Ligand Phase Separation-Promoted, "Squeezing-Out" Mode Explaining the Mechanism and Implications of Neutral Nanoparticles That Escaped from Lysosomes.

Neutral nanomaterials functionalized with PEG or similar molecules have been popularly employed as nanomedicines. Compared to positive counterparts that are capable of harnessing the well-known proton sponge effect to facilitate their escape from lysosomes, it is yet unclear how neutral substances got their entry into the cytosol. In this study, by taking PEGylated, neutral Au nanospheres as an example, we systematically investigated their time-dependent translocation postuptake. Specifically, we harnessed dissipative particle dynamics simulations to uncover how nanospheres bypass lysosomal entrapment, wherein a mechanism termed as "squeezing-out" mode was discovered. We next conducted a comprehensive investigation on how nanomaterials implicate lysosomes in terms of integrity and functionality. By using single-molecule imaging, specific preservation of PEG-terminated with targeting moieties in lysosomes supports the "squeezing-out" mode as the mechanism underlying the lysosomal escape of nanomaterials. All evidence points out that such a process is benign to lysosomes, wherein the escape of nanomaterials proceeds at the expense of targeting moieties loss. Furthermore, we proved that by fine-tuning of the efficacy of nanomaterials escaping from lysosomes, modulation of distinct pathways and metabolic machinery can be achieved readily, thereby offering us a simple and robust tool to implicate cells.

Large-Scale Continuous and In Situ Photosynthesis of Hydrogen Peroxide by Sulfur-Functionalized Polymer Catalyst for Water Treatment.

Photocatalytic H2 O2 generation system based on polymer catalyst receives increasing attention in recent years; however, the insufficient charge separation efficiency and low oxygen adsorption/activation capacity severely limit their potential application. In this study, a sulfur (C=S) functionalized polymer catalyst is reported through a green water-mediated and catalyst-free multi-component reactions (MCRs) route. The sulfur functional group endows the polymer with a suitable energy band and facilitates the separation of photogenerated electron-hole pair. The reported polymer achieves a high H2 O2 production efficiency (3132 µmol g-1 h-1 ) in pure water without oxygen aeration. To demonstrate their potential in in situ wastewater treatment, a panel reactor system (20×20 cm) is constructed for large-scale production of H2 O2 , which realizes continuous degradation of emerging pollutants including antibiotics and bisphenol A under natural sunlight irradiation condition. The H2 O2 utilization efficiency of the photo-self-Fenton system using in situ generated H2 O2 is found 7.9 times higher than that of the traditional photo-Fenton system. This study offers new insights in green synthesis and design of functional polymer photocatalyst, and demonstrates the feasibility of panel reactor system for large-scale continuous H2 O2 photocatalytic production and water treatment.

A novel S-scheme heterojunction of Cd0.5Zn0.5S/BiOCl with oxygen defects for antibiotic norfloxacin photodegradation: Performance, mechanism, and intermediates toxicity evaluation.

S-scheme heterojunction structure can endow the photocatalysts with high-performance photo-degradation of pharmaceuticals and personal care products (PPCPs) since it can remain the photogenerated electrons/holes with stronger redox ability. Herein, an integrative S-scheme heterojunction photocatalyst building from Cd0.5Zn0.5S nanoparticles and BiOCl microflowers with oxygen vacancies (OVs) was developed. Moreover, the in-situ grown process ensures the firm contact and intense electron coupling between BiOCl and Cd0.5Zn0.5S. As a result, Cd0.5Zn0.5S/BiOCl exhibited a significant reinforcement of photo-activity and stability for the abatement of antibiotic norfloxacin, manifesting a 2.8-fold or 9.6-fold enhancement compared to pristine Cd0.5Zn0.5S or BiOCl. Cd0.5Zn0.5S/BiOCl also shows good resistance to alkaline, sodium salts and humic acid. The performance of Cd0.5Zn0.5S/BiOCl to photocatalytically degrade other PPCPs with different molecular structures was further confirmed. At last, the ability of Cd0.5Zn0.5S/BiOCl for PPCPs de-toxicity was verified by evaluating the toxicity of norfloxacin and its degradation intermediate. This study demonstrates a new S-scheme heterojunction photocatalyt for efficient removal of PPCPs as well as provides some insights into developing high-performance metal sulfide solid-solution-based S-scheme heterojunctions for water decontamination.

