Ubiquitin-like prokaryotic MoaD as a fusion tag for expression of heterologous proteins in Escherichia coli

BACKGROUND: Eukaryotic ubiquitin and SUMO are frequently used as tags to enhance the fusion protein expression in microbial host. They increase the solubility and stability, and protect the peptides from proteolytic degradation due to their stable and highly conserved structures. Few of prokaryotic ubiquitin-like proteins was used as fusion tags except ThiS, which enhances the fusion expression, however, reduces the solubility and stability of the expressed peptides in E. coli. Hence, we investigated if MoaD, a conserved small sulfur carrier in prokaryotes with the similar structure of ubiquitin, could also be used as fusion tag in heterologous expression in E. coli. RESULTS: Fusion of MoaD to either end of EGFP enhanced the expression yield of EGFP with a similar efficacy of ThiS. However, the major parts of the fusion proteins were expressed in the aggregated form, which was associated with the retarded folding of EGFP, similar to ThiS fusions. Fusion of MoaD to insulin chain A or B did not boost their expression as efficiently as ThiS tag did, probably due to a less efficient aggregation of products. Interestingly, fusion of MoaD to the murine ribonuclease inhibitor enhanced protein expression by completely protecting the protein from intracellular degradation in contrast to ThiS fusion, which enhanced degradation of this unstable protein when expressed in E. coli. CONCLUSIONS: Prokaryotic ubiquitin-like protein MoaD can act as a fusion tag to promote the fusion expression with varying mechanisms, which enriches the arsenal of fusion tags in the category of insoluble expression

Orbital-angular-momentum fluorescence emission based on photon–electron interaction in a vortex field of an active optical fiber

We develop a model of interaction between photons and electrons in an active vortex field, which can generate a fluorescence spectrum with the characteristics of orbital angular momentum (OAM). In an active optical fiber, our findings generalize the notion of photon–electron interaction and point to a new kind of OAM-mode broad-spectrum light source, which could be interpreted in two processes: one microscopically is the excitation of OAM-carrying photons based on the photon–electron interaction; the other macroscopically is the emission and transmission of a donut-shaped fluorescence in a vortex field with a spiral phase wavefront in a ring-core active fiber. Here we present a straightforward experimental method that the emission of broad-spectrum fluorescence with an OAM feature is actualized and validated in a ring-core erbium-doped fiber. The spectrum has a broad spectral width up to 50 nm. Furthermore, four wavelengths are extracted from the fluorescence spectrum and superimposed with their corresponding Gaussian beams, from which the spiral-shaped interferograms of OAM modes in a broad spectrum are identified with high purity. The application of the OAM-based fluorescence light source may range from classical to quantum information technologies, and enable high-capacity communication, high-sensitivity sensing, high-resolution fluorescence imaging, etc

FGL2 deficiency alleviates maternal inflammation-induced blood-brain barrier damage by blocking PI3K/NF-κB mediated endothelial oxidative stress

IntroductionThe impairment of blood-brain barrier (BBB) is one of the key contributors to maternal inflammation induced brain damage in offspring. Our previous studies showed Fibrinogen-like protein 2 (FGL2) deficiency alleviated maternal inflammation induced perinatal brain damage. However, its role in BBB remains undefined.MethodsLipopolysaccharide (LPS) was intraperitoneally injected to dams at Embryonic day 17 to establish maternal inflammation model. FGL2 knockout mice and primary brain microvascular endothelial cells (BMECs) were used for the in-vivo and in-vitro experiments. BBB integrity was assessed by sodium fluorescein extravasation and tight junction (TJ) protein expression. Oxidative stress and the activation of PI3K/NF-κB pathway were evaluated to explore the mechanisms underlying.ResultsUpon maternal inflammation, BBB integrity was remarkedly reduced in neonatal mice. Meanwhile, FGL2 expression was consistently increased in BBB-impaired brain as well as in LPS-treated BMECs. Moreover, FGL2 deficiency attenuated the hyperpermeability of BBB, prevented the decline of TJ proteins, and reduced the cytokine expressions in LPS-exposed pups. Mechanistically, the indicators of oxidative stress, as well as the activation of PI3K/NF-κB pathway, were upregulated after LPS exposure in vivo and in vitro. FGL2 deletion decreased the generation of ROS and NO, reduced the endothelial iNOS and NOX2 expressions, and suppressed the PI3K/NF-κB pathway activation. Besides, inhibition of PI3K by LY294002 decreased the oxidative stress in LPS-treated wild-type BMECs. While, overexpression of PI3K by lentivirus reemerged the induction of NOX2 and iNOS as well as NF-κB activation in FGL2-deleted BMECs.ConclusionOur findings indicate that FGL2 deficiency alleviates the maternal inflammation-induced BBB disruption by inhibiting PI3K/NF-κB mediated oxidative stress in BMECs. Targeting FGL2 may provide a new therapy for prenatal brain damage of offspring

