The ligand binding domain of a type IV pilus chemoreceptor PilJ has a different fold from that of another PilJ-type receptor McpN.

Bacterial chemoreceptors sense the extracellular signals and regulate bacterial motilities, biofilm formation, etc. The periplasmic ligand binding domains of chemoreceptors occur as different structural folds and recognize a diversity of chemical molecules. In Pseudomonas aeruginosa (PAO1), two bacterial chemoreceptors, McpN (PA2788) and PilJ (PA0411), are proposed to both contain a PilJ-like ligand-binding domain (LBD) (Pfam motif PF13675) and involved in nitrate chemotaxis and type IV pilus-mediated motility, respectively. The LBDs of McpN and PilJ consist of 135 and 263 residues, respectively, and share very low sequence identity, suggesting they might occur as different structures. Here, we found that PilJ-LBD folded into an HBM module, the same as the sensor domains of McpS-LBD and TorS-LBD, but it differed from that of McpN-LBD. We also observed a trimer in SEC and AUC and proposed a trimeric model based on the crystal structure. Based on the sequence, we classified the Pfam containing McpN-LBD and PilJ-LBD into three classes: sPilJ (single PilJ) represented by McpN-LBD with only one PilJ domain, dPilJ (dual PilJ) that contained dual PilJ domains, and hPilJ (hybrid PilJ) that comprises of a PilJ domain and another non-PilJ domain. Our work indicates a significant structural difference between the ligand binding domains of PilJ and McpN and will help our further study on both kinds of chemoreceptors.

MicroRNA-148a-3p in pericyte-derived extracellular vesicles improves erectile function in diabetic mice by promoting cavernous neurovascular regeneration.

BACKGROUND: To investigate the regulatory role of microRNA (miR)-148a-3p in mouse corpus cavernous pericyte (MCPs)-derived extracellular vesicles (EVs) in the treatment of diabetes-induced erectile dysfunction (ED). METHODS: Mouse corpus cavernous tissue was used for MCP primary culture and EV isolation. Small-RNA sequencing analysis was performed to assess the type and content of miRs in MCPs-EVs. Four groups of mice were used: control nondiabetic mice and streptozotocin-induced diabetic mice receiving two intracavernous injections (days - 3 and 0) of phosphate buffered saline, MCPs-EVs transfected with reagent control, or MCPs-EVs transfected with a miR-148a-3p inhibitor. miR-148a-3p function in MCPs-EVs was evaluated by tube-formation assay, migration assay, TUNEL assay, intracavernous pressure, immunofluorescence staining, and Western blotting. RESULTS: We extracted EVs from MCPs, and small-RNA sequencing analysis showed miR-148a-3p enrichment in MCPs-EVs. Exogenous MCPs-EV administration effectively promoted mouse cavernous endothelial cell (MCECs) tube formation, migration, and proliferation, and reduced MCECs apoptosis under high-glucose conditions. These effects were significantly attenuated in miR-148a-3p-depleted MCPs-EVs, which were extracted after inhibiting miR-148a-3p expression in MCPs. Repetitive intracavernous injections of MCPs-EVs improved erectile function by inducing cavernous neurovascular regeneration in diabetic mice. Using online bioinformatics databases and luciferase report assays, we predicted that pyruvate dehydrogenase kinase-4 (PDK4) is a potential target gene of miR-148a-3p. CONCLUSIONS: Our findings provide new and reliable evidence that miR-148a-3p in MCPs-EVs significantly enhances cavernous neurovascular regeneration by inhibiting PDK4 expression in diabetic mice.

Bioactive polysaccharides from Momordica charantia as functional ingredients: a review of their extraction, bioactivities, structural-activity relationships, and application prospects.