Adaptive Neural Finite-Time Control of Non-Strict Feedback Nonlinear Systems With Non-Symmetrical Dead-Zone.

The control design method for a class of non-strict feedback nonlinear systems is studied in this brief considering uncertain nonlinearities and unknown non-symmetrical input dead-zone. Combining with the finite-time command filtered backstepping (FCFB) technique, a novel finite-time adaptive control approach is proposed in which a neural network-based methodology is adopted to cope with the uncertain nonlinearities in the non-strict feedback form. The input dead-zone model is transformed into a simple linear system with unknown gain and bounded disturbance which is estimated by an adaptive factor. Using the finite-time Lyapunov theory, the system convergence is proved. And the effectiveness of the proposed control scheme is verified through comparative numerical simulations.

Designing oxygen vacancy mediated bismuth molybdate (Bi2MoO6)/N-rich carbon nitride (C3N5) S-scheme heterojunctions for boosted photocatalytic removal of tetracycline antibiotic and Cr(VI): Intermediate toxicity and mechanism insight.

Polymeric N-rich carbon nitride of C3N5 is being utilized as a new visible-light-driven catalyst due to its narrower bandgap (∼2.0 eV). Building step-scheme (S-scheme) heterojunction by coupling with other semiconductors especially those own oxygen vacancies (OVs) can further upgrade the photocatalytic performance of C3N5-based photocatalysts. Herein, a novel S-scheme heterojunction of OVs mediated Bi2MoO6/C3N5 was fabricated by in-situ growing Bi2MoO6 nanoparticles with OVs on C3N5 nanosheets. Benefiting from the efficient separation and transfer of high energetic charge carriers by S-scheme charge migration, enriched structural defects, as well as the close contact by the in-situ growth, the heterojunction exhibited superior visible-light photocatalytic performance toward the removal of tetracycline (TC) and Cr(VI) than C3N5, Bi2MoO6, and their mechanical mixture under visible light. The TC degradation routes and the bio-toxicity evolution of TC were explored. Moreover, the photocatalytic mechanism for TC decomposition and Cr(VI) reduction over Bi2MoO6/C3N5 with OVs were elucidated. This work presents a newfangled vision for designing promising C3N5-based S-scheme heterojunction photocatalysts for pollution control.

Immunostimulatory and antioxidant activities of the selenized polysaccharide from edible Grifola frondosa.

Grifola frondosa polysaccharide (GFP2) was extracted and purified by anion-exchange chromatography. A selenized G. frondosa polysaccharide, SeGFP2, was modified in selenylation by nitric acid-sodium selenite (HNO3-Na2SeO3) method. Structural features were investigated, and the lymphocyte proliferation and antioxidant activities were compared taking GFP2 as control. SeGFP2 with a molecular weight of 2.12 × 104 Da was composed of mannose, glucose, and galactose with a ratio of 3.5:11.8:1.0. A typical absorption of selenium ester was observed in SeGFP2 molecule. SeGFP2 was proposed as a branched polysaccharide, which consisted of 1,3-D-Glcp, 1,6-D-Glcp, 1,4,6-D-Galp, and 1,3,6-D-Manp. SeGFP2 showed a linear filamentous structure with some branches. SeGFP2 could significantly promote T- or B-lymphocyte proliferation and the enhancement was higher than GFP2. The in vitro antioxidant activities of SeGFP2 were more potent than GFP2. These present data suggested that selenylation could significantly improve the lymphocyte proliferation and in vitro antioxidant activities of GFP2.

Rationally designed tetra (4-carboxyphenyl) porphyrin/graphene quantum dots/bismuth molybdate Z-scheme heterojunction for tetracycline degradation and Cr(VI) reduction: Performance, mechanism, intermediate toxicity appraisement.

