Gene set correlation enrichment analysis for interpreting and annotating gene expression profiles.

Pathway analysis, including nontopology-based (non-TB) and topology-based (TB) methods, is widely used to interpret the biological phenomena underlying differences in expression data between two phenotypes. By considering dependencies and interactions between genes, TB methods usually perform better than non-TB methods in identifying pathways that include closely relevant or directly causative genes for a given phenotype. However, most TB methods may be limited by incomplete pathway data used as the reference network or by difficulties in selecting appropriate reference networks for different research topics. Here, we propose a gene set correlation enrichment analysis method, Gscore, based on an expression dataset-derived coexpression network to examine whether a differentially expressed gene (DEG) list (or each of its DEGs) is associated with a known gene set. Gscore is better able to identify target pathways in 89 human disease expression datasets than eight other state-of-the-art methods and offers insight into how disease-wide and pathway-wide associations reflect clinical outcomes. When applied to RNA-seq data from COVID-19-related cells and patient samples, Gscore provided a means for studying how DEGs are implicated in COVID-19-related pathways. In summary, Gscore offers a powerful analytical approach for annotating individual DEGs, DEG lists, and genome-wide expression profiles based on existing biological knowledge.

Cysteine Regulates Oxidative Stress and Glutathione-Related Antioxidative Capacity before and after Colorectal Tumor Resection.

Cysteine might scavenge free radicals and is a limiting substrate for the cellular synthesis of glutathione (GSH). We investigated the association of cysteine with oxidative stress and GSH-related antioxidant capacity in colorectal cancer (CRC) patients. Plasma samples were drawn from 66 patients 1 day before (pre-resection) and 4 weeks after resection (post-resection). Tumor and adjacent normal tissues were collected. We measured levels of plasma and tissue cysteine, homocysteine, oxidative stress indicators (malondialdehyde, MDA; advanced oxidation protein products, AOPP), GSH, and antioxidant enzyme activities. After tumor resection, patients had significantly higher levels of plasma cysteine, homocysteine, MDA, AOPP, and GSH-related antioxidant enzyme activities when compared with pre-resection. Levels of cysteine, homocysteine, AOPP and all antioxidant capacity indicators in tumor tissue were significantly higher than those levels in the adjacent normal tissue. Plasma cysteine levels measured at pre-resection were positively associated with MDA levels in the tumor and in the adjacent normal tissues. Cysteine levels in tumor and adjacent normal tissues were significantly associated with tissue levels of homocysteine, almost as indicators of oxidative stress and antioxidant capacities. Cysteine in the circulation was likely utilized to mediate GSH-related antioxidant capacity and further cope with increased oxidative stress in tumor and adjacent normal tissues.

Design, synthesis and molecular docking study of α-triazolylsialosides as non-hydrolyzable and potent CD22 ligands.

Ligand 1 was the first reported example of monomeric high-affinity synthetic CD22 ligand that regulated B cell activation in vitro, augmented antibody production and regulated immune responses in mice. Replacing O-glycoside linkage of 1 by nitrogen of triazole by click reaction afforded compounds which are as potent as the parent compound. The synthesis of the new compounds is straightforward with fewer synthetic steps and higher yield. Such a strategy provided stable ligand that can bind avidly and can be conjugated to drugs for B-cell targeting or multimeric formation. The new compounds were screened for their affinity to CD22, using surface plasmon resonance (SPR). Compound 12 was obtained as a bioisosteric analogue and an anomerically stable imitation of 1. It was, also, screened for MAG to test for selectivity and analyzed by molecular docking and dynamic simulation to explore the potential binding modes and source of selectivity within CD22. Our results could enable the development of small molecule drug capable of modulating the activity of CD22 in autoimmune diseases and malignancies derived from B-cells.

Identification of the PCA29 gene signature as a predictor in prostate cancer.

