Insights into heat response mechanisms in Clematis species: physiological analysis, expression profiles and function verification.

KEY MESSAGE: Our results provide insights into heat response mechanisms among Clematis species. Overexpressing CvHSFA2 enhanced the heat resistance of yeast and silencing NbHSFA2 reduced the heat resistance of tobacco. Clematis species are commonly grown in western and Japanese gardens. Heat stress can inhibit many physiological processes mediating plant growth and development. The mechanism regulating responses to heat has been well characterized in Arabidopsis thaliana and some crops, but not in horticultural plants, including Clematis species. In this study, we found that Clematis alpina 'Stolwijk Gold' was heat-sensitive whereas Clematis vitalba and Clematis viticella 'Polish Spirit' were heat-tolerant based on the physiological analyses in heat stress. Transcriptomic profiling identified a set of heat tolerance-related genes (HTGs). Consistent with the observed phenotype in heat stress, 41.43% of the differentially expressed HTGs between heat treatment and control were down-regulated in heat-sensitive cultivar Stolwijk Gold, but only 9.80% and 20.79% of the differentially expressed HTGs in heat resistant C. vitalba and Polish Spirit, respectively. Co-expression network, protein-protein interaction network and phylogenetic analysis revealed that the genes encoding heat shock transcription factors (HSFs) and heat shock proteins (HSPs) may played an essential role in Clematis resistance to heat stress. Two clades of heat-induced CvHSFs were further identified by phylogenetic tree, motif analysis and qRT-PCR. Ultimately, we proposed that overexpressing CvHSFA2-2 could endow yeast with high temperature resistance and silencing its homologous gene NbHSFA2 reduced the heat resistance of tobacco. This study provides first insights into the diversity of the heat response mechanisms among Clematis species.

A Cysteine-Mediated Synthesis of Red Phosphorus Nanosheets.

Among phosphorus-based nanomaterials, layered black phosphorus and violet phosphorus have been actively explored in the past decade. However, methods for the synthesis of red phosphorus nanosheets (RPNSs) is lacking, even though red phosphorus (RP) is commercially available at low cost and has excellent chemical stability at room temperature. We report an efficient strategy for fabrication of RPNSs and doped RPNSs using cysteine as a reducing reagent. Data from in vitro and in vivo studies suggested that RPNSs can trigger production of reactive oxygen species, DNA damage, and subsequent autophagy-mediated cell death in a shape-dependent manner. Our findings provide a method for construction of layered RP nanomaterials and they present a unique mechanism for the application of phosphorus-based materials in nanomedicines.

An Inorganic Biopolymer Polyphosphate Controls Positively Charged Protein Phase Transitions.

Polyphosphate (PolyP) is one of the most compact inorganic polyanionic biopolymers that participates in various physiological processes. However, the mechanism of the interaction between polyP and proteins remains poorly understood. Herein, we report that polyP can interact with positively charged green fluorescent protein, +36GFP, resulting in liquid-liquid phase separation (LLPS) by intermolecular electrostatic interactions in cells. Upon nutrient deprivation, genetically engineered Citrobacter freundii accumulates intracellular polyP at a rate of 210 µm min-1 , resulting in the compartmentation of +36GFP at the cell poles within 1 h. Medium chain-length polyP (60-mer) could induce the formation of +36GFP coacervates in vitro at a protein concentration as low as 200 nm, which is of the same magnitude as native proteins. In contrast, shorter polyP (14-mer) could not induce LLPS under the same conditions. This may offer a general approach to manipulate protein-protein interactions through LLPS.

Arsenene: A Potential Therapeutic Agent for Acute Promyelocytic Leukaemia Cells by Acting on Nuclear Proteins.