Constructing novel Z-scheme heterojunctions is an effective strategy for obtaining high-performance photocatalysts. Herein, tetra (4-carboxyphenyl) porphyrin (TCPP) and graphene quantum dots (GQDs) were loaded on the surface of Bi2MoO6 (BMO) to fabricate novel Z-scheme heterojunctions of TCPP/G/BMO. Especially, TCPP/G/BMO-2 showed an exceptional visible-light photoactivity for tetracycline (TC) degradation (81.0%, 40 min) and Cr(VI) reduction (90.7%, 60 min), respectively by 2.38 folds and 2.96 folds enhancement compared to sole Bi2MoO6. The substantial enhancement of activity is attributed to the synergy effect of the Z-scheme charge transfer mechanism and the improved light absorption. The degradation pathways of TC were inferred by determining the generated intermediates using high performance liquid chromatography-mass spectrometry (HPLC-MS), and the toxicity of the transformation products was assessed by Toxicity Estimation Software Tool (T.E.S.T). Overall, on the basis of trapping experiments and electron spin resonance spectra (ESR) analysis, the photocatalytic mechanisms of Cr(VI) reduction and TC degradation by the TCPP/G/BMO Z-scheme heterojunction was proposed. This work indicates TCPP/G/BMO could be a promising photocatalyst for wastewater treatment.

Facile construction of novel organic-inorganic tetra (4-carboxyphenyl) porphyrin/Bi2MoO6 heterojunction for tetracycline degradation: Performance, degradation pathways, intermediate toxicity analysis and mechanism insight.

Developing durable photocatalysts with highly efficient antibiotics degradation is crucial for environment purification. Herein, tetra (4-carboxyphenyl) porphyrin (TCPP) was loaded onto the surface of Bi2MoO6 microspheres to gain hierarchical organic-inorganic TCPP/Bi2MoO6 (TCPP/BMO) heterojunctions via a facile impregnation strategy. The catalytic properties of these catalysts were comprehensively investigated through the photodegradation of tetracycline hydrochloride (TC) under visible light. Among all the TCPP/BMO heterojunctions, the highest photodegradation rate constant (0.0278 min-1) was achieved with 0.25 wt% TCPP (TCPP/BMO-2), which was approximately 1.15 folds greater than that of pristine Bi2MoO6 and far superior to pure TCPP. The extremely high photocatalytic performance is attributed to the interfacial interaction between TCPP and Bi2MoO6, which favors the efficient separation of charge carriers and the enhancement of visible-light absorbance. TCPP/BMO-2 possesses high mineralization capability and good recycling performance. Photo-induced O2-, h+, and OH were mainly responsible for the degradation of TC. The degradation pathways of TC and toxicity of degradation intermediates were analyzed based on the intermediates detected by the high performance liquid chromatography-mass spectrometer (HPLC-MS) and the toxicity assessment by the quantitative structure-activity relationship (QSAR) prediction. A possible photocatalytic mechanism over TCPP/BMO is proposed. This work offers an insight in developing the porphyrin-based organic-inorganic heterojunctions for effectively remedying pharmaceutical wastewater.

Dual-Motor Synchronization Control Design Based on Adaptive Neural Networks Considering Full-State Constraints and Partial Asymmetric Dead-Zone.

This paper proposes a command filtering backstepping (CFB) scheme with full-state constraints by leading into time-varying barrier Lyapunov functions (T-BLFs) for a dual-motor servo system with partial asymmetric dead-zone. Firstly, for the convenience of the controller design, the conventional partial asymmetric dead-zone model was replaced with a new smooth differentiable model owing to its non-smoothness. Secondly, neural networks (NNs) were utilized to approximate the nonlinearity that exists in the dead-zone model, improving the control performance. In addition, CFB was utilized to deal with the inherent computational explosion problem of the traditional backstepping method, and an error compensation mechanism was introduced to further reduce the filtering errors. Then, by applying the T-BLF to the CFB process, the states of the system never violated the prescribed constraints, and all signals in the dual-motor servo system were bounded. The tracking error and synchronization error could converge to a small desired neighborhood of the origin. In the end, the effectiveness of the proposed control scheme was verified through simulations.

