Transcription factor Foxm1 plays a critical role during embryonic development and its expression is repressed during retinoic acid (RA)-induced differentiation of pluripotent P19 embryonal carcinoma cells at the early stage, correlated with downregulation of expression of pluripotency markers. To study whether Foxm1 participates in the maintenance of pluripotency of stem cells, we knock down Foxm1 expression in P19 cells and identify that Oct4 are regulated directly by Foxm1. Knockdown of Foxm1 also results in spontaneous differentiation of P19 cells to mesodermal derivatives, such as muscle and adipose tissues. Maintaining Foxm1 expression prevents the downregulation of pluripotency-related transcription factors such as Oct4 and Nanog during P19 cell differentiation. Furthermore, overexpression of FOXM1 alone in RA-differentiated P19 cells (4 days) or human newborn fibroblasts restarts the expression of pluripotent genes Oct4, Nanog and Sox2. Together, our results suggest a critical involvement of Foxm1 in maintenance of stem cell pluripotency.

The genetic diversity and relationships among ethnic minority populations of southwest China were investigated using seven polymorphic restriction enzyme sites in the β-globin gene cluster. The haplotypes of 1392 chromosomes from ten ethnic populations living in southwest China were determined. Linkage equilibrium and recombination hotspot were found between the 5' sites and 3' sites of the β-globin gene cluster. 5' haplotypes 2 (+---), 6 (-++-+), 9 (-++++) and 3' haplotype FW3 (-+) were the predominant haplotypes. Notably, haplotype 9 frequency was significantly high in the southwest populations, indicating their difference with other Chinese. The interpopulation differentiation of southwest Chinese minority populations is less than those in populations of northern China and other continents. Phylogenetic analysis shows that populations sharing same ethnic origin or language clustered to each other, indicating current β-globin cluster diversity in the Chinese populations reflects their ethnic origin and linguistic affiliations to a great extent. This study characterizes β-globin gene cluster haplotypes in southwest Chinese minorities for the first time, and reveals the genetic variability and affinity of these populations using β-globin cluster haplotype frequencies. The results suggest that ethnic origin plays an important role in shaping variations of the β-globin gene cluster in the southwestern ethnic populations of China.

The tea green leafhopper Empoasca onukii Matsuda (Hemiptera: Cicadellidae), the orange spiny whitefly, Aleurocanthus spiniferus (Quaintanca) (Hemiptera: Aleyrodidae), and the green plant bugs Apolygus lucorum Meyer-Dür (Hemiptera: Miridae) are the important piercing-sucking herbivores in tea trees Camellia sinensis (L.) O. Kuntze (Theaceae). The goal of this study was to evaluate the laboratory toxicities and field control efficacies of botanical insecticides including matrine, azadirachtin, veratrine, and pyrethrin to three tea pests. Via leaf-dip bioassay, toxicity tests with botanical insecticides indicated that there were significant differences between the LC50 values for botanical insecticides within the same insect species. Matrine had the highest toxicity to E. onukii, A. spiniferus, and A. lucorum with the LC50 values of 2.35, 13.10, and 44.88 mg/liter, respectively. Field tests showed that, among four botanical insecticides, matrine at dose of 9 g a.i. ha-1 can significantly reduce the numbers of E. onukii and A. spiniferus and the infestation of A. lucorum on the tea plants. Furthermore, botanical insecticides matrine and azadirachtin had no obvious influence on the coccinellids, spiders, and parasitoids densities in tea plantations. The results of this study indicated that use of botanical insecticides, such as matrine, has the potential to manipulate the population of E. onukii, A. spiniferus, and A. lucorum and will be an effective and environmentally compatible strategy for the control of tea pests.

Biological control mechanisms of plant diseases have been intensively studied. However, how plant pathogens respond to and resist or alleviate biocontrol agents remains largely unknown. In this study, a comparative transcriptome analysis was performed to elucidate how the pathogen of sclerotinia stem rot, Sclerotinia sclerotiorum, responds and resists to the biocontrol agent, Bacillus amyloliquefaciens. Results revealed that a total of 2,373 genes were differentially expressed in S. sclerotiorum samples treated with B. amyloliquefaciens fermentation broth (TS) when compared to control samples (CS). Among these genes, 2,017 were upregulated and 356 were downregulated. Further analyses indicated that various genes related to fungal cell wall and cell membrane synthesis, antioxidants, and the autophagy pathway were significantly upregulated, including glucan synthesis, ergosterol biosynthesis pathway, fatty acid synthase, heme-binding peroxidase related to oxidative stress, glutathione S-transferase, ABC transporter, and autophagy-related genes. These results suggest that S. sclerotiorum recruits numerous genes to respond to or resist the biocontrol of B. amyloliquefaciens. Thus, this study serves as a valuable resource regarding the mechanisms of fungal pathogen resistance to biocontrol agents.