Arsenene has recently emerged as a promising new two-dimensional material for biomedical applications because of its excellent optical and electronic properties. Herein, novel 2D arsenene nanosheets were synthesized and shown to be effective against NB4 promyelocytic leukaemia (APL) cells (82 % inhibition) as well as inducing apoptosis while showing no toxicity towards normal cells. The high zeta potential, small size, and the planar structure were crucial to the toxicity of the materials. Label-free proteomic profiling analysis suggested that arsenene affected nuclear DNA replication, nucleotide excision repair, and pyrimidine metabolism pathways by downregulating the DNA polymerases POLE, POLD1, POLD2, and POLD3. Mass spectrometric studies showed that arsenene bound mainly to nuclear nucleotide acid binding proteins in NB4 cells and further cellular fluorescence studies revealed that the arsenene destroyed the nuclei. In vivo toxicity tests in mice also indicated the physiological biosafety of arsenene.

Melt-processable polyvinyl alcohol/lignin composites with improved strength via synergistic plasticization of lignin.

The characteristics of multi-hydroxyl structure and strong hydrogen bonding in polyvinyl alcohol (PVA) make its melting point close to its decomposition temperature, causing melt-processing difficulty. In this work, following the plasticization of small-molecule primary plasticizer acetamide, lignin was demonstrated as a green secondary plasticizer in realizing the melt processing and simultaneous reinforcement of PVA. During the plasticization process, lignin was able to combine with the hydroxyl groups of PVA, so as to destroy the hydrogen bonds and regularity of the PVA chains. The synergistic plasticization effect of lignin dramatically reduced the melting point of PVA from 185 °C to 151 °C. The thermal processing window of PVA composites was expanded from 50 °C to roughly 80 °C after introducing lignin. In contrast to acetamide, the addition of lignin significantly increased the tensile strength and Young's modulus of the composites to 71 MPa and 1.34 GPa, respectively. Meanwhile, lignin helped to hinder the migration of acetamide via hydrogen bonds. With the addition of lignin, the composites also displayed enhanced hydrophobicity and excellent UV shielding performance. The strategy of synergistic plasticization of lignin provides a feasible basis for the practical application of lignin in melt-processable PVA materials with good comprehensive properties.

Importance of secondary decomposition in the accurate prediction of daily-scale ozone pollution by machine learning.

Machine learning (ML) models have been proven as a reliable tool in predicting ambient pollution concentrations at various places in the world. However, their performance in predicting the maximum daily 8-h averaged ozone (MDA8 O3), the metric often used for O3 pollution assessment and management, is relatively poorer. This is largely resulted from more irregular data fluctuations of the MDA8 O3 levels governed collectively by the synoptic condition, local photochemistry, and long-range transport. In order to improve the prediction accuracy of MDA8 O3, this study developed a secondary decomposition ML model framework which coupled the complete ensemble empirical mode decomposition with adaptive noise (CEEMDAN) as the primary decomposition, the variational mode decomposition (VMD) as secondary decomposition, and the gate recurrent unit (GRU) ML model. By applying this secondary decomposition model framework on MDA8 O3 prediction for the first time, we showed that the prediction accuracy of MDA8 O3 is largely improved from R2 of 0.46 and RMSE of 30.4 µg/m3 for GRU without decomposition to R2 of 0.91 and RMSE of 12.6 µg/m3 over the Pearl River Delta of China. We also found that the prediction accuracy rate of O3 pollution non-attainments, an essential indicator for initiating contingency O3 pollution control, improved greatly from 14.9 % for GRU without decomposition to 72.5 %. The performance of O3 pollution non-attainment prediction is relatively higher in southwestern PRD, which is mainly due to greater number and severity of O3 non-attainments in southwestern cities located downwind of the emission hotspot area at central PRD. This study underscored the importance of secondary decomposition in accurately predicting daily-scale O3 concentration and non-attainments over the PRD, which can be extended to other photochemically active region worldwide to improve their O3 prediction accuracy and assist in O3 contingency control.

Exploiting the protein corona: coating of black phosphorus nanosheets enables macrophage polarization via calcium influx.