Vaccarin enhances intestinal barrier function in type 2 diabetic mice.

AIMS: Hyperglycemia and insulin resistance drive intestinal barrier dysfunction in type 2 diabetes (T2DM). Vaccarin, the main active component in the semen of traditional Chinese medicine Vaccaria has a definite effect on T2DM mice. The purpose of this study was to investigate whether vaccarin can enhance the intestinal barrier function in T2DM. MAIN METHODS: The T2DM mice model was established by streptozocin and high-fat diet. Vaccarin at a dose of 1 mg/kg/day was administered. We evaluated the effects of vaccarin on gut microbiota and intestinal barrier function by 16S rRNA sequencing, Western blot, quantitative fluorescent PCR (qPCR), and morphological observation. Moreover, we constructed a single layer of the human intestinal epithelium model to determine the effect of vaccarin in vitro. RESULTS: The experimental results showed that vaccarin alleviated inflammatory mediators in serum and intestinal tissue of mice (P < 0.05), which may depend on the improvement of tight junctions and gut microbiota (P < 0.05). Activation of extracellular regulated protein kinases (Erk1/2) stimulated myosin light chain kinase (MLCK). By inhibiting ERK expression (P < 0.05), vaccarin had similar effects to ERK inhibitors. In addition, the regulation of tight junction barriers also involved the abovementioned pathways in vivo. CONCLUSION: Vaccarin could protect the intestinal barrier by inhibiting the ERK/MLCK signaling pathway and modulate the composition of the microbiota. These results suggested that vaccarin may be an effective candidate for improving intestinal barrier changes in T2DM.

Regulation of the yeast formin Bni1p by the actin-regulating kinase Prk1p.

The formin family of proteins promotes the assembly of linear actin filaments in the cells of diverse eukaryotic organisms. The predominant formins in mammalian cells are self-inhibited by an intramolecular interaction between two terminal domains and are activated by the binding of the Rho GTPases and other factors. In this study, we show that Bni1p, a formin required for the assembly of actin cables in budding yeast, is also regulated by an autoinhibitory mechanism and phosphorylation by the actin regulatory kinase Prk1p, and possibly Ark1p as well, plays a key role in unlocking the inhibition. Bni1p is phosphorylated by Prk1p at three [L/V/I]xxxxTG motifs in vitro, and the phosphorylation is sufficient to activate Bni1p by disrupting its intramolecular interaction. This finding extends the roles of Prk1p in the regulation of actin dynamics to be associated with both anterograde and retrograde transport pathways, i.e. exocytosis and endocytosis, in yeast.

Negative regulation of the actin-regulating kinase Prk1p by patch localization-induced autophosphorylation.

The Prk1 family of protein kinases are important regulators of endocytosis and actin cytoskeleton in some eukaryotic cells. In budding yeast, Prk1p phosphorylates numerous endocytic proteins including Pan1p and Sla1p. Prk1p has been observed to undergo autophosphorylation in vivo. In this study, we determined the sites and underlying role of the autophosphorylation. Two sites located in the noncatalytic region were identified to be the autophosphorylation sites. When the sites were mutated, the non-autophosphorylatable Prk1p phosphorylated Pan1p and Sla1p more efficiently than the wild-type kinase, suggesting a negative effect of the autophosphorylation. In addition, the dynamic properties of actin and the coat complex were also altered in the autophosphorylation mutant cells. Interestingly, the autophosphorylation of Prk1p was dependent on cortical localization of the kinase and could be induced by phosphorylated Sla1p. These results suggest that the autophosphorylation of Prk1p may represent a feedback mechanism possibly involved in fine-tuning the pace of progression during actin-coupled endocytosis.

A novel septin-associated protein, Syp1p, is required for normal cell cycle-dependent septin cytoskeleton dynamics in yeast.

