Transcriptional Protein Research Articles

AP-1-dependent fibrosis: Exploring its potential role in the pathogenesis of placental transmogrification of the lung (PTL) via tissue-level transcriptome analysis

Placental transmogrification of the lung (PTL) is a rare pulmonary condition characterized by the presence of immature placental villous structures. The etiology and molecular mechanisms underlying this disease remain largely unknown. This functional study aimed to identify the molecular signatures in the pathogenesis of PTL via comprehensive transcriptome analysis. Comparative transcriptomic assessment of PTL tissue and stromal cells showed differential expression of 257 genes in PTL tissue and 189 genes in stromal cells. Notably, several transcription factors and regulators, including FOSB, FOS, JUN, and ATF3, were upregulated in PTL tissue. Additionally, genes associated with the extracellular matrix and connective tissue, such as COL1A1, MMP2, and SPARC, were significantly altered, indicating possible fibrotic changes. Gene set enrichment analysis highlighted the role of vascular development and extracellular matrix organization, and the Activator Protein-1 (AP-1) transcription factor was significantly activated in PTL tissue. Furthermore, the analysis highlighted an overlap of 25 genes between PTL tissue and stromal cells, underscoring the importance of shared molecular pathways in the pathogenesis of PTL. Among the shared genes, JUND, COL4A2, COL6A2, IGFBP5, and IGFBP7 were consistently upregulated, highlighting the possible involvement of AP-1-mediated signaling and fibrotic changes in the pathogenesis of PTL. The present findings pave the way for further research into the molecular mechanisms underlying PTL and offer novel insights for therapeutic interventions. Given the rarity of PTL, these molecular findings represent a significant step forward in our understanding this enigmatic disease.

Photoregulatory protein kinases fine-tune plant photomorphogenesis by directing a bifunctional phospho-code on HY5 in Arabidopsis

Photomorphogenesis is a light-dependent plant growth and development program. As the core regulator of photomorphogenesis, ELONGATED HYPOCOTYL 5 (HY5) is affected by dynamic changes in its transcriptional activity and protein stability; however, little is known about the mediators of these processes. Here, we identified PHOTOREGULATORY PROTEIN KINASE 1 (PPK1), which interacts with and phosphorylates HY5 in Arabidopsis, as one such mediator. The phosphorylation of HY5 by PPK1 is essential to establish high-affinity binding with B-BOX PROTEIN 24 (BBX24) and CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1), which inhibit the transcriptional activity and promote the degradation of HY5, respectively. As such, PPKs regulate not only the binding of HY5 to its target genes under light conditions but also HY5 degradation when plants are transferred from light to dark. Our data identify a PPK-mediated phospho-code on HY5 that integrates the molecular mechanisms underlying the regulation of HY5 to precisely control plant photomorphogenesis.

PAX1 represses canonical Wnt signaling pathway and plays dual roles during endoderm differentiation

BackgroundPaired box 1 (PAX1) is a transcription factor and essential for the development of pharyngeal pouches-derived tissues, including thymus. PAX1 mutations are identified in Severe Combined Immunodeficiency (SCID) patients with Otofaciocervical Syndrome Type 2 (OTFCS2). However, despite the critical roles of PAX1 in embryonic development and diseases, detailed insights into its molecular mode of action are critically missing.MethodsThe repressing roles of PAX1 and SCID associated mutants on Wnt signaling pathway were investigated by luciferase reporter assays, qRT-PCR and in situ hybridization in HEK293FT, HCT116 cells and zebrafish embryos, respectively. Co-immunoprecipitation (co-IP) and western blotting assays were carried out to identify the molecular mechanisms underlying PAX1’s role on Wnt signaling pathway. hESC based endoderm differentiation, flow cytometry, high-throughput sequencing data analysis, and qRT-PCR assays were utilized to determine the roles of PAX1 during endoderm differentiation.ResultsHere, we show that PAX1 represses canonical Wnt signaling pathway in vertebrate cells. Mechanically, PAX1 competes with SUMO E3 ligase PIASy to bind to TCF7L2, thus perturbing TCF7L2 SUMOylation level, further reducing its transcriptional activity and protein stability. Moreover, we reveal that PAX1 plays dual roles in hESC-derived definitive and foregut/pharyngeal endoderm cells, which give rise to the thymus epithelium, by inhibiting Wnt signaling. Importantly, our data show PAX1 mutations found in SCID patients significantly compromise the suppressing ability of PAX1 on Wnt signaling.ConclusionsOur study presents a novel molecular mode of action of PAX1 in regulation of canonical Wnt signaling and endoderm differentiation, thus providing insights for the molecular basis of PAX1 associated SCID, offering better understanding of the behavior of PAX1 in embryogenesis.