Momordica charantia L. is a well-known medicine and food homology plant with high pharmaceutical and nutritional values. Polysaccharides are carbohydrate polymers connected by glycosidic bonds, one of the key functional ingredients of M. charantia. Recently, M. charantia polysaccharides (MCPs) have attracted much attention from industries and researchers due to their anti-oxidant, anti-tumor, anti-diabetes, anti-bacteria, immunomodulatory, neuroprotection, and organ protection activities. However, the development and utilization of MCPs-based functional foods and medicines were hindered by the lack of a deeper understanding of the structure-activity relationship (SAR), structural modification, applications, and safety of MCPs. Herein, we provide an overview of the extraction, purification, structural characterization, bioactivities, and mechanisms of MCPs. Besides, SAR, toxicities, application, and influences of the modification associated with bioactivities are spotlighted, and the potential development and future study direction are scrutinized. This review provides knowledge and research underpinnings for the further research and application of MCPs as therapeutic agents and functional food additives.

Photocatalytic Activity of TiO2/g-C3N4 Nanocomposites for Removal of Monochlorophenols from Water.

This research employed g-C3N4 nanosheets in the hydrothermal synthesis of TiO2/g-C3N4 hybrid photocatalysts. The TiO2/g-C3N4 heterojunctions, well-dispersed TiO2 nanoparticles on the g-C3N4 nanosheets, are effective photocatalysts for the degradation of monochlorophenols (MCPs: 2-CP, 3-CP, and 4-CP) which are prominent water contaminants. The removal efficiency of 2-CP and 4-CP reached 87% and 64%, respectively, after treatment of 25 ppm CP solutions with the photocatalyst (40TiO2/g-C3N4, 1 g/L) and irradiation with UV-Vis light. Treatment of CP solutions with g-C3N4 nanosheets or TiO2 alone in conjunction with irradiation gave removal efficiencies lower than 50%, which suggests the two act synergically to enhance the photocatalytic activity of the 40TiO2/g-C3N4 nanocomposite. Superoxide and hydroxyl radicals are key active species produced during CP photodegradation. In addition, the observed nitrogen and Ti3+ defects and oxygen vacancies in the TiO2/g-C3N4 nanocomposites may improve the light-harvesting ability of the composite and assist preventing rapid electron-hole recombination on the surface, enhancing the photocatalytic performance. In addition, interfacial interactions between the MCPs (low polarity) and thermally exfoliated carbon nitride in the TiO2/g-C3N4 nanocomposites may also enhance MCP degradation.

Marine collagen peptides: A novel biomaterial for the healing of oral mucosal ulcers.

The purpose of this study was to analyze the therapeutic effects of marine collagen peptides (MCPs) from tilapia skin on oral mucosal ulcers in a rat model. CCK-8 and wound healing assays were performed in vitro to evaluate proliferation and migration of L929 cells after treatment with MCPs. The effects of MCPs on the healing of oral mucosal ulcers in a rat model were macroscopically and microscopically analyzed in vivo. Results showed that MCPs promoted proliferation and migration of L929 cells. Moreover, 75%MCPs enhanced the ulcer healing process, suppressed inflammatory response and up-regulated the expression levels of vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF). MCPs are potentially used as a new therapeutic strategy for oral mucosal ulceration.

Peritoneal GATA6+ macrophage drives hepatic immunopathogenesis and maintains the Treg cell niche in the liver.

