Biological macromolecules must fold into native structures to gain functional activities. In living cells, proteins called molecular chaperones mediate productive folding by preventing undesired ...interactions and aggregation and by facilitating refolding of misfolded macromolecules into their bioactive forms. Inspired by natural molecular chaperones, artificial chaperones that mimic some features of their biological counterparts have been designed. This review describes recent progress in the development of artificial chaperones and their promising applications in enhancing macromolecular assembly of proteins, polypeptides, and nucleic acids.
The environmental DNA (eDNA) technique is expected to become a powerful, non-invasive tool for estimating the distribution and biomass of organisms. This technique was recently shown to be applicable ...to aquatic vertebrates by collecting extraorganismal DNA floating in the water or absorbed onto suspended particles. However, basic information on eDNA release rate is lacking, despite it being essential for practical applications. In this series of experiments with bluegill sunfish (Lepomis macrochirus), we examined the effect of fish developmental stage on eDNA release rate. eDNA concentration reached equilibrium 3 days after the individual fish were introduced into the separate containers, enabling calculation of the eDNA release rate (copies h-1) from individual fish on the assumption that the number of eDNA released from the fish per unit time equals total degradation in the container (copies h-1). The eDNA release rate was 3-4 times higher in the adult (body weight: 30-75 g) than in the juvenile group (0.5-2.0 g). Such positive relationship between fish size and eDNA release rate support the possibility of biomass rather than density estimation using eDNA techniques. However, the eDNA release rate per fish body weight (copies h-1 g-1) was slightly higher in the juvenile than the adult group, which is likely because of the ontogenetic reduction in metabolic activity. Therefore, quantitative eDNA data should be carefully interpreted to avoid overestimating biomass when the population is dominated by juveniles, because the age structure of the focal population is often variable and unseen in the field. eDNA degradation rates (copies l-1 h-1), calculated by curve fitting of time-dependent changes in eDNA concentrations after fish removal, were 5.1-15.9% per hour (half-life: 6.3 h). This suggests that quantitative eDNA data should be corrected using a degradation curve attained in the target field.
The growth of lamellar crystals has been studied in particular for spherulites in polymeric materials. Even though such spherulitic structures and their growth are of crucial importance for the ...mechanical and optical properties of the resulting polymeric materials, several issues regarding the residual stress remain unresolved in the wider context of crystal growth. To gain further insight into micro-mechanical forces during the crystallization process of lamellar crystals in polymeric materials, herein, we introduce tetraarylsuccinonitrile (TASN), which generates relatively stable radicals with yellow fluorescence upon homolytic cleavage at the central C-C bond in response to mechanical stress, into crystalline polymers. The obtained crystalline polymers with TASN at the center of the polymer chain allow not only to visualize the stress arising from micro-mechanical forces during polymer crystallization via fluorescence microscopy but also to evaluate the micro-mechanical forces upon growing polymer lamellar crystals by electron paramagnetic resonance, which is able to detect the radicals generated during polymer crystallization.
Recent studies in streams and ponds have demonstrated that the distribution and biomass of aquatic organisms can be estimated by detection and quantification of environmental DNA (eDNA). In more open ...systems such as seas, it is not evident whether eDNA can represent the distribution and biomass of aquatic organisms because various environmental factors (e.g., water flow) are expected to affect eDNA distribution and concentration. To test the relationships between the distribution of fish and eDNA, we conducted a grid survey in Maizuru Bay, Sea of Japan, and sampled surface and bottom waters while monitoring biomass of the Japanese jack mackerel (Trachurus japonicus) using echo sounder technology. A linear model showed a high R(2) value (0.665) without outlier data points, and the association between estimated eDNA concentrations from the surface water samples and echo intensity was significantly positive, suggesting that the estimated spatial variation in eDNA concentration can reflect the local biomass of the jack mackerel. We also found that a best-fit model included echo intensity obtained within 10-150 m from water sampling sites, indicating that the estimated eDNA concentration most likely reflects fish biomass within 150 m in the bay. Although eDNA from a wholesale fish market partially affected eDNA concentration, we conclude that eDNA generally provides a 'snapshot' of fish distribution and biomass in a large area. Further studies in which dynamics of eDNA under field conditions (e.g., patterns of release, degradation, and diffusion of eDNA) are taken into account will provide a better estimate of fish distribution and biomass based on eDNA.