Egg Protein Transferrin-Derived Peptides IRW and IQW Regulate Citrobacter rodentium-Induced, Inflammation-Related Microbial and Metabolomic Profiles

Bioactive peptides that target the gastrointestinal tract can strongly affect the health of animals and humans. This study aimed to evaluate the abilities of two peptides derived from egg albumin transferrin, IRW and IQW, to treat enteritis in a mouse model of Citrobacter rodentium-induced colitis by evaluating serum metabolomics and gut microbes. Forty-eight mice were randomly assigned to six groups: basal diet (CTRL), intragastric administration Citrobacter rodentium (CR), basal diet with 0.03%IRW (IRW), CR with 0.03% IRW (IRW+CR), basal diet with 0.03%IQW (IQW) and CR with 0.03% IQW (IQW+CR). CR administration began on day 10 and continued for 7 days. After 14 days of IRW and IQW treatment, serum was collected and subjected to a metabolomics analysis. The length and weight of each colon were measured, and the colon contents were collected for 16srRNA sequencing. The colons were significantly longer in the CR group, compared to the CTRL group. A serum metabolomics analysis revealed no significant difference in microbial diversity between the six groups. Compared with the CTRL group, the proportions of Firmicutes and Actinobacteria species decreased significantly and the proportions of Bacteroidetes and Proteobacteria species increased in the CR group. There were no significant differences between the CTRL and other groups. The serum metabolomics analysis revealed that Infected by CR increased the levels of oxalic acid, homogentisic acid and prostaglandin but decreased the levels of L-glutamine, L-acetyl carnitine, 1-methylhistidine and gentisic acid. Therefore, treatment with IRW and IQW was shown to regulate the intestinal microorganisms associated with colonic inflammation and serum metabolite levels, thus improving intestinal health

Chloroplast genomes in seven Lagerstroemia species provide new insights into molecular evolution of photosynthesis genes

Lagerstroemia indica is an important commercial tree known for the ornamental value. In this study, the complete chloroplast genome sequence of Lagerstroemia indica “Pink Velour” (Lagerstroemia “Pink Velour”) was 152,174 bp in length with a GC content of 39.50%. It contained 85 protein coding genes (PCGs), 37 tRNAs, and 8 rRNA genes. 207 simple sequence repeats (SSRs) and 31 codons with relative synonymous codon (RSCU)value > 1 were detected. Phylogenetic analysis divided 10 Lagerstroemia species into evolutionary branches of clade A and clade B. We conducted a comparative analysis of Lagerstroemia “Pink Velours” complete chloroplast genome with the genomes of six closely related Lagerstroemia species from different origins. The structural features of all seven species were similar, except for the deletion of ycf1 nucleobases at the JSA boundary. The large single-copy (LSC) and the small single-copy (SSC) had a higher sequence divergence than the IR region, and 8 genes that were highly divergent (trnK-UUU, petN, psbF, psbJ, ndhE, ndhD, ndhI, ycf1) had been identified and could be used as molecular markers in future studies. High nucleotide diversity was present in genes belonging to the photosynthesis category. Mutation of single nucleic acid was mainly influenced by codon usage. The value percentage of nonsynonymous substitutions (Ka) and synonymous substitutions (Ks) in 6 Lagerstroemia species revealed that more photosynthesis genes have Ka or Ks only in Lagerstroemia fauriei, Lagerstroemia limii, and Lagerstroemia subcostata. These advances will facilitate the breeding of closely related Lagerstroemia species and deepen understanding on climatic adaptation of Lagerstroemia plants

The effects and mechanisms of SLC34A2 in tumorigenesis and progression of human non-small cell lung cancer

BACKGROUND: SLC34A2 with highest expressions in lung, small intestine and kidney encoded a type 2b sodium-dependent phosphate transporter (NaPi-IIb). In lung, SLC34A2 only expressed in the apical membrane of type II alveolar epithelium cells (ATII cells) and played a pivotal role during the fetal lung development and embryonic development. ATII cells acting as multifunctional stem cells might transform into NSCLC after undergoing exogenous or endogenous factors. Increasing evidences showed that the genes performing critical roles during embryogenesis were also expressed during the development of cancer. In addition, recent research found the expression of SLC34A2 had a significant difference between the surgical samples of NSCLC and normal tissues, and SLC34A2 was down-regulated in lung adenocarcinoma cell line A549 and up-regulation expression of SLC34A2 could significantly inhibit cell viability and invasion of A549 in vitro. These results suggested SLC34A2 might play an important role in the development of NSCLC. However, the role of SLC34A2 in tumorigenesis and progression of NSCLC remains unknown. RESULTS: Our study found that SLC34A2 was also significantly down-regulated in 14/15 of examined NSCLC tissues. Moreover, we found that expressions of SLC34A2 were reduced in six NSCLC cell lines for the first time. Our result also revealed a dramatic inhibitory effects of SLC34A2 on cell growth, migration and invasion of several NSCLC cell lines. SLC34A2 also strongly inhibited tumor growth and metastasis ability in A549 subcutaneous tumor model and lung metastasis model, respectively. Further studies found that the suppressive effects of SLC34A2 on tumorigenesis and progression might be associated with the down-regulation of related protein in PI3K/Akt and Ras/Raf/MEK signal pathway. CONCLUSIONS: For the first time, our data indicated that SLC34A2 could exert significantly suppressive effects on tumorigenesis and progression of NSCLC. SLC34A2 might provide new insights for further understanding the early pathogenesis of human NSCLC

