Explore the vast range of titles available.

Riemerella anatipestifer infection is a critical disease that is a major threat to the poultry industry worldwide. The adhesion and invasion of host cells are key steps in the primary stages of bacterial infection. However, the outer membrane proteins that mediate these events in R. anatipestifer are poorly characterized. In this study, the PorV and PosF proteins, as well as the previously described OMP71 protein, were identified as important mediators of the adhesion and invasion of duck embryo fibroblast (DEF) cells by R. anatipestifer . Affinity chromatography-based surface proteomics was used to screen for adhesion proteins. The surface proteins on DEF cells were labelled with biotin-avidin to enrich for outer membrane proteins of R. anatipestifer, which generated 11 candidate proteins that were tested further. Protein adhesion and blocking assays and polyclonal antiserum inhibition analysis revealed that the PorV, PosF, and OMP71 proteins are adhesion factors. Knockout of porV or posF reduced the adhesion and invasion of R. anatipestifer in DEF cells. Moreover, the pathogenicity of the mutant strains was significantly attenuated, which supports the hypothesis that PorV and PosF are important virulence factors required for the pathogenicity of R. anatipestifer . The PorV protein is a key component of the type IX secretory system and is responsible for transporting effector substrates to the extracellular environment, whereas PosF belongs to the porin superfamily of barrel-shaped transmembrane proteins. This is the first description that PorV is an adhesin involved in host‒microbial interactions, which represents a breakthrough in pathogenicity studies of R. anatipestifer and other members of Flavobacteriaceae .

To explore and isolate genes related to duck embryonic fibroblast cells (DEFs) post-infected with duck enteritis virus (DEV), a cDNA library was established using SMART (Switching Mechanism At 5′ end of the RNA Transcript) technique coupling with a homologous recombination method. The cells were harvested and total RNA was extracted at 48 h post infection. Then the mRNAs were purified and reverse transcribed to first-strand cDNAs using oligo (dT) primers (CDS III). Subsequently, long distance-PCR was performed, the double-stranded cDNAs were purified, and a transformation assay was carried out in that order. Eventually, a high qualitative library was successfully established according to an evaluation on quality. The transformation efficiency was about 2.33 × 10 6 transformants/4.34 μg pGADT7-Rec (>1.0 × 10 6 ). The cell density of the library was 1.75 × 10 9  cells/mL (>2×10 7  cells/mL). The titer of the primary cDNA library and amplified cDNA library was 6.75 × 10 5 and 2.33 × 10 7  CFU/mL respectively. The numbers for the primary cDNA library and amplified cDNA library were 1.01 × 10 7 and 1.14 × 10 9 , respectively, and the recombinant rate was 97.14 %. The sequence results of 27 randomly picked independent clones revealed the insert ranged from 0.323 to 2.017 kb with an average insert size of 0.807 kb. Full-length transcripts of DEV-CHv LORF3, UL26 and UL35 genes were acquired through sequence similarity analysis from the non-redundant nucleic acid or protein database. Five polyA sites were identified in the DEV-CHv genome. Also, a new transcript of 668 bp was found between the IRS gene and US1 gene of the DEV-CHv genome. Thus, we concluded that the constructed cDNA library will be a useful tool in proteomic analysis of interactions between the DEV and host DEFs, and discovery of biomarkers studies on the mechanism of DEV and subsequently exploitation original vaccines and antiviral drugs to prevent or cure diseases.

p53 protein turnover through the ubiquitination pathway is a vital mechanism in the regulation of its transcriptional activity; however, little is known about p53 turnover through proteasome-independent pathway（s）. The diges tive organ expansion factor （Det） protein is essential for the development of digestive organs. In zebrafish, loss of function of defselectively upregulates the expression of p53 response genes, which raises a question as to what is the relationship between Def and p53. We report here that Def is a nucleolar protein and that loss of function of defleads to the upregulation of p53 protein, which surprisingly accumulates in the nucleoli. Our extensive studies have dem- onstrated that Def can mediate the degradation of p53 protein and that this process is independent of the proteasome pathway, but dependent on the activity of Calpain3, a cysteine protease. Our findings define a novel nucleolar path- way that regulates the turnover function of p53, which will advance our understanding of p53＇s role in organogenesis and tumorigenesis.