Exploration the interaction of cadmium and copper toxic effects in pakchoi (Brassica chinensis L) roots through combinatorial transcriptomic and weighted gene co-expression network analysis

The interaction between cadmium(Cd) and copper(Cu) during combined pollution can lead to more complex toxic effects on humans and plants.However, there is still a lack of sufficient understanding regarding the types of interactions at the plant molecular level and the response strategies of plants to combined pollution. To assess this, we investigated the phenotypic and transcriptomic patterns of pakchoi (Brassica chinensis L) roots in response to individual and combined pollution of Cd and Cu. The results showed that compared to single addition, the translocation factor of heavy metals in roots significantly decreased (p < 0.05) under the combined addition, resulting in higher accumulation of Cd and Cu in the roots. Transcriptomic analysis of pakchoi roots revealed that compared to single pollution, there were 312 and 1926 differentially expressed genes (DEGs) specifically regulated in the Cd2Cu20 and Cd2Cu100 combined treatments, respectively. By comparing the expression of these DEGs among different treatments, we found that the combined pollution of Cd and Cu mainly affected the transcriptome of the roots in an antagonistic manner. Enrichment analysis indicated that pakchoi roots upregulated the expression of genes involved in glucosetransferase activity, phospholipid homeostasis, proton transport, and the biosynthesis of phenylpropanoids and flavonoids to resist Cd and Cu combined pollution. Using weighted gene co-expression network analysis (WGCNA), we identified hub genes related to the accumulation of Cd and Cu in the roots, which mainly belonged to the LBD, thaumatin-like protein, ERF, MYB, WRKY, and TCP transcription factor families. This may reflect a transcription factor-driven trade-off strategy between heavy metal accumulation and growth in pakchoi roots. Additionally, compared to single metal pollution, the expression of genes related to Nramp, cation/H+ antiporters, and some belonging to the ABC transporter family in the pakchoi roots was significantly upregulated under combined pollution. This could lead to increased accumulation of Cd and Cu in the roots. These findings provide new insights into the interactions and toxic mechanisms of multiple metal combined pollution at the molecular level in plants.

Ferulic acid alleviates avermectin induced renal injury in carp by inhibiting inflammation, oxidative stress and apoptosis

Avamectin (AVM), a macrolide antibiotic, is widely used in fisheries, agriculture, and animal husbandry, however, its irrational use poses a great danger to aquatic organisms. Ferulic acid (FA) is a natural chemical found in the cell walls of plants. It absorbs free radicals from the surrounding environment and acts as an antioxidant. However, the protective effect of FA against kidney injury caused by AVM has not been demonstrated. In this study, 60 carp were divided into the control group, AVM group (2.404 μg/L), FA+AVM group and FA group (400 mg/kg). Pathological examination, quantitative real-time PCR (qPCR), reactive oxygen species (ROS) and western blot were used to evaluate the preventive effect of FA on renal tissue injury after AVM exposure. Histological findings indicated that FA significantly reduced the swelling and infiltration of inflammatory cells in the kidney tissues of carp triggered by AVM. Dihydroethidium (DHE) fluorescent probe assay showed that FA inhibited the accumulation of kidney ROS. Biochemical results showed that FA significantly increased glutathione (GSH) content, total antioxidant capacity (T-AOC) and catalase (CAT) activity, and decreased intracellular malondialdehyde (MDA) content. In addition, western blot results revealed that the protein expression levels of Nrf2 and p-NF-κBp65 in the carp kidney were inhibited by AVM, but reversed by the FA. The qPCR results exhibited that FA significantly increased the mRNA levels of tgf-β1 and il-10, while significantly down-regulated the gene expression levels of tnf-α, il-6 and il-1β. These data suggest that FA can reduce oxidative stress and renal tissue inflammation induced by AVM. At the same time, FA inhibited the apoptosis of renal cells induced by AVM by decreasing the transcription level and protein expression level of Bax, and increasing the transcription level and protein expression level of Bcl2, PI3K and AKT. This study provides preliminary evidence for the theory that FA reduces the level of oxidative stress, inflammation response and kidney tissue damage caused by apoptosis in carp, providing a theoretical basis for the prevention and treatment of the AVM.