Black phosphorus nanosheets (BPNSs) have substantially promoted biomedical nanotechnology due to their unique photothermal and chemotherapeutic properties. However, there is still a limited molecular understanding of the effects of bio-nano interfaces on BPNSs and the subsequent impacts on physiological systems. Here, it is shown that black phosphorus-corona complexes (BPCCs) could function as immune modulators to promote the polarization of macrophages. Mechanistically, BPCCs could interact with calmodulin to activate stromal interaction molecule 2 and facilitate Ca2+ influx in macrophages, which induced the activation of p38 and NF-κB and polarized M0 macrophages to the M1 phenotype. As a result, BPCC-activated macrophages show greater migration towards cancer cells, 1.3-1.9 times higher cellular cytotoxicity and effective phagocytosis of cancer cells. These findings offer insights into the development of potential and unique applications of corona on BPNSs in nanomedicine.

Protein Corona and Immune Responses of Borophene: A Comparison of Nanosheet-Plasma Interface with Graphene and Phosphorene.

Borophene has emerged as a type of two-dimensional monoelemental nanomaterials with excellent drug loading capacity and photothermal properties. Here, we demonstrated the adsorption of plasma proteins onto borophene nanosheets (B NSs) and the promoted immune responses of macrophage by the B NS-corona complex. We discovered that plasma proteins changed the surface identities of B NSs. Using proteomics analysis, 46.5% of the proteins bound to B NSs (94 plasma proteins) were immune-relevant proteins. Uptake of B NSs by phagolysosomes was observed, and the plasma corona promoted the uptake. In comparison with graphene and phosphorene, we found that 32 plasma proteins appeared on all of the three nanosheets. The proportion of immune-relevant proteins in graphene-corona and phosphorene-corona was 41.3% and 75.6%, respectively. The components of the adsorbed immune-relevant proteins show diversity, which influence the immune responses of these nanosheets. Phosphorene-corona showed the most remarkable immunoregulatory behavior in these nanosheets. For the first time, we compared the highly complex protein corona at the nanosheet-plasma interface of three key 2D monoelemental nanosheets. Our study helps to understand the interaction between borophene and biological systems and provides a theoretical basis for the development and application of borophene in the biomedical field.

Proteome and transcriptome reveal the involvement of heat shock proteins and antioxidant system in thermotolerance of Clematis florida.

Clematis florida Thun (CfT) is an ornamental and medicinal plant. It is a cold resistant but heat sensitive species and deserves to be further investigated to improve its adaptability to heat stress. Exploring the molecular mechanism potential via an omic-analysis constitutes a promising approach towards improving heat tolerance of CfT. Two CfT lines, heat resistance (HR) and heat sensitive (HS), with differential thermotolerance capacities were used for the integrative analyses of proteomics and transcriptomes. Transcriptomes analysis showed that various pathways were significantly enriched including plant hormone signal transduction and carbon fixation pathways in prokaryotes. Proteomics study revealed the enrichment of some other pathways comprising antioxidant activity and carbohydrates metabolism. Based on combined transcriptomes and proteomics analyses and following heat stress treatment, a total of 1724 annotated genes were overlapped between both CfT lines. Particularly, 84 differential expressed genes (DEGs) were overlapped in both CfT lines. Fifteen out of these 84 genes were up-regulated solely for HR line (PS) but not for HS one (SG). This strongly suggests a potential prominent role for these genes in the thermotolerance process in PS line. We corroborate that two Hsps (Hsp18 and Hsp70) out of 20 detected proteins with higher expression levels in PS than in SG based on either global transcripts or proteins levels. According to the transcriptomes and proteomics analyses, 6 proteins and their corresponding genes were found to be significantly abundant in HR line (PS). Data are available via ProteomeXchange with identifier PXD018192. The expressions levels of these 6 genes were checked also for both CfT lines to evaluate their potential contributions in the heat tolerance process. Thus, their expression levels were approximately 2~4 times higher in HR than in HS line. We provided as well a representative schematic model to highlight the key genes involved in ROS scavenging and photorespiratory pathway in CfT. This model could be helpful also in understanding the mechanism of heat tolerance in CfT.

Revealing the immune perturbation of black phosphorus nanomaterials to macrophages by understanding the protein corona.