Prostate cancer (PCa) is the second leading cause of cancer death among men worldwide. About 70% of PCa patients were diagnosed at later stage, and metastasis has been observed. Additionally, the cure rate of PCa closely relies on the early diagnosis with biomarkers. The identification of biomarkers for diagnosis and prognosis is an urgent clinical issue for PCa. Here, we developed a novel scoring strategy, including cluster score (CS) and predicting score (PS), to identify 29 PCa genes (called PCa29) for early diagnostic biomarkers from two datasets in Gene Expression Omnibus. The result indicates that PCa29 can discriminate between normal and tumor tissues and are specific for prostate cancer. To validate PCa29, we found that 97% of PCa29 were consistently significant with these gene expressions in The Cancer Genome Atlas; furthermore, ∼ 70% of PCa29 are consensus to the protein expression in The Human Protein Atlas. Finally, we examined 10 genes in PCa29 on three PCa cell lines by real-time quantitative polymerase chain reaction. The experimental results show that the trend of the differential PCa29 expression is consistent with the analyzed results from our novel scoring method. We believe that our method is useful and PCa29 are potential biomarkers that provide the clues to develop targeting therapy for PCa.

A homologous mapping method for three-dimensional reconstruction of protein networks reveals disease-associated mutations.

BACKGROUND: One of the crucial steps toward understanding the associations among molecular interactions, pathways, and diseases in a cell is to investigate detailed atomic protein-protein interactions (PPIs) in the structural interactome. Despite the availability of large-scale methods for analyzing PPI networks, these methods often focused on PPI networks using genome-scale data and/or known experimental PPIs. However, these methods are unable to provide structurally resolved interaction residues and their conservations in PPI networks. RESULTS: Here, we reconstructed a human three-dimensional (3D) structural PPI network (hDiSNet) with the detailed atomic binding models and disease-associated mutations by enhancing our PPI families and 3D-domain interologs from 60,618 structural complexes and complete genome database with 6,352,363 protein sequences across 2274 species. hDiSNet is a scale-free network (γ = 2.05), which consists of 5177 proteins and 19,239 PPIs with 5843 mutations. These 19,239 structurally resolved PPIs not only expanded the number of PPIs compared to present structural PPI network, but also achieved higher agreement with gene ontology similarities and higher co-expression correlation than the ones of 181,868 experimental PPIs recorded in public databases. Among 5843 mutations, 1653 and 790 mutations involved in interacting domains and contacting residues, respectively, are highly related to diseases. Our hDiSNet can provide detailed atomic interactions of human disease and their associated proteins with mutations. Our results show that the disease-related mutations are often located at the contacting residues forming the hydrogen bonds or conserved in the PPI family. In addition, hDiSNet provides the insights of the FGFR (EGFR)-MAPK pathway for interpreting the mechanisms of breast cancer and ErbB signaling pathway in brain cancer. CONCLUSIONS: Our results demonstrate that hDiSNet can explore structural-based interactions insights for understanding the mechanisms of disease-associated proteins and their mutations. We believe that our method is useful to reconstruct structurally resolved PPI networks for interpreting structural genomics and disease associations.

Dual thio-digalactoside-binding modes of human galectins as the structural basis for the design of potent and selective inhibitors.

Human galectins are promising targets for cancer immunotherapeutic and fibrotic disease-related drugs. We report herein the binding interactions of three thio-digalactosides (TDGs) including TDG itself, TD139 (3,3'-deoxy-3,3'-bis-(4-[m-fluorophenyl]-1H-1,2,3-triazol-1-yl)-thio-digalactoside, recently approved for the treatment of idiopathic pulmonary fibrosis), and TAZTDG (3-deoxy-3-(4-[m-fluorophenyl]-1H-1,2,3-triazol-1-yl)-thio-digalactoside) with human galectins-1, -3 and -7 as assessed by X-ray crystallography, isothermal titration calorimetry and NMR spectroscopy. Five binding subsites (A-E) make up the carbohydrate-recognition domains of these galectins. We identified novel interactions between an arginine within subsite E of the galectins and an arene group in the ligands. In addition to the interactions contributed by the galactosyl sugar residues bound at subsites C and D, the fluorophenyl group of TAZTDG preferentially bound to subsite B in galectin-3, whereas the same group favored binding at subsite E in galectins-1 and -7. The characterised dual binding modes demonstrate how binding potency, reported as decreased Kd values of the TDG inhibitors from µM to nM, is improved and also offer insights to development of selective inhibitors for individual galectins.