An overview of the two-component system GarR/GarS role on antibiotic production in Streptomyces coelicolor

The Streptomyces genus comprises Gram-positive bacteria known to produce over two-thirds of the antibiotics used in medical practice. The biosynthesis of these secondary metabolites is highly regulated and influenced by a range of nutrients present in the growth medium. In Streptomyces coelicolor, glucose inhibits the production of actinorhodin (ACT) and undecylprodigiosin (RED) by a process known as carbon catabolite repression (CCR). However, the mechanism mediated by this carbon source still needs to be understood. It has been observed that glucose alters the transcriptomic profile of this actinobacteria, modifying different transcriptional regulators, including some of the one- and two-component systems (TCSs). Under glucose repression, the expression of one of these TCSs SCO6162/SCO6163 was negatively affected. We aimed to study the role of this TCS on secondary metabolite formation to define its influence in this general regulatory process and likely establish its relationship with other transcriptional regulators affecting antibiotic biosynthesis in the Streptomyces genus. In this work, in silico predictions suggested that this TCS can regulate the production of the secondary metabolites ACT and RED by transcriptional regulation and protein–protein interactions of the transcriptional factors (TFs) with other TCSs. These predictions were supported by experimental procedures such as deletion and complementation of the TFs and qPCR experiments. Our results suggest that in the presence of glucose, the TCS SCO6162/SCO6163, named GarR/GarS, is an important negative regulator of the ACT and RED production in S. coelicolor.Key points• GarR/GarS is a TCS with domains for signal transduction and response regulation• GarR/GarS is an essential negative regulator of the ACT and RED production• GarR/GarS putatively interacts with and regulates activators of ACT and RED

Molecular Analysis of Antibiotic Resistance in CarbapenemResistant Pseudomonas aeruginosa Isolates Isolated from Clinical Samples