The recognition of the scorpion toxin maurotoxin (MTX) by the voltage-gated potassium (Kv1) channels, Kv1.1, Kv1.2, and Kv1.3, has been studied by means of Brownian dynamics (BD) simulations. All of the 35 available structures of MTX in the Protein Data Bank (http://www.rcsb.org/pdb) determined by nuclear magnetic resonance were considered during the simulations, which indicated that the conformation of MTX significantly affected both the recognition and the binding between MTX and the Kv1 channels. Comparing the top five highest-frequency structures of MTX binding to the Kv1 channels, we found that the Kv1.2 channel, with the highest docking frequencies and the lowest electrostatic interaction energies, was the most favorable for MTX binding, whereas Kv1.1 was intermediate, and Kv1.3 was the least favorable one. Among the 35 structures of MTX, the 10th structure docked into the binding site of the Kv1.2 channel with the highest probability and the most favorable electrostatic interactions. From the MTX-Kv1.2 binding model, we identified the critical residues for the recognition of these two proteins through triplet contact analyses. MTX locates around the extracellular mouth of the Kv1 channels, making contacts with its beta-sheets. Lys23, a conserved amino acid in the scorpion toxins, protrudes into the pore of the Kv1.2 channel and forms two hydrogen bonds with the conserved residues Gly401(D) and Tyr400(C) and one hydrophobic contact with Gly401(C) of the Kv1.2 channel. The critical triplet contacts for recognition between MTX and the Kv1.2 channel are Lys23(MTX)-Asp402(C)(Kv1), Lys27(MTX)-Asp378(D)(Kv1), and Lys30(MTX)-Asp402(A)(Kv1). In addition, six hydrogen-bonding interactions are formed between residues Lys23, Lys27, Lys30, and Tyr32 of MTX and residues Gly401, Tyr400, Asp402, Asp378, and Thr406 of Kv1.2. Many of them are formed by side chains of residues of MTX and backbone atoms of the Kv1.2 channel. Five hydrophobic contacts exist between residues Pro20, Lys23, Lys30 and Tyr32 of MTX and residues Asp402, Val404, Gly401, and Arg377 of the Kv1.2 channel. The simulation results are in agreement with the previous molecular biology experiments and explain the binding phenomena between MTX and Kv1 channels at the molecular level. The consistency between the results of the BD simulations and the experimental data indicated that our three-dimensional model of the MTX-Kv1.2 channel complex is reasonable and can be used in additional biological studies, such as rational design of novel therapeutic agents blocking the voltage-gated channels and in mutagenesis studies in both the toxins and the Kv1 channels. In particular, both the BD simulations and the molecular mechanics refinements indicate that residue Asp378 of the Kv1.2 channel is critical for its recognition and binding functionality toward MTX. This phenomenon has not been appreciated in the previous mutagenesis experiments, indicating this might be a new clue for additional functional study of Kv1 channels.

Alginate is known to prevent elimination of Pseudomonas aeruginosa biofilms. Alginate lyase (AlgL) might therefore facilitate treatment of Pseudomonas aeruginosa-infected cystic fibrosis patients. However, the catalytic activity of wild-type AlgL is not sufficiently high. Therefore, molecular modeling and site-directed mutagenesis of AlgL might assist in enzyme engineering for therapeutic development. AlgL, isolated from Azotobacter vinelandii, catalyzes depolymerization of alginate via a β-elimination reaction. AlgL was modeled based on the crystal structure template of Sphingomonas AlgL species A1-III. Based on this computational analysis, AlgL was subjected to site-directed mutagenesis to improve its catalytic activity. The kcat/Km of the K194E mutant showed a nearly 5-fold increase against the acetylated alginate substrate, as compared to the wild-type. Double and triple mutants (K194E/K245D, K245D/K319A, K194E/K245D/E312D, and K194E/K245D/K319A) were also prepared. The most potent mutant was observed to be K194E/K245D/K319A, which has a 10-fold improved kcat value (against acetylated alginate) compared to the wild-type enzyme. The antibiofilm effect of both AlgL forms was identified in combination with piperacillin/tazobactam (PT) and the disruption effect was significantly higher in mutant AlgL combined with PT than wild-type AlgL. However, for both the wild-type and K194E/K245D/K319A mutant, the use of the AlgL enzyme alone did not show significant antibiofilm effect.