Context In this study, we evaluate the geometrical, absorption, optoelectronic, electronic, nonlinear optical (NLO) and thermodynamic properties of dibenzo[b,def]chrysene molecule derivatives by means of DFT and TD-DFT simulations. In view of the aim of producing new high-performance materials for non-linear optics (NLO) by doping test, two types of doping were used. We obtained six derivatives by doping with organic dopants (Nitro, amide and ticyanoethenyl) and mixed alkali metal (potassium) and organic dopants. Doping with organic dopants produced molecules A, B and C, respectively when substituting one hydrogen with nitro (NO 2 ), amide (CONH 2 ) and tricyanoethenyl (C 5 N 3 ) groups, while mixed doping involved considering A, B and C and then substituting two hydrogens with two potassiums to obtain compounds D, E and F respectively. The negative values of the various interaction energies calculated for all the doped molecules show that they are all stable, but also that molecules C and F are the most stable in the case of both dopings. The gap energies calculated at the B3LYP level of theory are all below 3 eV, which means that all the molecules obtained are semiconductors. Better still, compounds C and F, with gap energies of 1.852 eV and 1.204 eV, respectively, corresponding to decreases of 35.67% and 58.18% in gap energy compared with the pristine molecule, are more reactive than the other doped molecules. Mixed doping is therefore a highly effective way of narrowing the energy gap and boosting the semiconducting character and reactivity of organic materials. Optoelectronic properties have also been improved, with refractive index values higher than those of the reference material, glass. This shows that our compounds could be used under very high electric field conditions of the order of 4.164 × 10 9  V.m −1 for C and 7.410 × 10 9  V.m −1 for F the highest values at the B3LYP level of theory. The maximum first-order hyperpolarizability values for both types of doping are obtained at the CAM-B3LYP level of theory by C: β mol = 92.088 × 10 −30 esu and by F: β mol = 129.449 × 10 −30 esu, and second-order values are also given by these same compounds. These values are higher than the reference value, which is urea, making our compounds potential candidates for high-performance NLO applications. In dynamic mode and at a frequency of 1064 nm, at the CAM-B3LYP level of theory, the highest dynamic hyperpolarizability coefficients were obtained by C and F. Hyper-Rayleigh scattering β HRS , coefficients of the electro-optical Pockel effect (EOPE), EFISHG, third-order NLO-response degree four-wave mixing γ DFWM , quadratic nonlinear refractive index n2 were also calculated. The maximum values of n 2 are obtained by C (6.13 × 10 –20 m 2 /W) and F (6.60 × 10 –20 m 2 /W), these values are 2.24 times higher than that of fused silica which is the reference for degenerate four-wave mixing so our molecules could also have applications in optoelectronics as wavelength converters, optical pulse modulators and optical switches. Methods Using the DFT method, we were able to determine the optimized and stable electronic structures of doped dibenzo[b,def]chrysene derivatives in the gas phase. We limited ourselves to using the proven B3LYP and CAMB3LYP levels of theory for calculating electronic properties, and non-linear optics with the 6-311G +  + (d,p) basis set, which is a large basis set frequently used for these types of compound. Gaussian 09 software was used to run our calculations, and Gauss View 6.0.16 was used to visualize the output files. TD-DFT was also used to determine absorption properties at the B3LYP level of theory, using the same basis set.

Recovery of liver mass to a healthy liver donor by compensatory regeneration after partial hepatectomy (PH) is a prerequisite for liver transplantation. Synchronized cell cycle reentry of the existing hepatocytes after PH is seemingly a hallmark of liver compensatory regeneration. Although the molecular control of the PH-triggered cell cycle reentry has been extensively studied, little is known about how the synchronization is achieved after PH. The nucleolus-localized protein cleavage complex formed by the nucleolar protein Digestive-organ expansion factor (Def) and cysteine proteinase Calpain 3 (Capn3) has been implicated to control wounding healing during liver regeneration through selectively cleaving the tumor suppressor p53 in the nucleolus. However, whether the Def-Capn3 complex participates in regulating the synchronization of cell cycle reentry after PH is unknown. In this report, we generated a zebrafish capn3b null mutant ( capn3b ∆19∆14 ). The homozygous mutant was viable and fertile, but suffered from a delayed liver regeneration after PH. Delayed liver regeneration in capn3b ∆19∆14 was due to disruption of synchronized cell proliferation after PH. Mass spectrometry (MS) analysis of nuclear proteins revealed that a number of negative regulators of cell cycle are accumulated in the capn3b ∆19∆14 liver after PH. Moreover, we demonstrated that Check-point kinase 1 (Chk1) and Wee1, two key negative regulators of G2 to M transition, are substrates of Capn3. We also demonstrated that Chk1 and Wee1 were abnormally accumulated in the nucleoli of amputated capn3b ∆19∆14 liver. In conclusion, our findings suggest that the nucleolar-localized Def-Capn3 complex acts as a novel regulatory pathway for the synchronization of cell cycle reentry, at least partially, through inactivating Chk1 and Wee1 during liver regeneration after PH.

Digestive organs originate from the endoderm. Morphogenesis of the digestive system is precisely controlled by multiple factors that dictate the cell fate and behavior so that the specific digestive organs are timely formed in the right place and develop into right size and structure. We showed previously that digestive organ expansion factor (def) is a gene whose expression is enriched in the liver, pancreas and intestine. Loss-of-function of def in the def(hi429) mutant confers hypoplastic digestive organs partly due to alteration of expression of genes related to the p53 pathway. However, the molecular mechanism for the involvement of Def in the organogenesis of digestive organs is still largely unknown. For example, it is not known whether Def regulates specific pathways in a specific organ. To address this question, we generated four independent Tg(fabp10a:def) transgenic fish lines which over-expressed Def specifically in the liver. We characterized Tg-I, one of the transgenic lines, in detail with genetic, molecular and histological approaches. We found that Tg-I restored the liver but not exocrine pancreas and intestine development in the def(hi429) mutant. However, Tg-I adult fish in the wild type (WT) background exhibits reduced liver-to-body ratio and all four transgenic lines conferred abnormal intrahepatic structure. Microarray data analysis showed that certain specific functional pathways were affected in the liver of Tg-I. These results demonstrate that Def functions in a cell autonomous manner during early liver development and aberrant Def protein expression might lead to disruption of the structural integrity of a normal adult liver.