Pseudomonas aeruginosa is an opportunistic pathogen that causes increased morbidity and mortality in risky patient groups. Nowadays, carbapenem resistance has become a threat and resistance genes are spreading among species through mobile genetic elements. The dissemination of carbapenemases among P.aeruginosa is a serious public health concern due to its limited options for the treatment of bacterial infections. In this study, it was aimed to investigate the molecular epidemiology of 47 carbapenem resistant P.aeruginosa (CRPA) isolates derived from various clinical samples from the Central Laboratory Bacteriology Unit of Kocaeli University Research and Training Hospital between October 2021 and March 2023. The rates of resistance to the antibiotics, some carbapenemase and virulence genes, conjugative resistance plasmids, integron gene cassette contents and the clonal similarity of the isolates were investigated and then epidemiologically evaluated. In the study, identification of the bacterial isolates and their susceptibility to some antibiotics (imipenem, meropenem, aztreonam, amikacin, netilmicin, tobramycin, piperacillin, piperacillin/tazobactam, ceftazidime, cefepime, ciprofloxacin and levofloxacin) were determined by the VITEK® 2 Compact automated system. Metallo-beta-lactamase (MBL) production of the isolates was demonstrated by the imipenem/meropenem-EDTA (IMP/MEM-EDTA) combined disc method. Conjugation experiments were performed by the broth mating method. Alkali lysis method was used in plasmid DNA isolations. Co-transferred antibiotic resistances in transconjugants were detected by disc diffusion method. Carbapenemase genes (blaIMP , blaVIM , blaNDM , blaKPC and blaOXA-48 ), integron gene cassettes (class 1 and class 2) and virulence genes (lasR and rhlR) were screened by specific polymerase chain reactions (PCRs). Clonal relationships of the CRPA isolates were investigated by evaluating the DNA f ingerprintings obtained from the ERIC (enterobacterial repetitive intergenic consensus)-PCR assay. The highest resistance rate of the isolates were to levofloxacin, while the lowest resistance rates were observed against tobramycin, gentamicin and amikacin. MBL production was detected in 25 (53.2%) isolates. In conjugation experiments, 12 (25.5%) isolates were detected to harbour conjugative resistance plasmids. In 90% of the CRPA isolates, lasR and rhlR biofilm genes (encoding for the transcriptional activator protein) were detected by PCR. The blaVIM gene was detected in six (12.8%) isolates. The blaNDM gene was detected in five (10.6%) isolates and the blaOXA-48 gene was detected in three (6.4%) isolates. The blaKPC and blaIMP genes were not detected in CRPA isolates. It was determined that two (16.6%) of the isolates that carried the blaVIM gene, one (8.3%) carried the blaNDM gene and one (8.3%) carried the blaOXA-48 gene contained conjugative plasmids.In integron-specific PCRs, intI1 gene was positive in 39 (82.9%) isolates, while class 1 integron gene cassettes were detected in 24 isolates (51%). IntI1 positive six isolates were found to harbour class 1 integron gene cassettes-bearing conjugative plasmids. Class 2 integrons were not found in the CRPA isolates. Dendrogram analysis of ERIC-PCR patterns showed that there was no clonal similarity between the CRPA isolates and the isolates did not spread by cross-contamination. As a result, it has been observed that most of the CRPA isolates which have the potential to form biofilms, are highly resistant to other antibiotic groups other than carbapenems and can co-transfer some resistances (ceftazidime, cefepime, ciprofloxacin, levofloxacin, piperacillin-tazobactam) with conjugative resistance plasmids. It is thought that it would be useful to follow molecular epidemiology in the resistance gene reservoirs of these strains which have the potential to cause epidemics in the clinical arena.

Transpiration of neurological phenomena in cancer: Long‐term depression (LTD) abrogates cancer stem cell memory and sensitizes it to metabolic therapy

AbstractTherapy resistance and recurrence are irrefutably ascribed to the unique attributes of the cancer stem cell (CSC) population. Therefore, discerning the underlying mechanism is imperative in arbitrating a strategic therapeutic approach. In this study, we have observed the coexistence of neuron‐like cell (NLC) subpopulations in different cancer cell lines. The NLC subset was further validated by transfection studies and was found to be positive for PSD‐95. As a proof of concept, we have also demonstrated the transpiration of neurological phenomena such as long‐term potentiation (LTP) and long‐term depression (LTD) in cancer. We have utilized PMA drug and IL‐13 cytokine to study LTD and LTP, respectively, and observed a correlation between the memory marker (GLUN2B) and stemness markers (CD95 and CD58). Moreover, the transcriptional protein pCREB, which plays a crucial role in memory formation, was found to influence CD44 protein levels. These findings suggest that the instigation of LTD can impair cellular memory and sensitize CSC to the metabolic inhibitor, 3Bromopyruvate. Furthermore, this study interfaces the neurobiology of cancer and nanomedicine, wherein we have used biomimetic nanocarriers for the active targeting of drugs and demonstrate their ability to augment therapeutic efficiency.

Protein degradation in auxin response.