Cocaine binds and inhibits dopamine transporter (DAT), norepinephrine transporter (NET) and serotonin transporter. The residues forming cocaine binding sites are unknown. RTI-113, a cocaine analog, is 100× more potent at inhibiting DAT than inhibiting NET. Here we show that removing the hydroxyl group from residue Tyr151 in NET by replacing it with Phe, the corresponding residue in DAT, increased the sensitivity of NET to RTI-113, while the reverse mutation in DAT decreased the sensitivity of DAT to RTI-113. In contrast, RTI-31, another cocaine analog having the same structure as RTI-113 but with the phenyl group at the 2β position replaced by a methyl group, inhibits the transporter mutants equally well whether a hydroxyl group is present at the residue or not. The data suggest that this residue contributes to cocaine binding site and is close to the 2β position of cocaine analogs. These results are consistent with our previously proposed cocaine-DAT binding model where cocaine initially binds to a site that does not overlap with, but is close to, the dopamine-binding site. Computational modeling and molecular docking yielded a binding model that explains the observed changes in RTI-113 inhibition potencies.

Population migrations in Southwest and South China have played an important role in the formation of East Asian populations and led to a high degree of cultural diversity among ethnic minorities living in these areas. To explore the genetic relationships of these ethnic minorities, we systematically surveyed the variation of 10 autosomal STR markers of 1,538 individuals from 30 populations of 25 ethnic minorities, of which the majority were chosen from Southwest China, especially Yunnan Province. With genotyped data of the markers, we constructed phylogenies of these populations with both D(A) and D(C) measures and performed a principal component analysis, as well as a clustering analysis by structure. Results showed that we successfully recovered the genetic structure of analyzed populations formed by historical migrations. Aggregation patterns of these populations accord well with their linguistic affiliations, suggesting that deciphering of genetic relationships does in fact offer clues for study of ethnic differentiation.

Transcription factor FOXM1 is involved in stimulating cell proliferation, enhancing DNA damage repair, promoting metastasis of cancer cells, and the inhibition of FOXM1 has been shown to prevent the initiation and progression of multiple cancers and FOXM1 is considered to be an effective target for tumor therapeutic drug development. The N-terminus of FOXM1 has been found to prevent transcriptional activities of FOXM1 and to mediate the interaction between FOXM1 and SMAD3.

Many essential oils (EOs) regulate the quorum-sensing (QS) system of pathogens and inhibit the virulence expression. Interference with QS can potentially reduce bacterial multidrug resistance and aid the biological control of bacterial disease. In the present work, the antibacterial and anti-QS activities of Cinnamomum camphora leaf EO were investigated. A total of 23 chemical components with relative levels ≥0.11%, including a large number of terpene compounds, were identified in C. camphora leaf EO by gas chromatography-mass spectrometry (GC-MS). The principal component was linalool, followed by eucalyptol, with relative levels of 51.57% and 22.07%, respectively. The minimum inhibitory concentration (MIC) and antibacterial activity of C. camphora EO were examined, and P. aeruginosa and E. coli ATCC25922 showed the highest and lowest sensitivity to C. camphora EO, respectively. Tests of QS inhibitory activity revealed that C. camphora EO significantly decreased the production of violacein and biofilm biomass in C. violaceum, with the maximum inhibition rates of 63% and 77.64%, respectively, and inhibited the biofilm formation and swarming movement, independent of affecting the growth of C. violaceum. Addition of C. camphora EO also resulted in downregulation of the expression of the acyl-homoserine lactones (AHL) synthesis gene (cviI) and transcription regulator (cviR), and inhibited the expression of QS-regulated virulence genes, including vioA, vioB, vioC, vioD, vioE, lasA, lasB, pilE3, and hmsHNFR. Collectively, the prominent antibacterial activity and anti-QS activities clearly support that C. camphora EO acts as a potential antibacterial agent and QS inhibitor in the prevention of bacterial contamination.

