenolic stilbenoid and verified to possess a large variety of biological activities. DA neurotoxicity was assessed via behavior test and DA neuronal quantification. The movement disorders of dyskinesia were detected by the abnormal involuntary movements scores analysis. Effects of RES on glial cells-elicited neuroinflammation were also explored. Data showed that RES attenuated dyskinesia induced by L-DOPA without affecting L-DOPA's anti-parkinsonian effects. Furthermore, RES generated neuroprotection against long term treatment of L-DOPA-induced DA neuronal damage. Meanwhile, RES reduced protein expression of dyskinesia molecular markers, ΔFOS B and ERK, in the striatum. Also, there was a strong negative correlation between DA system damage and ΔFOS B level in the striatum. In addition, RES inhibited microglia and astroglia activation in substantia nigra and subsequent inflammatory responses in the striatum during L-DOPA treatment. RES alleviates dyskinesia induced by L-DOPA and these beneficial effects are closely associated with protection against DA neuronal damage and inhibition of glial cells-mediated neuroinflammatory reactions.

Ellagic Acid Protects Dopamine Neurons via Inhibition of NLRP3 Inflammasome Activation in Microglia.

Neuroinflammation plays a crucial role in the pathological process of Parkinson's disease (PD). Nod-like receptor protein 3 (NLRP3) inflammasome was highly located in microglia and involved in the process of neuroinflammation. Activation of the NLRP3 inflammasome has been confirmed to contribute to the progression of PD. Thus, inhibition of NLRP3 inflammasome activation could be an important breakthrough point on PD therapy. Ellagic acid (EA) is a natural polyphenol that has been widely found in soft fruits, nuts, and other plant tissues with anti-inflammatory, antioxidant, and neuroprotective properties. However, the mechanisms underlying EA-mediated anti-inflammation and neuroprotection have not been fully elucidated. In this study, a lipopolysaccharide- (LPS-) induced rat dopamine (DA) neuronal damage model was performed to determine the effects of EA on the protection of DA neurons. In addition, the DA neuronal MN9D cell line and microglial BV-2 cell line were employed to explore whether EA-mediated neuroprotection was through an NLRP3-dependent mechanism. Results indicated that EA ameliorated LPS-induced DA neuronal loss in the rat substantia nigra. Further, inhibition of microglial NLRP3 inflammasome signaling activation was involved in EA-generated neuroprotection, as evidenced by the following observations. First, EA reduced NLRP3 inflammasome signaling activation in microglia and subsequent proinflammatory cytokines' excretion. Second, EA-mediated antineuroinflammation and further DA neuroprotection from LPS-induced neurotoxicity were not shown upon microglial NLRP3 siRNA treatment. In conclusion, this study demonstrated that EA has a profound effect on protecting DA neurons against LPS-induced neurotoxicity via the suppression of microglial NLRP3 inflammasome activation.

Ellagic acid protects dopamine neurons from rotenone-induced neurotoxicity via activation of Nrf2 signalling.

Parkinson's disease (PD) is the second most prevalent central nervous system (CNS) degenerative disease. Oxidative stress is one of key contributors to PD. Nuclear factor erythroid-2-related factor 2 (Nrf2) is considered to be a master regulator of many genes involved in anti-oxidant stress to attenuate cell death. Therefore, activation of Nrf2 signalling provides an effective avenue to treat PD. Ellagic acid (EA), a natural polyphenolic contained in fruits and nuts, possesses amounts of pharmacological activities, such as anti-oxidant stress and anti-inflammation. Recent studies have confirmed EA could be used as a neuroprotective agent in neurodegenerative diseases. Here, mice subcutaneous injection of rotenone (ROT)-induced DA neuronal damage was performed to investigate EA-mediated neuroprotection. In addition, adult Nrf2 knockout mice and different cell cultures including MN9D-enciched, MN9D-BV-2 and MN9D-C6 cell co-cultures were applied to explore the underlying mechanisms. Results demonstrated EA conferred neuroprotection against ROT-induced DA neurotoxicity. Activation of Nrf2 signalling was involved in EA-mediated DA neuroprotection, as evidenced by the following observations. First, EA activated Nrf2 signalling in ROT-induced DA neuronal damage. Second, EA generated neuroprotection with the presence of astroglia and silence of Nrf2 in astroglia abolished EA-mediated neuroprotection. Third, EA failed to produce DA neuroprotection in Nrf2 knockout mice. In conclusion, this study identified EA protected against DA neuronal loss via an Nrf2-dependent manner.