The signaling molecule auxin sits at the nexus of plant biology where it coordinates essentially all growth and developmental processes. Auxin molecules are transported throughout plant tissues and are capable of evoking highly specific physiological responses by inducing various molecular pathways. In many of these pathways, proteolysis plays a crucial role for correct physiological responses. This review provides a chronology of the discovery and characterization of the auxin receptor, which is a fascinating example of separate research trajectories ultimately converging on the discovery of a core auxin signaling hub that relies on degradation of a family of transcriptional inhibitor proteins-the Aux/IAAs. Beyond describing the "classical" proteolysis-driven auxin response system, we explore more recent examples of the interconnection of proteolytic systems, which target a range of other auxin signaling proteins, and auxin response. By highlighting these emerging concepts, we provide potential future directions to further investigate the role of protein degradation within the framework of auxin response.

Ancestral TALE homeobox protein transcription factor regulates actin dynamics and cellular activities of protozoan parasite Entamoeba invadens.

Entamoeba histolytica causes invasive amoebiasis, an important neglected tropical disease with a significant global health impact. The pathogenicity and survival of E. histolytica and its reptilian equivalent, Entamoeba invadens, relies on its ability to exhibit efficient motility, evade host immune responses, and exploit host resources, all of which are governed by the actin cytoskeleton remodeling. Our study demonstrates the early origin and the regulatory role of TALE homeobox protein EiHbox1 in actin-related cellular processes. Several genes involved in different biological pathways, including actin dynamics are differentially expressed in EiHbox1 silenced cells. EiHbox1 silenced parasites showed disrupted F-actin organization and loss of cellular polarity. EiHbox1's presence in the anterior region of migrating cells further suggests its involvement in maintaining cellular polarity. Loss of polarized morphology of EiHbox1 silenced parasites leads to altered motility from fast, directionally persistent, and highly chemotactic to slow, random, and less chemotactic, which subsequently leads to defective aggregation during encystation. EiHbox1 knockdown also resulted in a significant reduction in phagocytic capacity and poor capping response. These findings highlight the importance of EiHbox1 of E. invadens in governing cellular processes crucial for their survival, pathogenicity, and evasion of the host immune system.

Inhalation exposure-induced toxicity and disease mediated via mTOR dysregulation.

Environmental air pollution is a global health concern, associated with multiple respiratory and systemic diseases. Epidemiological supports continued urbanization and industrialization increasing the prevalence of inhalation exposures. Exposure to these inhaled pollutants induces toxicity via activation of numerous cellular mechanisms including oxidative stress, autophagy, disrupted cellular metabolism, inflammation, tumorigenesis, and others contributing to disease development. The mechanistic target of rapamycin (mTOR) is a key regulator involved in various cellular processes related to the modulation of metabolism and maintenance of homeostasis. Dysregulation of mTOR occurs following inhalation exposures and has also been implicated in many diseases such as cancer, obesity, cardiovascular disease, diabetes, asthma, and neurodegeneration. Moreover, mTOR plays a fundamental role in protein transcription and translation involved in many inflammatory and autoimmune diseases. It is necessary to understand inhalation exposure-induced dysregulation of mTOR since it is key regulator which may contribute to numerous disease processes. This mini review evaluates the available literature regarding several types of inhalation exposure and their impacts on mTOR signaling. Particularly we focus on the mTOR signaling pathway related outcomes of autophagy, lipid metabolism, and inflammation. Furthermore, we will examine the implications of dysregulated mTOR pathway in exposure-induced diseases. Throughout this mini review, current gaps will be identified related to exposure-induced mTOR dysregulation which may enable the targeting of mTOR signaling for the development of therapeutics.

Management of an older Marshall-Smith syndrome patient: a review of literature of MSS and craniosynostosis.

Marshall-Smith Syndrome (MSS) is a rare progressive developmental disorder that severely impairs a patient's intellectual development and physical health. The only known cause for MSS is a mutation in the nuclear factor 1 X (NFIX) gene. This mutation affects neuronal development and protein transcription. Historically, most patients with MSS do not survive beyond 3years of age. Reports of ocular findings are limited. We report a case of a 9-year-old MSS patient with progressive craniosynostosis, elevated intracranial pressure, and catastrophic ocular complications. A comprehensive PubMed literature search from 2018 to August 2022 updating a previous review of older literature produced 72 articles relating to MSS, which are reviewed.