Acute lung injury (ALI) is characterized by protein-rich pulmonary edema, critical hypoxemia, and influx of pro-inflammatory cytokines and cells. There are currently no effective pharmacon therapies in clinical practice. Syndecan-1 in endothelial cells has potential to protect barrier function of endothelium and suppress inflammation response. Thus, the present study was to identify whether exosomes with encapsulation of syndecan-1 could achieve ideal therapeutic effects in ALI. Exosomes were isolated from the conditional medium of lentivirus-transfected mouse pulmonary microvascular endothelial cells (MPMVECs) and characterized by nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM), and western blotting. ALI mouse models were induced via intratracheal administration of lipopolysaccharide (LPS) and treated with exosomes. Lung edema, inflammation, and glycocalyx thickness were examined. The possible mechanism was verified by immunoblotting in MPMVECs. The purified exosomes included SDC1-high-Exos and SDC1-low-Exos which loaded with up-regulated syndecan-1 and down-regulated syndecan-1 respectively. Compared with SDC1-low-Exos, administration of SDC1-high-Exos could ameliorate lung edema and inflammation, attenuate number of cells and protein levels in bronchoalveolar lavage fluid (BALF), and preserve glycocalyx. Furthermore, SDC1-high-Exos also mitigated the expression of pro-inflammatory cytokines such as IL-1β, TNF-α, and IL-6 following LPS challenge. In MPMVECs, SDC1-high-Exos decreased stress fiber formation and ameliorated monolayer hyper-permeability after LPS stimulation. Western blotting analysis demonstrated that FAK/p190RhoGAP/RhoA/ROCK/NF-κB signaling pathway may be involved in LPS-induced ALI. In conclusion, SDC1-high-Exos play a pivotal role in ameliorating LPS-stimulated ALI models and may be served as a potential therapeutic agent for clinical application in the future.

Hand, foot, and mouth disease (HFMD) is a common childhood disease, which is usually caused by enterovirus A (EV-A) serotypes. Enterovirus A71 (EV-A71) and coxsackievirus A16 (CV-A16) are the main etiologic agents. Multiple serotypes of enterovirus B serotypes (EV-B) have been detected in outbreaks or sporadic cases of HFMD.

In this study, we investigated the spontaneous neural plasticity on the contralateral side in hypertensive rats, including the expression of nerve growth factors (synaptophysin [SYN] and growth-associated protein 43 [GAP-43]), and the association between nerve fiber sprouting and redistribution, and the recovery of motor functions following sensorimotor cortical infarction.

The bacterial endotoxin or lipopolysaccharide (LPS) leads to the extensive vascular endothelial cells (EC) injury under septic conditions. Guanine nucleotide exchange factor-H1 (GEF-H1)/ROCK signaling not only involved in LPS-induced overexpression of pro-inflammatory mediator in ECs but also implicated in LPS-induced endothelial hyper-permeability. However, the mechanisms behind LPS-induced GEF-H1/ROCK signaling activation in the progress of EC injury remain incompletely understood. GEF-H1 localized on microtubules (MT) and is suppressed in its MT-bound state. MT disassembly promotes GEF-H1 release from MT and stimulates downstream ROCK-specific GEF activity. Since glycogen synthase kinase (GSK-3beta) participates in regulating MT dynamics under pathologic conditions, we examined the pivotal roles for GSK-3beta in modulating LPS-induced activation of GEF-H1/ROCK, increase of vascular endothelial permeability and severity of acute lung injury (ALI). In this study, we found that LPS induced human pulmonary endothelial cell (HPMEC) monolayers disruption accompanied by increase in GSK-3beta activity, activation of GEF-H1/ROCK signaling and decrease in beta-catenin and ZO-1 expression. Inhibition of GSK-3beta reduced HPMEC monolayers hyper-permeability and GEF-H1/ROCK activity in response to LPS. GSK-3beta/GEF-H1/ROCK signaling is implicated in regulating the expression of beta-catenin and ZO-1. In vivo, GSK-3beta inhibition attenuated LPS-induced activation of GEF-H1/ROCK pathway, lung edema and subsequent ALI. These findings present a new mechanism of GSK-3beta-dependent exacerbation of lung micro-vascular hyper-permeability and escalation of ALI via activation of GEF-H1/ROCK signaling and disruption of intracellular junctional proteins under septic condition.

Human echovirus 12 (E-12) belongs to the enterovirus B species. To date, only one full-length genome sequence of E-12 (prototype strain Travis) is available in the GenBank database. This study determined the complete sequence of three E-12 strains, which were isolated from the stools of three healthy children in Yunnan, China, in 2013. We revealed that the three Yunnan E-12 strains had only 80.8-80.9% nucleotide identity and 96.4-96.8% amino acid identity with the Travis strain based on pairwise comparisons of the complete genome nucleotide and amino acid sequences. The three Yunnan strains shared 99.7% nucleotide identity and 99.1-99.5% amino acid similarity. Phylogenetic and similarity plot analyses showed that intertypic recombination occurred in the non-structural regions of the three Yunnan E-12 strains. This is the first report of the complete genome sequence of E-12 in China and it enriches the complete genome sequences of E-12 in the GenBank database.