Multi-component nucleic acid enzymes (MNAzymes) are allosteric deoxyribozymes that are activated upon binding of a specific nucleic acid effector. MNAzyme activity is limited due to an insufficient ...assembly of the MNAzyme and its turnover. In this work, we describe the successful improvement of MNAzyme reactivity and selectivity by addition of cationic copolymers, which exhibit nucleic acid chaperone-like activity. The copolymer allowed a 210-fold increase in signal activity and a 95-fold increase in the signal-to-background selectivity of MNAzymes constructed for microRNA (miRNA) detection. The selectivity of the MNAzyme for homologous miRNAs was demonstrated in a multiplex format in which isothermal reactions of two different MNAzymes were performed. In addition, the copolymer permitted miRNA detections even in the presence of a ribonuclease which is ubiquitous in environments, indicating the protective effect of the copolymer against ribonucleases.
Photodynamic therapy is a new type of anti‐tumor therapy with excellent therapeutic effects and minor side effects. The key factor for photodynamic therapy is highly efficient loading and protection ...of photosensitizers. Covalent organic framework is a new type of organic porous material with rich sources and has huge development potential in the loading of photosensitizers. However, the π–π interaction between the rigid monomers inevitably causes aggregation and quenching between photosensitizers, which in turn affects the rate of reactive oxygen production. Here, newly designed cationic flexible organic framework nanoparticles (PEI‐Por NPs) are synthesized via one‐step method with PEI25K and meso‐tetra(p‐formylphenyl)porphyrin under microwave irradiation. The structure of the flexible organic framework can effectively inhibit the aggregation and quenching of porphyrin. In addition, PEI‐Por NPs had excellent gene transfection ability both in vitro and in vivo. Excellent antitumor effect can be achieved by combining PEI‐Por NPs’ photodynamic therapy capacity and PEI‐Por NPs‐mediated PD‐L1 gene silencing with the guidance of fluorescence imaging and photoacoustic imaging. This cationic flexible organic framework material combines the advantages of flexible building units and rigid monomers, which provides a basis for the development of nano‐photosensitizers and excellent gene carriers, and has great potential for clinical application.
This article provides a method for preparing cationic flexible organic framework nanoparticles (PEI‐Por NPs), which has good gene transfection ability and photodynamic effect. Further, effective tumor suppression is achieved by combining photodynamic therapy and gene immunotherapy. The preparation of nanoparticles based on flexible polymers and rigid porphyrin monomers provides new ideas for the design of high‐efficiency photosensitizers and gene carriers.
The kinetics of photoinduced reactions can be approached by laser flash photolysis techniques. Although such techniques allow for a detailed understanding of the important photophysics of molecules, ...they normally require a substantial amount of sample for measurements (>1 nmol), and thus, they are difficult to apply to analytical and diagnostic applications. The photophysics of a fluorescent molecule can be accessed by monitoring the kinetics of the fluctuation of fluorescence, which is called blinking. Blinking is a phenomenon that can be monitored only if molecules are observed at the single‐molecule level. In bulk solution, blinking kinetics can be measured by using fluorescence correlation spectroscopy (FCS), which normally requires more than 105 times less sample than that required for laser flash photolysis. Blinking is controlled to extract fruitful microenvironmental information around a fluorescent molecule, by using a method named kinetic analysis based on the control of fluorescence blinking (KACB). This Concept highlights the adaption of the KACB method to investigate the local conformation of DNA with less than 1 pmol of DNA sample.
Blink, don't FRET! The kinetics of photoinduced reactions can be accessed through fluorescence correlation spectroscopy. By measuring the OFF time during blinking, the lifetimes of photoinduced intermediates, such as triplet, radical ion, and isomerized states, can be determined.