The source of macrophages that contribute to human liver disease remains poorly understood. The purpose of this study is to investigate the functional mechanism of peritoneal macrophages in the development of hepatic immunopathology. By performing the natural infection with the blood fluke Schistosoma japonicum (S. japonicum) and the chemically carbon tetrachloride (CCl4 )-induced liver injured mouse model, we identified the peritoneal cavity as an essential source of hepatic macrophages. Here, we show that a large number of F4/80+ macrophages was accumulated in the peritoneal cavity during liver injury. An unknown source population of macrophages, which highly expressed GATA6 that is specific to peritoneal macrophages, was found to exist in the injured livers. Peritoneal macrophage deletion by injection with clodronate-containing liposomes led to an attenuated hepatic pathology and the inflammatory microenvironment, while adoptive transfer of macrophages into the abdominal cavity, by contrast, results in restoring liver pathology. Importantly, there are set genes of monocyte chemoattractant protein (MCP)-1, -2, and -3 that are highly related to recruit GATA6+ macrophages during S. japonicum infection, while administration of bindarit, a selective inhibitor of MCPs synthesis, dramatically decreased the hepatic expression of GATA6+ macrophages and thus attenuated hepatic pathology. Furthermore, in vivo study showed that peritoneal macrophages promote hepatic immunopathology is dependent on the accumulation of regulatory T cells (Tregs) in the liver. Altogether, these data provide the first clear evidence that GATA6+ peritoneal macrophages play critical roles in both the formation of hepatic immunopathology and the accumulation of Tregs cells.

Chemotactic Responses of Xanthomonas with Different Host Ranges.

Xanthomonas citri pv. citri (Xcc) (X. citri subsp. citri) type A is the causal agent of citrus bacterial canker (CBC) on most Citrus spp. and close relatives. Two narrow-host-range strains of Xcc, Aw and A*, from Florida and Southwest Asia, respectively, infect only Mexican lime (Citrus aurantifolia) and alemow (C. macrophylla). In the initial stage of infection, these xanthomonads enter via stomata to reach the apoplast. Herein, we investigated the differences in chemotactic responses for wide and narrow-host-range strains of Xcc A, X. euvesicatoria pv. citrumelonis (X. alfalfae subsp. citrumelonis), the causal agent of citrus bacterial spot, and X. campestris pv. campestris, the crucifer black rot pathogen. These strains of Xanthomonas were compared for carbon source use, the chemotactic responses toward carbon compounds, chemotaxis sensor content, and responses to apoplastic fluids from Citrus spp. and Chinese cabbage (Brassica pekinensis). Different chemotactic responses occurred for carbon sources and apoplastic fluids, depending on the Xanthomonas strain and the host plant from which the apoplastic fluid was derived. Differential chemotactic responses to carbon sources and citrus apoplasts suggest that these Xanthomonas strains sense host-specific signals that facilitate their location and entry of stomatal openings or wounds.

Finding novel chemoreceptors that specifically sense and trigger chemotaxis toward polycyclic aromatic hydrocarbons in Novosphingobium pentaromativorans US6-1.

Bacterial chemotaxis can improve the efficiency of aromatic compound degradation, however, knowledge of how bacteria sense high-molecular-weight polycyclic aromatic hydrocarbons (HMW-PAHs), is limited. Here, the chemotactic responses of Novosphingobium pentaromativorans US6-1 to 9 aromatic compounds were investigated. The results showed that US6-1 chemotactically responded to phenanthrene (PHE), pyrene (PYR), benzo[a]pyrene (BaP) and their six metabolites. Six methyl-accepting chemotaxis proteins (MCPs) were annotated from US6-1 genome, four of which contained putative ligand-binding domains (LBDs). To confirm whether these four MCPs were involved in triggering chemotaxis toward PAHs, the MCP mutants were constructed. Observations showed a loss of the chemotactic responses to benzoate, phthalate, PHE and BaP only in the mutant ∆mcp03030. Surface plasmon resonance (SPR) assays further confirmed that MCP03030LBD specifically bound phthalate, PHE, PYR and BaP, while MCP18870LBD bound only PYR. The mutant ∆mcp03030-∆mcp18870 was then constructed and was shown to have lost the chemotactic response to 5 aromatic compounds. Combined with the effects of outer membrane transporter deletion on chemotaxis and MCP deletion on the PAH degradation, our study demonstrated that the chemoreceptors MCP03030 and MCP18870 can recognize PAHs and their metabolites in the periplasm, triggering metabolism-dependent and metabolism-independent chemotaxis, and be linked with HMW-PAH biodegradation.

Endocytosis of nutrient transporters in fungi: The ART of connecting signaling and trafficking.