Disrupting protein-protein interactions (PPIs) has emerged as a promising strategy for cancer drug development. Interfering peptides disrupting PPIs can be rationally designed based on the structures of natural sequences mediating these interactions. Transcription factor FOXM1 overexpresses in multiple cancers and is considered an effective target for cancer therapeutic drug development. Using a rational design approach, we have generated a peptide library from the FOXM1 C-terminal sequence and screened FOXM1-binding peptides. Combining FOXM1 binding and cell inhibitory results, we have obtained a FOXM1-targeting interfering peptide M1-20 that is optimized from the natural parent peptide to the D-retro-inverso peptide. With improved stability characteristics, M1-20 inhibits proliferation and migration, and induces apoptosis of cancer cells. Mechanistically, M1-20 inhibits FOXM1 transcriptional activities by disrupting its interaction between the MuvB complex and the transcriptional co-activator CBP. These are consistent with the results that M1-20 suppresses cancer progression and metastasis without noticeable toxic and side effects in wild-type mice. These findings reveal that M1-20 has the potential to be developed as an anti-cancer drug candidate targeting FOXM1.

Insights into the interacting mode of CXCR4 with SDF-1alpha are crucial in understanding the structural and functional characteristics of CXCR4 receptor. In this paper a computational pipeline, integrating protein structure prediction, molecular dynamics simulations, automated molecular docking, and Brownian dynamics simulations were employed to investigate the dynamic and energetic aspects of CXCR4 associating with SDF-1alpha. The entire simulation revealed the surface distribution feature of electrostatic potentials and conformational "open-close" process of the receptor. The possible binding conformation of CXCR4 was identified, and the CXCR4-SDF-1alpha binding complex was generated. Arg188-Glu277 salt bridge plays an important role for both the extracellular domain conformational change and SDF-1alpha binding. Two binding sites were mapped at the extracellular domain (Site 1) and inside the transmembrane domain (Site 2), which are composed of conserved residues. Sites 1 and 2 contribute approximately 60% and 40% to the binding affinity with SDF-1alpha, respectively. The binding model is in agreement with most of the experimental data. Transmembrane VI has more significant motion in the harmonious conformational transition of CXCR4 during SDF-1alpha binding, which may be possibly associated with signal transduction. Based on the modeling and simulation, a binding mechanism hypothesis between CXCR4 and SDF-1alpha and its relationship to the signal transduction has been proposed.

Pathogen-driven balancing selection determines the richness of human leukocyte antigen (HLA) alleles. Changes in the pathogen spectrum may cause corresponding changes in HLA loci. Approximately 700 years ago, a Mongolian population moved from the north of China to the Yunnan region in the south of China. The pathogen spectrum in the south of China differs from that in the north. In this study, changes in the HLA genes in the Yunnan Mongolian population, as well as the underlying mechanism, were investigated. A sequence-based typing method (SBT) was used to genotype HLA-DRB1 in 470 individuals from two Mongolian populations and another five ethnic groups. Meanwhile, 10 autosomal short tandem repeats (STRs) were genotyped to assess the influence of genetic background on HLA-DRB1 frequencies. The frequencies of certain alleles changed significantly in the Mongolian population that migrated to Yunnan. For example, DRB1*12:02:01 increased from 6.1% to 35.4%. STR analysis excluded the possibility of a recent bottleneck and indicated that 50% of the genetic consistency between northern and southern Mongolians; Tajima's D value for HLA-DRB1 exon2 and dN/dS analysis showed that the HLA-DRB1 genes in both Mongolian populations were under balancing selection. However, the sites under natural selection changed. We proposed that the dramatically change of HLA frequencies in southern Mongolian was caused by a combination of inter-population gene flow and natural selection. Certain diseases specific to the south of China, such as malaria, may be the driving force behind the enhanced DRB1*12:02:01 frequency.

The mechanism through which death-associated protein kinase 1 (DAPK1) causes hepatocellular carcinoma (HCC) progression remains unclear. In this study, we aimed to identify key proteins that were altered after DAPK1 knockout.

To investigate the feasibility and safety of remote ischemic postconditioning (RIPC) in acute ischemic stroke patients after intravenous recombinant tissue plasminogen activator (rt-PA) thrombolysis (IVT).