The ubiquitin-proteasome pathway degrades ubiquitinated proteins to remove damaged or misfolded protein and thus plays an important role in the maintenance of many important cellular processes. ...Because the pathway is also crucial for tumor cell growth and survival, proteasome inhibition by specific inhibitors exhibits potent antitumor effects in many cancer cells. xCT, a subunit of the cystine antiporter system x
c
−
, plays an important role in cellular cysteine and glutathione homeostasis. Several recent reports have revealed that xCT is involved in cancer cell survival; however, it was unknown whether xCT affects the cytotoxic effects of proteasome inhibitors. In this study, we found that two stress-inducible transcription factors, Nrf2 and ATF4, were upregulated by proteasome inhibition and cooperatively enhance human xCT gene expression upon proteasome inhibition. In addition, we demonstrated that the knockdown of xCT by small interfering RNA (siRNA) or pharmacological inhibition of xCT by sulfasalazine (SASP) or (S)-4-carboxyphenylglycine (CPG) significantly increased the sensitivity of T24 cells to proteasome inhibition. These results suggest that the simultaneous inhibition of both the proteasome and xCT could have therapeutic benefits in the treatment of bladder tumors.
Recent studies have disclosed the function of enhancer RNAs (eRNAs), which are long non-coding RNAs transcribed from gene enhancer regions, in transcriptional regulation. However, it remains unclear ...whether eRNAs are involved in the regulation of human heme oxygenase-1 gene (HO-1) induction. Here, we report that multiple nuclear-enriched eRNAs are transcribed from the regions adjacent to two human HO-1 enhancers (i.e. the distal E2 and proximal E1 enhancers), and some of these eRNAs are induced by the oxidative stress-causing reagent diethyl maleate (DEM). We demonstrated that the expression of one forward direction (5' to 3') eRNA transcribed from the human HO-1 E2 enhancer region (named human HO-1enhancer RNA E2-3; hereafter called eRNA E2-3) was induced by DEM in an NRF2-dependent manner in HeLa cells. Conversely, knockdown of BACH1, a repressor of HO-1 transcription, further increased DEM-inducible eRNA E2-3 transcription as well as HO-1 expression. In addition, we showed that knockdown of eRNA E2-3 selectively down-regulated DEM-induced HO-1 expression. Furthermore, eRNA E2-3 knockdown attenuated DEM-induced Pol II binding to the promoter and E2 enhancer regions of HO-1 without affecting NRF2 recruitment to the E2 enhancer. These findings indicate that eRNAE2-3 is functional and is required for HO-1 induction.
•Alginate shielding improved the stability and decreased the cytotoxicity of PEI/DNA complexes.•Ca2+/(Alg/PEI/DNA) nanoparticles exhibited enhanced stability compared to Alg/PEI/DNA ...complexes.•Ca2+/(Alg/PEI/DNA) nanoparticles showed improved transfection efficiency compared to PEI/DNA complexes.
In gene therapy, the stability of gene delivery system is essential during the systemic circulation. To improve the stability of PEI/DNA complexes, sodium alginate (Alg) was chosen as the optimal shielding material from four different polysaccharides. Then Alg/PEI/DNA complexes were further crosslinked by calcium ions (Ca2+) to form Ca2+/(Alg/PEI/DNA) nanoparticles. Compared to PEI/DNA complexes and Alg/PEI/DNA complexes, Ca2+/(Alg/PEI/DNA) nanoparticles exhibited enhanced stability, which was confirmed by the in vitro and in vivo evaluations. Furthermore, the pharmacokinetic study indicated that Ca2+/(Alg/PEI/DNA) nanoparticles exhibited longer circulation time in blood, which would be beneficial to the enhanced permeability and retention (EPR) effect of nanoparticles and could realize improved nanoparticles accumulation at tumor site. Overall, the Ca2+/(Alg/PEI/DNA) nanoparticles based on alginate shielding and Ca2+ crosslinking might be a promising platform in gene delivery and tumor treatment.
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