Changes in the mental health status of the general Chinese population during the COVID-19 pandemic: A longitudinal study

The study is based on a longitudinal evaluation of the public, during the initial COVID-19 outbreak in China and 8 months after. It aimed to explore the changes in the mental health of the public at the beginning of the pandemic and during the regular epidemic prevention and control. An online survey questionnaire was used to collect data during the initial COVID-19 outbreak (February 10, 2020–February 18, 2020; T1) and 8 months after the outbreak (October 21, 2020–December 29, 2020; T2). Psychological distress was assessed using the Patient Health Questionnaire-9 (PHQ-9), Self-rating Anxiety Scale (SAS), and Post-traumatic Stress Disorder Checklist (PCL-5). A chi-square test was used to compare the changes in the depression and anxiety scores at T1 and T2, and the correlation between symptoms was analyzed through Spearman's rank correlation. In T1, 1,200 people were recruited, while 168 people responded in T2. Depression (48.2–31.0%; p=0.001) and anxiety (17.9–9.5%; p = 0.026) symptoms decreased over time; two participants developed post-traumatic stress disorder (PTSD) in T2. The scores of the PHQ-9 scale and the SAS scale were both positively correlated with the score of the PCL-5 scale and negatively correlated with sleep time. During the COVID-19 pandemic, part of the general population's anxiety and depression significantly reduced with time, and they rarely developed PTSD. PTSD occurrence was related to severe depression and anxiety

Generation and Comprehensive Analysis of Host Cell Interactome of the PA Protein of the Highly Pathogenic H5N1 Avian Influenza Virus in Mammalian Cells

Accumulating data have identified the important roles of PA protein in replication and pathogenicity of influenza A virus (IAV). Identification of host factors that interact with the PA protein may accelerate our understanding of IAV pathogenesis. In this study, using immunoprecipitation assay combined with liquid chromatography-tandem mass spectrometry, we identified 278 human cellular proteins that might interact with PA of H5N1 IAV. Gene Ontology annotation revealed that the identified proteins are highly associated with viral translation and replication. Further KEGG pathway analysis of the interactome profile highlighted cellular pathways associated with translation, infectious disease, and signal transduction. In addition, Diseases and Functions analysis suggested that these cellular proteins are highly related with Organismal Injury and Abnormalities and Cell Death and Survival. Moreover, two cellular proteins (nucleolin and eukaryotic translation elongation factor 1-alpha 1) identified both in this study and others were further validated to interact with PA using co-immunoprecipitation and co-localization assays. Therefore, this study presented the interactome data of H5N1 IAV PA protein in human cells which may provide novel cellular target proteins for elucidating the potential molecular functions of PA in regulating the lifecycle of IAV in human cells

CpG_MI: a novel approach for identifying functional CpG islands in mammalian genomes

CpG islands (CGIs) are CpG-rich regions compared to CpG-depleted bulk DNA of mammalian genomes and are generally regarded as the epigenetic regulatory regions in association with unmethylation, promoter activity and histone modifications. Accurate identification of CpG islands with epigenetic regulatory function in bulk genomes is of wide interest. Here, the common features of functional CGIs are identified using an average mutual information method to differentiate functional CGIs from the remaining CGIs. A new approach (CpG mutual information, CpG_MI) was further explored to identify functional CGIs based on the cumulative mutual information of physical distances between two neighboring CpGs. Compared to current approaches, CpG_MI achieved the highest prediction accuracy. This approach also identified new functional CGIs overlapping with gene promoter regions which were missed by other algorithms. Nearly all CGIs identified by CpG_MI overlapped with histone modification marks. CpG_MI could also be used to identify potential functional CGIs in other mammalian genomes, as the CpG dinucleotide contents and cumulative mutual information distributions are almost the same among six mammalian genomes in our analysis. It is a reliable quantitative tool for the identification of functional CGIs from bulk genomes and helps in understanding the relationships between genomic functional elements and epigenomic modifications