Comprehensive modelling of the Neurospora circadian clock and its temperature compensation.

Circadian clocks provide an internal measure of external time allowing organisms to anticipate and exploit predictable daily changes in the environment. Rhythms driven by circadian clocks have a temperature compensated periodicity of approximately 24 hours that persists in constant conditions and can be reset by environmental time cues. Computational modelling has aided our understanding of the molecular mechanisms of circadian clocks, nevertheless it remains a major challenge to integrate the large number of clock components and their interactions into a single, comprehensive model that is able to account for the full breadth of clock phenotypes. Here we present a comprehensive dynamic model of the Neurospora crassa circadian clock that incorporates its key components and their transcriptional and post-transcriptional regulation. The model accounts for a wide range of clock characteristics including: a periodicity of 21.6 hours, persistent oscillation in constant conditions, arrhythmicity in constant light, resetting by brief light pulses, and entrainment to full photoperiods. Crucial components influencing the period and amplitude of oscillations were identified by control analysis. Furthermore, simulations enabled us to propose a mechanism for temperature compensation, which is achieved by simultaneously increasing the translation of frq RNA and decreasing the nuclear import of FRQ protein.

Yeast-expressed classical swine fever virus glycoprotein E2 induces a protective immune response.

Classical swine fever (CSF) is an economically important swine disease worldwide. The glycoprotein E2 of classical swine fever virus (CSFV) is a viral antigen that can induce a protective immune response against CSF. A recombinant E2 protein was constructed using the yeast Pichia pastoris expression system and evaluated for its vaccine efficacy. The yeast-expressed E2 (yE2) was shown to have N-linked glycosylation and to form homodimer molecules. Four 6-week-old specified-pathogen-free (SPF) piglets were intramuscularly immunized with yE2 twice at 3-week intervals. All yE2-vaccinated pigs could mount an anamnestic response after booster vaccination with neutralizing antibody titers ranging from 1:96 to 1:768. Neutralizing antibody titers at 10 weeks post booster vaccination ranged from 1:16 to 1:64. At this time, the pigs were subjected to challenge infection with a dose of 1x10(5)TCID(50) (50% tissue culture infective dose) virulent CSFV strain. At 1 week post challenge infection, all of the yE2-immunized pigs were alive and without symptoms or signs of CSF. Neutralizing antibody titers at this time ranged from 1:4,800 to 1:12,800 and even to 1:51,200 one week later. In contrast, the control pigs continuously exhibited signs of CSF and had to be euthanized because of severe clinical symptoms at 6 days post challenge infection. All of the yE2-vaccinated pigs were E(rns) antibody negative and had seroconverted against E(rns) by post challenge day 11, suggesting that yE2 is a potential DIVA (differentiating infected from vaccinated animals) vaccine. The yeast-expressed E2 protein retains correct immunogenicity and is able to induce a protective immune response against CSFV infection.

Expression of the porcine circovirus type 2 capsid protein subunits and application to an indirect ELISA.

Porcine circovirus type 2 (PCV2) is considered to be associated with post-weaning multisystemic wasting syndrome (PMWS), which is a newly emerged economically important swine disease. The entire coding region of open reading frame 2 (ORF2), encoding the viral capsid protein (Cap), of PCV2 was cloned and sequenced from the clinical specimen obtained from PMWS-affected piglets. Six recombinant subunits, A-F, spanning the defined regions of Cap were produced by E. coli expression system and used as antigens for testing their reactivities with swine sera in the indirect ELISA. The recombinant Cap subunit-based ELISA was evaluated by examining a panel of 12 PCV2-negative and 26 PCV2-positive sera. When the positive/negative cut-off value was set at the mean value of negative sera plus 3 standard deviations, all subunits-based ELISA demonstrated 100% specificities. The N-terminal subunits, A and B, revealed poor reactivity with positive swine sera, whereas, greater immunoreactivity was observed for the C-terminal subunits of which subunits C and D demonstrated good sensitivities of 96.2% and 84.6%, respectively. The recombinant Cap subunits possessing defined antigenicity are easy to produce and the subunit-based ELISA was developed with a high specificity and sensitivity that may provide a useful method for routine serodiagnosis of PCV2 infection.