Regulation of flagellar motor switching by c-di-GMP phosphodiesterases in Pseudomonas aeruginosa.

The second messenger cyclic diguanylate (c-di-GMP) plays a prominent role in regulating flagellum-dependent motility in the single-flagellated pathogenic bacterium Pseudomonas aeruginosa The c-di-GMP-mediated signaling pathways and mechanisms that control flagellar output remain to be fully unveiled. Studying surface-tethered and free-swimming P. aeruginosa PAO1 cells, we found that the overexpression of an exogenous diguanylate cyclase (DGC) raises the global cellular c-di-GMP concentration and thereby inhibits flagellar motor switching and decreases motor speed, reducing swimming speed and reversal frequency, respectively. We noted that the inhibiting effect of c-di-GMP on flagellar motor switching, but not motor speed, is exerted through the c-di-GMP-binding adaptor protein MapZ and associated chemotactic pathways. Among the 22 putative c-di-GMP phosphodiesterases, we found that three of them (DipA, NbdA, and RbdA) can significantly inhibit flagellar motor switching and swimming directional reversal in a MapZ-dependent manner. These results disclose a network of c-di-GMP-signaling proteins that regulate chemotactic responses and flagellar motor switching in P. aeruginosa and establish MapZ as a key signaling hub that integrates inputs from different c-di-GMP-signaling pathways to control flagellar output and bacterial motility. We rationalized these experimental findings by invoking a model that postulates the regulation of flagellar motor switching by subcellular c-di-GMP pools.

The MapZ-Mediated Methylation of Chemoreceptors Contributes to Pathogenicity of Pseudomonas aeruginosa.

The pathogenic bacterium Pseudomonas aeruginosa is notorious for causing acute and chronic infections in humans. The ability to infect host by P. aeruginosa is dependent on a complex cellular signaling network, which includes a large number of chemosensory signaling pathways that rely on the methyl-accepting chemotaxis proteins (MCPs). We previously found that the second messenger c-di-GMP-binding adaptor MapZ modulates the methylation of an amino acid-detecting MCP by directly interacting with a chemotaxis methyltransferase CheR1. The current study further expands our understanding of the role of MapZ in regulating chemosensory pathways by demonstrating that MapZ suppresses the methylation of multiple MCPs in P. aeruginosa PAO1. The MCPs under the control of MapZ include five MCPs (Aer, CtpH, CptM, PctA, and PctB) for detecting oxygen/energy, inorganic phosphate, malate and amino acids, and three MCPs (PA1251, PA1608, and PA2867) for detecting unknown chemoattractant or chemorepellent. Chemotaxis assays showed that overexpression of MapZ hampered the taxis of P. aeruginosa toward chemoattractants and scratch-wounded human cells. Mouse infection experiments demonstrated that a dysfunction in MapZ regulation had a profound negative impact on the dissemination of P. aeruginosa and resulted in attenuated bacterial virulence. Together, the results imply that by controlling the methylation of various MCPs via the adaptor protein MapZ, c-di-GMP exerts a profound influence on chemotactic responses and bacterial pathogenesis.

Tunable dual-core liquid-filled photonic crystal fibers for dispersion compensation.

We have theoretically investigated the dispersion characteristics of dispersion compensating fibers based on dual-core liquid-filled PCFs. A very high negative chromatic dispersion value D = -19000 ps/(nm-km) can be achieved at 1.55-microm wavelength by an appropriate design. By varying the geometry of the PCF and the index of the filling liquid, the phase-matching wavelength and dispersion values are shown to be well tuned to desired values. The proposed structure also demonstrates good tunable properties with operation temperature for optical communication systems.