Septins are a family of GTP-binding proteins whose heterooligomeric complex is the basic structural element of the septin filaments found in many eukaryotic organisms. In budding yeast, septins are mainly confined at the mother-daughter junction and are required for cell morphogenesis and division. Septins undergo assembly and disassembly in accordance with the progression of the cell cycle. In this report, we identified the yeast protein Syp1p as a new regulator of septin dynamics. Syp1p colocalizes with septins throughout most of the cell cycle. Syp1p interacts with the septin subunit Cdc10p and can be precipitated by Cdc10p and Cdc12p. In the syp1Delta mutant, both formation of a complete septin ring at the incipient bud site and disassembly of the septin ring in later stages of cell division are significantly delayed. In addition, overexpression of Syp1p causes marked acceleration of septin disassembly. The fluorescence recovery after photobleaching (FRAP) assay further showed that Syp1p promotes septin turnover in different cell cycle stages. These results suggest that Syp1p is involved in the regulation of cell cycle-dependent dynamics of the septin cytoskeleton in yeast.

A novel function of Arp2p in mediating Prk1p-specific regulation of actin and endocytosis in yeast.

The yeast protein Pan1p plays essential roles in actin cytoskeleton organization and endocytosis. It couples endocytosis with actin polymerization through its dual function in endocytic complex assembly and activation of the actin polymerization initiation complex Arp2/3p. Phosphorylation of Pan1p and other components of the endocytic complex by the kinase Prk1p leads to disassembly of the coat complex and the termination of vesicle-associated actin polymerization. A homologous kinase, Ark1p, has also been implicated in this regulatory process. In this study, we investigated the distinct roles of Prk1p and Ark1p. We found that the nonkinase domains determined the functional specificity of the two kinases. A short region located adjacent to the kinase domain unique to Prk1p was found to be required for the kinase to interact with Arp2p. Further studies demonstrated that the Prk1p-Arp2p interaction is critical for down-regulation of Pan1p. These findings reveal that, in addition to its role in the nucleation of actin polymerization, Arp2p also mediates what appears to be an auto-regulatory mechanism possibly adapted for efficient coordination of actin assembly and disassembly during endocytosis.

Scd5p mediates phosphoregulation of actin and endocytosis by the type 1 phosphatase Glc7p in yeast.

Pan1p plays essential roles in both actin and endocytosis in yeast. It interacts with, and regulates the function of, multiple endocytic proteins and actin assembly machinery. Phosphorylation of Pan1p by the kinase Prk1p down-regulates its activity, resulting in disassembly of the endocytic vesicle coat complex and termination of vesicle-associated actin polymerization. In this study, we focus on the mechanism that acts to release Pan1p from phosphorylation inhibition. We show that Pan1p is dephosphorylated by the phosphatase Glc7p, and the dephosphorylation is dependent on the Glc7p-targeting protein Scd5p, which itself is a phosphorylation target of Prk1p. Scd5p links Glc7p to Pan1p in two ways: directly by interacting with Pan1p and indirectly by interacting with the Pan1p-binding protein End3p. Depletion of Glc7p from the cells causes defects in cell growth, actin organization, and endocytosis, all of which can be partially suppressed by deletion of the PRK1 gene. These results suggest that Glc7p antagonizes the activity of the Prk1p kinase in regulating the functions of Pan1p and possibly other actin- and endocytosis-related proteins.

Pan1p: an actin director of endocytosis in yeast.

The yeast protein Pan1p plays a key role in actin-driven endocytosis. The molecular architecture enables the protein to perform multivalent tasks. First, Pan1p acts as a central scaffold for assembly of coat complex at the endocytic sites through its binding to multiple endocytic proteins. Secondly, Pan1p is also required for normal actin cytoskeleton organization and dynamics at the cell cortex. It is capable of F-actin binding and promoting the Arp2/3-mediated actin nucleation via its WH2 and acid domains. Pan1p, therefore, is responsible for the mechanism of coupling the vesicle coat to actin network in the early steps of internalization. The function of Pan1p is under a negative regulation by the kinase Prk1p. Phosphorylation of Pan1p by Prk1p results in disassembly of the coat complex and dissociation of the vesicle from actin meshwork after internalization. The phosphorylation of Pan1p is possibly reversed by the type 1 phosphatase Glc7p, which will allow Pan1p to be reused for coat assembly in the next round of endocytosis.