Plasma membrane transporters play pivotal roles in the import of nutrients, including sugars, amino acids, nucleobases, carboxylic acids, and metal ions, that surround fungal cells. The selective removal of these transporters by endocytosis is one of the most important regulatory mechanisms that ensures a rapid adaptation of cells to the changing environment (e.g., nutrient fluctuations or different stresses). At the heart of this mechanism lies a network of proteins that includes the arrestin-related trafficking adaptors (ARTs) which link the ubiquitin ligase Rsp5 to nutrient transporters and endocytic factors. Transporter conformational changes, as well as dynamic interactions between its cytosolic termini/loops and with lipids of the plasma membrane, are also critical during the endocytic process. Here, we review the current knowledge and recent findings on the molecular mechanisms involved in nutrient transporter endocytosis, both in the budding yeast Saccharomyces cerevisiae and in some species of the filamentous fungus Aspergillus. We elaborate on the physiological importance of tightly regulated endocytosis for cellular fitness under dynamic conditions found in nature and highlight how further understanding and engineering of this process is essential to maximize titer, rate and yield (TRY)-values of engineered cell factories in industrial biotechnological processes.

Promotion of Momordica Charantia polysaccharides on neural stem cell proliferation by increasing SIRT1 activity after cerebral ischemia/reperfusion in rats.

The deacetylase SIRT1 has been reported to play a critical role in regulating neurogenesis, which may be an adaptive processes contributing to recovery after stroke. Our previous work showed that the antioxidant capacity of Momordica charantia polysaccharides (MCPs) could protect against cerebral ischemia/reperfusion (I/R) after stroke. However, whether the protective effect of MCPs on I/R injury is related to neural stem cell (NSC) proliferation remains unclear. In the present study, we designed invivo and invitro experiments to elucidate the underlying mechanisms by which MCPs promote endogenous NSC proliferation during cerebral I/R. Invivo results showed that MCPs rescued the memory and learning abilities of rats after I/R damage and enhanced NSC proliferation in the rat subventricular zone (SVZ) and subgrannular zone (SGZ) during I/R. Invitro experiments demonstrated that MCPs could stimulate the proliferation of C17.2 cells under oxygen-glucose deprivation (OGD) conditions. Further studies revealed that the proliferation-promoting mechanism of MCPs relied on increasing the activity of SIRT1, decreasing the level of acetylation of ß-catenin in the cytoplasm, and then triggering the translocation of ß-catenin into the nucleus. These data provide experimental evidence that the up-regulation of SIRT1 activity by MCPs led to an increased cytoplasmic deacetylation of ß-catenin, which promoted translocation of ß-catenin to the nucleus to participate in the signaling pathway involved in NSC proliferation. The present study reveals that MCPs function as a therapeutic drug to promote stroke recovery by increasing the activity of SIRT1, decreasing the level of acetylated ß-catenin, promoting the nuclear translocation of ß-catenin and thereby increasing endogenous NSC proliferation.

Momordica charantia polysaccharides modulate the differentiation of neural stem cells via SIRT1/Β-catenin axis in cerebral ischemia/reperfusion.