SUMOylation of GMFB regulates its stability and function in retinal pigment epithelial cells under hyperglycemia

BackgroundGlia maturation factor beta (GMFB) is a growth and differentiation factor that acts as an intracellular regulator of signal transduction pathways. The small ubiquitin-related modifier (SUMO) modification, SUMOylation, is a posttranslational modification (PTM) that plays a key role in protein subcellular localization, stability, transcription, and enzymatic activity. Recent studies have highlighted the importance of SUMOylation in the inflammation and progression of numerous diseases. However, the relationship between GMFB and SUMOylation is unclear. ResultsHere, we report for the first time that GMFB and SUMO1 are markedly increased in retinal pigment epithelial (RPE) cells at the early stage of diabetes mellitus (DM) under hyperglycemia. The GMFΒ protein could be mono-SUMOylated by SUMO1 at the K20, K35, K58 or K97 sites. SUMOylation of GMFB led to its increased protein stability and subcellular translocation. Furthermore, deSUMOylation of GMFΒ downregulates multiple signaling pathways, including the Jak-STAT signaling pathway, p38 pathway and NF-kappa B signaling pathway. ConclusionsThis work provides novel insight into the role of SUMOylated GMFB in RPE cells and provides a novel therapeutic target for diabetic retinopathy (DR).

Single-cell transcriptomics unveiled that early life BDE-99 exposure reprogrammed the gut-liver axis to promote a proinflammatory metabolic signature in male mice at late adulthood.

Polybrominated diphenyl ethers (PBDEs) are legacy flame retardants that bioaccumulate in the environment. The gut microbiome is an important regulator of liver functions including xenobiotic biotransformation and immune regulation. We recently showed that neonatal exposure to polybrominated diphenyl ether-99 (BDE-99), a human breast milk-enriched PBDE congener, up-regulated proinflammation-related and down-regulated drug metabolism-related genes predominantly in males in young adulthood. However, the persistence of this dysregulation into late adulthood, differential impact among hepatic cell types, and the involvement of the gut microbiome from neonatal BDE-99 exposure remain unknown. To address these knowledge gaps, male C57BL/6 mouse pups were orally exposed to corn oil (10 ml/kg) or BDE-99 (57 mg/kg) once daily from postnatal days 2-4. At 15 months of age, neonatal BDE-99 exposure down-regulated xenobiotic and lipid-metabolizing enzymes and up-regulated genes involved in microbial influx in hepatocytes. Neonatal BDE-99 exposure also increased the hepatic proportion of neutrophils and led to a predicted increase of macrophage migration inhibitory factor signaling. This was associated with decreased intestinal tight junction protein (Tjp) transcripts, altered gut environment, and dysregulation of inflammation-related metabolites. ScRNA-seq using germ-free (GF) mice demonstrated the necessity of a normal gut microbiome in maintaining hepatic immune tolerance. Microbiota transplant to GF mice using large intestinal microbiome from adults neonatally exposed to BDE-99 down-regulated Tjp transcripts and up-regulated several cytokines in large intestine. In conclusion, neonatal BDE-99 exposure reprogrammed cell type-specific gene expression and cell-cell communication in liver towards proinflammation, and this may be partly due to the dysregulated gut environment.

Discovery of Highly Potent and Efficient CBP/p300 Degraders with Strong In Vivo Antitumor Activity.

The transcriptional coactivator cAMP response element binding protein (CREB)-binding protein (CBP) and its homologue p300 have emerged as attractive therapeutic targets for human cancers such as acute myeloid leukemia (AML). Herein, we report the design, synthesis, and biological evaluation of a series of cereblon (CRBN)-recruiting CBP/p300 proteolysis targeting chimeras (PROTACs) based on the inhibitor CCS1477. The representative compounds 14g (XYD190) and 14h (XYD198) potently inhibited the growth of AML cells with low nanomolar IC50 values and effectively degraded CBP and p300 proteins in a concentration- and time-dependent manner. Mechanistic studies confirmed that 14g and 14h can selectively bind to CBP/p300 bromodomains and induce CBP and p300 degradation in bromodomain family proteins in a CRBN- and proteasome-dependent manner. 14g and 14h displayed remarkable antitumor efficacy in the MV4;11 xenograft model (TGI = 88% and 93%, respectively). Our findings demonstrated that 14g and 14h are useful lead compounds and deserve further optimization and activity evaluation for the treatment of human cancers.