Prk1p.

The protein kinase Prk1p (standing for p53 regulating kinase 1) of the yeast Saccharomyces cerevisiae is the prototype of a kinase family identified recently as important regulators of the actin cytoskeleton and endocytosis. These kinases all have a highly homologous serine/threonine kinase domain in their N-terminal region but share no significant homology in other regions. Prk1p also contains a proline-rich motif near its C-terminus that is required for the proper subcellular localization of the protein. The kinase activity of Prk1p has been confirmed by both in vitro and in vivo studies and shown to be essential for the protein's function. To date, several proteins that play essential roles in actin cytoskeleton organization and endocytosis have been identified as the regulatory targets of Prk1p. Phosphorylation on the [L/I/V/N]xx[Q/N/T/S]xTG motifs by Prk1p results in a down-regulation of the functions of these target proteins. The observation that many yeast proteins involved in the actin cytoskeleton organization and endocytosis contain the Prk1p phosphorylation motifs has led to the hypothesis that the Prk1p family of kinases are possibly the general regulators of the actin cytoskeleton and endocytosis in yeast.

The yeast dynamin-related GTPase Vps1p functions in the organization of the actin cytoskeleton via interaction with Sla1p.

Recent studies have suggested that the function of the large GTPase dynamin in endocytosis in mammalian cells may comprise a modulation of actin cytoskeleton. The role of dynamin in actin cytoskeleton organization in the yeast Saccharomyces cerevisiae has remained undefined. In this report, we found that one of the yeast dynamin-related proteins, Vps1p, is required for normal actin cytoskeleton organization. At both permissive and non-permissive temperatures, the vps1 mutants exhibited various degrees of phenotypes commonly associated with actin cytoskeleton defects: depolarized and aggregated actin structures, hypersensitivity to the actin cytoskeleton toxin latrunculin-A, randomized bud site selection and chitin deposition, and impaired efficiency in the internalization of membrane receptors. Over-expression of the GTPase mutants of vps1 also led to actin abnormalities. Consistent with these actin-related defects, Vps1p was found to interact physically, and partially co-localize, with the actin-regulatory protein Sla1p. The normal cellular localization of Sla1p required Vps1p and could be altered by over-expression of a region of Vps1p that was involved in the interaction with Sla1p. The same region also promoted mis-sorting of the vacuolar protein carboxypeptidase Y upon over-expression. These findings suggest that the functions of the dynamin-related protein Vps1p in actin cytoskeleton dynamics and vacuolar protein sorting are probably related to each other.

Identification of novel recognition motifs and regulatory targets for the yeast actin-regulating kinase Prk1p.

Prk1p is a serine/threonine kinase involved in the regulation of the actin cytoskeleton organization in the yeast Saccharomyces cerevisiae. Previously, we have identified LxxQxTG as the phosphorylation site of Prk1p. In this report, the recognition sequence for Prk1p is investigated more thoroughly. It is found that the presence of a hydrophobic residue at the position of P-5 is necessary for Prk1p phosphorylation and L, I, V, and M are all able to confer the phosphorylation at various efficiencies. The residue flexibility at P-2 has also been identified to include Q, N, T, and S. A homology-based three-dimensional model of the kinase domain of Prk1p provided some structural interpretations for these substrate specificities. The characterization of the [L/I/V/M]xx[Q/N/T/S]xTG motif led to the identification of a spectrum of potential targets for Prk1p from yeast genome. One of them, Scd5p, which contains three LxxTxTG motifs and is previously known to be important for endocytosis and actin organization, has been chosen to demonstrate its relationship with Prk1p. Phosphorylation of Scd5p by Prk1p at the three LxxTxTG motifs could be detected in vitro and in vivo, and deletion of PRK1 suppressed the defects in actin cytoskeleton and endocytosis in one of the scd5 mutants. These results allowed us to conclude that Scd5p is likely another regulatory target of Prk1p.