The increasing number of biological applications for black phosphorus (BP) nanomaterials has precipitated considerable concern about their interactions with physiological systems. Here we demonstrate the adsorption of plasma protein onto BP nanomaterials and the subsequent immune perturbation effect on macrophages. Using liquid chromatography tandem mass spectrometry, 75.8% of the proteins bound to BP quantum dots were immune relevant proteins, while that percentage for BP nanosheet-corona complexes is 69.9%. In particular, the protein corona dramatically reshapes BP nanomaterial-corona complexes, influenced cellular uptake, activated the NF-κB pathway and even increased cytokine secretion by 2-4-fold. BP nanomaterials induce immunotoxicity and immune perturbation in macrophages in the presence of a plasma corona. These findings offer important insights into the development of safe and effective BP nanomaterial-based therapies.

Controlled synthesis and size effects of multifunctional mesoporous silica nanosystem for precise cancer therapy.

Nanomaterials-based drug delivery systems display potent applications in cancer therapy, owing to the enhanced permeability and retention effect and diversified chemical modification. In this study, we have tailored and synthesized different sized mesoporous silica nanoparticles (MSNs) through reactant control to investigate the relevancy of nanoparticle size toward anticancer efficacy and suppressing cancer multidrug resistance. The different sized MSNs loaded with anticancer ruthenium complex (RuPOP) and conjugated with folate acid (FA) to enhance the selectivity between cancer and normal cells. The nanosystem (Ru@MSNs) can specifically recognize HepG2 hepatocellular carcinoma cells, thus enhance accumulation and selective cellular uptake. The smaller sized (20 nm) Ru@MSNs exhibit higher anticancer activity against HepG2 cells, while the larger sized (80 nm) Ru@MSNs exhibit higher inhibitory effect against DOX-resistant hepatocellular carcinoma cells (R-HepG2). Moreover, Ru@MSNs induced ROS overproduction in cancer cells, leading to DNA damage and p53 phosphorylation, consequently promoting cancer cells apoptosis. Ru@MSNs (80 nm) also inhibited ABCB1 and ABCG2 expression in R-HepG2 cells to prevent drug efflux, thus overcome multidrug resistance. Ru@MSNs also inhibited tumor growth in vivo without obvious toxicity in major organs of tumor-bearing nude mice. Taken together, these results verify the size effects of MSNs nanosystem for precise cancer therapy.

Tailoring Particle Size of Mesoporous Silica Nanosystem To Antagonize Glioblastoma and Overcome Blood-Brain Barrier.

The blood-brain barrier (BBB) is the main bottleneck to prevent some macromolecular substance entering the cerebral circulation, resulting the failure of chemotherapy in the treatment of glioma. Cancer nanotechnology displays potent applications in glioma therapy owing to their penetration across BBB and accumulation into the tumor core. In this study, we have tailored the particle size of mesoporous silica nanoparticles (MSNs) through controlling the hydrolysis rate and polycondensation degree of reactants, and optimized the nanosystem that could effectively penetrate BBB and target the tumor tissue to achieve enhanced antiglioma efficacy. The nanoparticle was conjugated with cRGD peptide to enhance its cancer targeting effect, and then used to load antineoplastic doxorubicin. Therefore, the functionalized nanosystem (DOX@MSNs) selectively recognizes and binds to the U87 cells with higher expression level of ανß3 integrin, sequentially enhancing the cellular uptake and inhibition to glioma cells, especially the particle size at 40 nm. This particle could rapidly enter cancer cells and was difficult to excrete outside the cells, thus leading to high drug accumulation. Furthermore, DOX@MSNs exhibited much higher selectivity and anticancer activity than free DOX and induced the glioma cells apoptosis through triggering ROS overproduction. Interestingly, DOX@MSNs at about 40 nm exhibited stronger permeability across the BBB, and could disrupt the VM-capability of glioma cells by regulating the expression of E-cadherin, FAK, and MMP-2, thus achieving satisfactory antiglioblastoma efficacy and avoiding the unwanted toxic side effects to normal brain tissue. Taken together, these results suggest that tailoring the particle size of MSNs nanosystem could be an effective strategy to antagonize glioblastoma and overcome BBB.