Genome-wide analysis and identification of stress-responsive genes of the CCCH zinc finger family in Capsicum annuum L.

The CCCH zinc finger gene family encodes a class of proteins that can bind to both DNA and RNA, and an increasing number of studies have demonstrated that the CCCH gene family plays a key role in growth and development and responses to environmental stress. Here, we identified 57 CCCH genes in the pepper (Capsicum annuum L.) genome and explored the evolution and function of the CCCH gene family in C. annuum. Substantial variation was observed in the structure of these CCCH genes, and the number of exons ranged from one to fourteen. Analysis of gene duplication events revealed that segmental duplication was the main driver of gene expansion in the CCCH gene family in pepper. We found that the expression of CCCH genes was significantly up-regulated during the response to biotic and abiotic stress, especially cold and heat stress, indicating that CCCH genes play key roles in stress responses. Our results provide new information on CCCH genes in pepper and will aid future studies of the evolution, inheritance, and function of CCCH zinc finger genes in pepper

The Preliminary Exploration of Micro-Friction Stir Welding Process and Material Flow of Copper and Brass Ultra-Thin Sheets

In the friction stir welding (FSW) of ultra-thin dissimilar metal sheets, different physical material properties, the reduction of plastic metal in the weld zone, and insufficient plastic metal flow lead to poor weld seam shapes and joint qualities. Therefore, it is necessary to study the flow behavior during the FSW of ultrathin sheets. In this study, micro friction stir welding (μFSW) was conducted and analyzed for the butt welding of 0.6-mm-thick ultrathin brass (H62-H) and pure copper (T2-Y) sheets. By analyzing the electric signals of the temperature and force during the welding process, testing the mechanical properties, and analyzing the metallography of the joint, the influences of the process parameters on the metal flow behavior during μFSW were studied. In the proper process conditions, the material preferentially migrated and concentric vortex flow occurred in the vicinity of the shoulder and tool pin action areas. The copper was pushed from the retreating side (RS) to the advancing side (AS) of the weld, allowing it to flow more fully. A mixture of both materials formed at the bottom of the weld nugget, and less migration occurred in the heat-affected zone of the AS at this time. The highest tensile strength can reach 194 MPa, accounting for 82.6% of the copper. The presence of brittle phases Cu5Zn8, AgZn3 and AgZn caused the hardness to fluctuate slightly

The Cellulosome System of Acetivibrio cellulolyticus Includes a Novel Type of Adaptor Protein and a Cell Surface Anchoring Protein

A scaffoldin gene cluster was identified in the mesophilic cellulolytic anaerobe Acetivibrio cellulolyticus. The previously described scaffoldin gene, cipV, encodes an N-terminal family 9 glycoside hydrolase, a family 3b cellulose-binding domain, seven cohesin domains, and a C-terminal dockerin. The gene immediately downstream of cipV was sequenced and designated scaB. The protein encoded by this gene has 942 amino acid residues and a calculated molecular weight of 100,358 and includes an N-terminal signal peptide, four type II cohesions, and a C-terminal dockerin. ScaB cohesins 1 and 2 are very closely linked. Similar, but not identical, 39-residue Thr-rich linker segments separate cohesin 2 from cohesin 3 and cohesin 3 from cohesin 4, and an 84-residue Thr-rich linker connects the fourth cohesin to a C-terminal dockerin. The scaC gene downstream of scaB codes for a 1,237-residue polypeptide that includes a signal peptide, three cohesins, and a C-terminal S-layer homology (SLH) module. A long, ca. 550-residue linker separates the third cohesin and the SLH module of ScaC and is characterized by an 18-residue Pro-Thr-Ala-Ser-rich segment that is repeated 27 times. The calculated molecular weight of the mature ScaC polypeptide (excluding the signal peptide) is 124,162. The presence of the cohesins and the conserved SLH module implies that ScaC acts as an anchoring protein. The ScaC cohesins are on a separate branch of the phylogenetic tree that is close to, but distinct from, the type I cohesins. Affinity blotting with representative recombinant probes revealed the following specific intermodular interactions: (i) an expressed CipV cohesin binds selectively to an enzyme-borne dockerin, (ii) a representative ScaB cohesin binds to the CipV band of the cell-free supernatant fraction, and (iii) a ScaC cohesin binds to the ScaB dockerin. The experimental evidence thus indicates that CipV acts as a primary (enzyme-recognizing) scaffoldin, and the protein was also designated ScaA. In addition, ScaB is thought to assume the role of an adaptor protein, which connects the primary scaffoldin (ScaA) to the cohesin-containing anchoring scaffoldin (ScaC). The cellulosome system of A. cellulolyticus thus appears to exhibit a special type of organization that reflects the function of the ScaB adaptor protein. The intercalation of three multiple cohesin-containing scaffoldins results in marked amplification of the number of enzyme subunits per cellulosome unit. At least 96 enzymes can apparently be incorporated into an individual A. cellulolyticus cellulosome. The role of such amplified enzyme incorporation and the resultant proximity of the enzymes within the cellulosome complex presumably contribute to the enhanced synergistic action and overall efficient digestion of recalcitrant forms of cellulose. Comparison of the emerging organization of the A. cellulolyticus cellulosome with the organizations in other cellulolytic bacteria revealed the diversity of the supramolecular architecture