BACKGROUND: Stroke is the leading cause of long-term motor disability and cognitive impairment. Recently, neurogenesis has become an attractive strategy for the chronic recovery of stroke. It is important to understand the molecular mechanism that promotes neural stem cell (NSC) neurogenesis for future NSC-based therapies. Our previous study showed that Momordica charantia polysaccharides (MCPs) exerted neuroprotective effects on stroke via their anti-oxidant and anti-inflammation activities. However, it remains unknown whether MCPs promote NSC neurogenesis after cerebral ischemic/reperfusion injury (IRI). METHODS: We investigated MCPs' function in differentiation of neural stem cells (NSCs) in vivo and in vitro experiments. Based on a middle cerebral artery occlusion (MCAO) rat model, the effect of MCPs on neuronal differentiation after MCAO was analyzed. Primary NSCs and neural stem cell line C17.2 were cultured and subjected to glutamate stimulation to establish the cell model of IRI. We evaluated the effect of MCPs on NSC differentiation in IRI cell model by Western blot and immunofluorescence staining. The SIRT1 activity of NSCs post glutamate stimulation was also evaluated by CELL SIRT1 COLORIMETRY ASSAY KIT. In addition, molecular mechanism was clarified by employing the activator and inhibitor of SIRT1. RESULTS: MCPs had no effects on the differentiation of neural stem cells under physiological conditions while shifted NSC differentiation potential from the gliogenic to neurogenic lineage under pathological conditions. Activation of SIRT1 with MCPs was responsible for the neuronal differentiation of C17.2-NSCs. The neuronal differentiation effect of MCPs was attributed to upregulation SIRT1-mediated deacetylation of ß-catenin. MCP-induced deacetylation via SIRT1 promoted nuclear accumulation of ß-catenin in NSCs. CONCLUSION: Our findings indicate that the deacetylation of ß-catenin by SIRT1 represents a critical mechanism of action of MCPs in promoting NSC neuronal differentiation. It provides an improved understanding of molecular mechanism underlying neuroprotective effects of MCPs in IRI, indicating its potential role on treating ischemic stroke especially chronic recovery.

An Efficient Certificateless Aggregate Signature Scheme for Blockchain-Based Medical Cyber Physical Systems.

Different from the traditional healthcare field, Medical Cyber Physical Systems (MCPS) rely more on wireless wearable devices and medical applications to provide better medical services. The secure storage and sharing of medical data are facing great challenges. Blockchain technology with decentralization, security, credibility and tamper-proof is an effective way to solve this problem. However, capacity limitation is one of the main reasons affecting the improvement of blockchain performance. Certificateless aggregation signature schemes can greatly tackle the difficulty of blockchain expansion. In this paper, we describe a two-layer system model in which medical records are stored off-blockchain and shared on-blockchain. Furthermore, a multi-trapdoor hash function is proposed. Based on the proposed multi-trapdoor hash function, we present a certificateless aggregate signature scheme for blockchain-based MCPS. The purpose is to realize the authentication of related medical staffs, medical equipment, and medical apps, ensure the integrity of medical records, and support the secure storage and sharing of medical information. The proposed scheme is highly computationally efficient because it does not use bilinear maps and exponential operations. Many certificateless aggregate signature schemes without bilinear maps in Internet of things (IoT) have been proposed in recent years, but they are not applied to the medical field, and they do not consider the security requirements of medical data. The proposed scheme in this paper has high computing and storage efficiency, while meeting the security requirements in MCPS.

Cargo encapsulation in bacterial microcompartments: Methods and analysis.

Metabolic engineers seek to produce high-value products from inexpensive starting materials in a sustainable and cost-effective manner by using microbes as cellular factories. However, pathway development and optimization can be arduous tasks, complicated by pathway bottlenecks and toxicity. Pathway organization has emerged as a potential solution to these issues, and the use of protein- or DNA-based scaffolds has successfully increased the production of several industrially relevant compounds. These efforts demonstrate the usefulness of pathway colocalization and spatial organization for metabolic engineering applications. In particular, scaffolding within an enclosed, subcellular compartment shows great promise for pathway optimization, offering benefits such as increased local enzyme and substrate concentrations, sequestration of toxic or volatile intermediates, and alleviation of cofactor and resource competition with the host. Here, we describe the 1,2-propanediol utilization (Pdu) bacterial microcompartment (MCP) as an enclosed scaffold for pathway sequestration and organization. We first describe methods for controlling Pdu MCP formation, expressing and encapsulating heterologous cargo, and tuning cargo loading levels. We further describe assays for analyzing Pdu MCPs and assessing encapsulation levels. These methods will enable the repurposing of MCPs as tunable nanobioreactors for heterologous pathway encapsulation.