Arsenite (As⁺³) and dibenzo[def,p]chrysene (DBC), a polycyclic aromatic hyrdrocarbon (PAH), are found in nature as environmental contaminants. Both are known to individually suppress the immune system of humans and mice. In order to determine their potential interactive and combined immunosuppressive effects, we examined murine bone marrow (BM) immune progenitor cells’ responses following combined oral exposures at very low levels of exposure to As⁺³ and DBC. Oral 5-day exposure to DBC at 1 mg/kg (cumulative dose) was found to suppress mouse BM lymphoid progenitor cells, but not the myeloid progenitors. Previously established no-effect doses of As⁺³ in drinking water (19 and 75 ppb for 30 days) produced more lymphoid suppression in the bone marrow when mice were concomitantly fed a low dose of DBC during the last 5 days. The lower dose (19 ppb) As⁺³ had a stronger suppressive effect with DBC than the higher dose (75 ppb). Thus, the interactive toxicity of As⁺³ and DBC in vivo could be As⁺³ dose dependent. In vitro, the suppressive interaction of As⁺³ and DBC was also evident at low concentrations (0.5 nM), but not at higher concentrations (5 nM) of As⁺³. These studies show potentially important interactions between As⁺³ and DBC on mouse BM at extremely low levels of exposure in vivo and in vitro.

BackgroundDue to their unique properties, gold nanoparticles (GNPs) have been proposed to be used for a wide range of applications, especially for photon radiation therapy. In addition to experimental works, there are worthwhile simulation-based studies focused on the investigation of the effect of parameters governing the dose enhancement due to the presence of GNPs in tissue. In a recently published study, we found that the distribution of GNPs in a single cell plays an important role in nucleus dose enhancement.MethodsThe present work investigates the sensitivity of dose enhancement of a macroscopic phantom to the modeling of GNPs at the cellular level by using the MCNPX Monte Carlo code. A human eye phantom containing the realistic structures and materials was simulated, with a typical tumor located in its corner filled with three different patterns of distribution of GNPs around the nuclei of the cells. The primary photons emit from a COMS eye plaque brachytherapy containing thirteen 131Cs seeds in the vicinity of the tumor.ResultsThe study was extended to estimate dose enhancement for various concentration, size, and density of the GNPs accumulated around the nuclei of the tumor. Moreover, the dose delivered to the healthy eye structures for different models has been investigated and discussed. The results show obvious differences between the dose enhancements in the tumor depending on the modeling of GNPs.ConclusionThe results emphasized that an appropriate small-scale model for the distribution of GNPs in the cell would be of high importance to estimate the degree of dose enhancement in a macroscopic phantom to provide a trustworthy prediction to move towards clinical application.

The intracellular levels of the p53 tumor suppressor protein are regulated through various pathways and involve numerous regulatory components. A recent study published in CellResearch identifies a proteasomeindependent pathway of p53 protein degradation in the nucleolus that is dependent on Def and Calpain3.

Peptide deformylase (PDF) is considered an attractive target for screening novel antibiotics. The PDF from Escherichia coli and Staphylococcus aureus are representative of the gram-negative species type of PDF (type I PDF) and the gram-positive species type of PDF (type II PDF), respectively. They could be used for screening broad-spectrum antibiotics. Herein, we cloned the def gene by PCR, inserted it into plasmid pET-22b-def, and transformed the plasmid into E. coli BL21 (DE3) cells, then the cells were induced by IPTG to express PDF. E. coli Ni 2+ -PDF was extracted and purified by ion-exchange chromatography and gel filtration chromatography. S. aureus PDFs were extracted and purified using the MagExtractor kit. The nickel form of S. aureus PDF was obtained by adding NiCl 2 to all reagents used for purification. Iron-enriched S. aureus PDF was obtained by adding FeCl 3 to the growth medium for E. coli BL21 (DE3) cells and adding FeCl 3 and catalase to all reagents used for purification. The activities of PDFs were analyzed, compared, and grouped according to the experimental conditions that produced optimal activity, and we used actinonin as an inhibitor of PDF and calculated the IC 50 value. We obtained high expression of E. coli and S. aureus PDF with high activity and stability. The function of PDFs was inhibited by actinonin in a dose-dependent manner. Results may be helpful for future mechanistic investigations of PDF as well as high-throughput screening for other PDF inhibitors.