Bromide Decorated Eco‐Friendly ZnSeTe/ZnSe/ZnS Quantum Dots for Efficient Blue Light‐Emitting Diodes

Abstract Multi‐shelled ZnSeTe/ZnSe/ZnS quantum dots (QDs) have served as a promising eco‐friendly emitter for blue quantum dot light‐emitting diodes (QLEDs). While extensive studies have concentrated on the optimization of the shell species and thickness of the multi‐shelled QDs to raise the QLED electroluminescent performance, very few reports focus on the QD surface states involving ligand and defect modulations which are essential for high‐performance QLEDs. Herein, the strategy of bromide decoration is theoretically and practically demonstrated to simultaneously diminish the QD surface defects by passivating the unsaturated Zn for strengthened carrier radiative recombination and removing the superabundant oleic acid through ligand exchange for efficient carrier transport. As a result, the merits of bromide decoration benefit a large increase in photoluminescence quantum yield (PLQY) from 39.7% to 86.2% at the wavelength of 443 nm, as well as a great enhancement of the device performance with over sevenfold improvement in external quantum efficiency (EQE) from 0.74% to 5.46% and a distinct decrease in turn‐on voltage from 6.7 to 5.9 V. Consequently, this work contributes an effective approach of the multi‐shelled QD surface decoration toward enhanced QLED performance

The Combination of R848 with Sorafenib Enhances Antitumor Effects by Reprogramming the Tumor Immune Microenvironment and Facilitating Vascular Normalization in Hepatocellular Carcinoma

Abstract Novel promising strategies for combination with sorafenib are urgently needed to enhance its clinical benefit and overcome toxicity in hepatocellular carcinoma (HCC). the molecular and immunomodulatory antitumor effects of sorafenib alone and in combination with the new immunotherapeutic agent R848 are presented. Syngeneic HCC mouse model is presented to explore the antitumor effect and safety of three sorafenib doses alone, R848 alone, or their combination in vivo. R848 significantly enhances the sorafenib antitumor activity at a low subclinical dose with no obvious toxic side effects. Furthermore, the combination therapy reprograms the tumor immune microenvironment by increasing antitumor macrophages and neutrophils and preventing immunosuppressive signaling. Combination treatment promotes classical M1 macrophage‐to‐FTH1high M1 macrophage transition. The close interaction between neutrophils/classical M1 macrophages and dendritic cells promotes tumor antigen presentation to T cells, inducing cytotoxic CD8+ T cell‐mediated antitumor immunity. Additionally, low‐dose sorafenib, alone or combined with R848, normalizes the tumor vasculature, generating a positive feedback loop to support the antitumor immune environment. Therefore, the combination therapy reprograms the HCC immune microenvironment and normalizes the vasculature, improving the therapeutic benefit of low‐dose sorafenib and minimizing toxicity, suggesting a promising novel immunotherapy (R848) and targeted therapy (tyrosine kinase inhibitors) combination strategy for HCC treatment

NEDD4-1 Regulates Migration and Invasion of Glioma Cells through CNrasGEF Ubiquitination In Vitro

<div><p>Neuronal precursor cell-expressed developmentally down-regulated 4-1 (NEDD4-1) plays a great role in tumor cell growth, but its function and mechanism in cell invasive behavior are totally unknown. Here we report that NEDD4-1 regulates migration and invasion of malignant glioma cells via triggering ubiquitination of cyclic nucleotide Ras guanine nucleotide exchange factor (CNrasGEF) using cultured glioma cells. NEDD4-1 overexpression promoted cell migration and invasion, while its downregulation specifically inhibited them. However, NEDD4-1 did not affect the proliferation and apoptosis of glioma cells. NEDD4-1 physically interacted with CNrasGEF and promoted its poly-ubiquitination and degradation. Contrary to the effect of NEDD4-1, CNrasGEF downregulation promoted cell migration and invasion, while its overexpression inhibited them. Importantly, downregulation of CNrasGEF facilitated the effect of NEDD4-1-induced cell migration and invasion. Interestingly, aberrant up-regulated NEDD4-1 showed reverse correlation with CNrasGEF protein level but not with its mRNA level in glioma tissues. Combined with the in vitro results, the result of glioma tissues indicated post-translationally modification effect of NEDD4-1 on CNrasGEF. Our study suggests that NEDD4-1 regulates cell migration and invasion through ubiquitination of CNrasGEF in vitro. </p> </div

The effect of NEDD4-1 on cell migration and invasion.

<p>A) NEDD4-1 overexpressing or downregulating efficacy in U251 glioma cells examined by RT-PCR. Left, Representative RT-PCR analysis. Twenty-four hours after transfection, total RNAs were extracted and underwent RT-PCR analysis. β-actin was used as internal control. Right, Quantitative analysis of relative mRNA levels of NEDD4-1 normalized to those of β-actin. B) NEDD4-1 overexpressing or downregulating efficacy in U251 glioma cells detected by western blotting. Left, Representative image of western blotting. Forty-eight hours after transfection, cells were lysed and protein extraction was underwent western blot analysis using NEDD4-1 antibody. β-actin was used as the loading control. Right, Quantitative analysis of relative protein levels of NEDD4-1 normalized to those of β-actin. C) Wound-healing assay of glioma U251 cells after NEDD4-1 overexpression or downregulation. Representative digital pictures were taken at 0h and 24h. Bar: 100 μm. D) Quantitative analysis of the number of migratory cells. E) Transwell invasion assay of glioma U251 cells after NEDD4-1 overexpression or downregulation. After 48 hours of transfection, cell suspension was added into the matrigel precoated transwell chambers and the cells invaded through the matrigel were stained and photoed. Bar: 50 μm. F) Quantitative analysis of the number of invasive cells. G) Wound-healing (up and middle panel) and transwell invasion assay (bottom panel) of glioma U87 cells after NEDD4-1 overexpression or downregulation. Bar: 100 μm. H & I) Quantitative analysis of the number of migratory (H) or invasive (I) cells. Results are mean ± SEM of three independent experiments in triplicate. *<i>P</i><0.05.</p

The effect of CNrasGEF on cell migration and invasion.

<p>A) Representative western blotting analysis of CNrasGEF knocking-down or overexpression efficacy in U251 glioma cells. B) Quantitative analysis of relative protein levels of CNrasGEF normalized to those of β-actin. C) Wound-healing assay (up and middle panel) and transwell invasion assay (bottom panel) of glioma U251 cells transfected with siRNAs or overexpression of CNrasGEF. D & E) Quantitative analysis of the number of migratory (D) or invasive (E) cells. F) Wound-healing (up and middle panel) and transwell invasion assay (bottom panel) of glioma U87 cells after CNrasGEF downregulation or overexpression. G & H) Quantitative analysis of the number of migratory (G) or invasive (H) cells. Data were mean ± SEM of three independent experiments in triplicate. * <i>P</i><0.05.Bar: 100 μm.</p

Expression of NEDD4-1 and CNrasGEF in glioma tissues.

<p>A) mRNA levels of NEDD4-1 and CNrasGEF in glioma and nontumorous brainspecimens was detected by RT-PCR. B) Quantitative analysis of the mRNA levels of NEDD4-1 and CNrasGEF normalized to those of β-actin. C) The mRNA relationship between the NEDD4-1 and CNrasGEF. D) Western blotting analysis of the total extracts isolated from human nontumorous brain and human glioma specimens using human NEDD4-1 and CNrasGEF antibodies. E) Quantitative analysis of the protein levels of NEDD4-1 and CNrasGEF normalized to those of β-actin. F) The protein relationship between the NEDD4-1 and CNrasGEF. r=-0.79. I) Representative hematoxylin-eosin staining and immunohistochemistry (IHC) analysis of NEDD4-1 and CNrasGEF expression in nontumorous brain tissue and glioma specimens of varying World Health Organization (WHO) grades. Normal brain tissue (a); diffuse astrocytoma, grade II (b); anaplastic astrocytoma, grade III (c); GBM, grade IV (d) are shown. IHC stainings were performed at least twice with similar stained patterns. Scale bar:100μm. J&K) Quantitativeanalysis of the percentage of NEDD4-1 and CNrasGEF positive cells. Data were derived from three independent experiments. ** <i>P</i><0.01 vs nontumorous.</p