Hedyotis diffusa Willd and Astragalus membranaceus May Exert Anti-colon Cancer Effects by Affecting AKTI Expression, as Determined by Network Pharmacology and Molecular Docking.

Network pharmacology is a novel approach that uses bioinformatics to predict multitarget drugs and ingredient-target interactions in various diseases. A thorough search of previously published studies revealed that Hedyotis diffusa Willd (HDW) and Astragalus membranaceus (AM) possess anticancer activity. Colon cancer (CC) is one of the most common malignant tumors of the digestive tract and occurs in the colon. Herein, we explored the effect of two drugs in the treatment of CC. The present study aimed to predict and verify the effect of these two drugs in the treatment of CC. To explore the molecular mechanisms of the "HDW-AM" drug in the treatment of CC, we analyzed its principal efficiency in terms of ingredients, target spots, and pathways via network pharmacology, molecular docking, and experimental verification. The ingredients and their gene target sites were searched and screened through the TCMSP platform according to specific filtering conditions. Subsequently, components corresponding to the gene targets were chosen to construct the drug component-target network. The GEO (Gene Expression Omnibus) dataset was used to collect and screen for gene chips under CC and normal conditions, obtain differential genes, and construct a volcano map. The intersection genes between drug and disease targets were screened, the ".tsv" file was downloaded from the STRING platform and imported into Cytoscape 3.8.0 for visualization, a protein-protein interaction (PPI) network was constructed, the core targets were identified, and the common components with core targets were docked through Autodock Tools-1.5.6. Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were carried out through the Metascape platform to determine the major pathways. The CCK-8 (Cell Counting Kit-8) assay verified the effect of AKT1 on cell proliferation after treatment with quercetin. After the screening, 3658 DEGs (1841 downregulated and 1817 upregulated) were obtained from the GSE75970 gene chip; 21 active components and 220 targets were identified from the drugs. Subsequently, ten core genes (including AKT1, P53, and CASP3) and six major components were screened. GO functional analysis and KEGG analysis revealed that "HDWAM" regulates cell migration and motility through the combination of a transcription regulator complex, membrane rafts, vesicle lumen, and protein kinases via the MAPK, PI3K-Akt, and IL17 signaling pathways. The molecular docking results suggested that quercetin binds to AKT1, TP53, TNF, and CASP3. HDW-AM may exert a therapeutic effect on CC by modulating AKT1, TP53, TNF, and CASP3 and through signaling pathways. A CCK-8 cytotoxicity assay verified that quercetin affects cell viability through AKT1. The current study provides a theoretical basis for an in-depth investigation into the molecular mechanism of the "HDW-AM" drug in CC treatment via network pharmacology, molecular docking, and experimental verification.

Lineage-specific gene duplication and expansion of DUF1216 gene family in Brassicaceae.

Proteins containing domain of unknown function (DUF) are prevalent in eukaryotic genome. The DUF1216 proteins possess a conserved DUF1216 domain resembling to the mediator protein of Arabidopsis RNA polymerase II transcriptional subunit-like protein. The DUF1216 family are specifically existed in Brassicaceae, however, no comprehensive evolutionary analysis of DUF1216 genes have been performed. We performed a first comprehensive genome-wide analysis of DUF1216 proteins in Brassicaceae. Totally 284 DUF1216 genes were identified in 27 Brassicaceae species and classified into four subfamilies on the basis of phylogenetic analysis. The analysis of gene structure and conserved motifs revealed that DUF1216 genes within the same subfamily exhibited similar intron/exon patterns and motif composition. The majority members of DUF1216 genes contain a signal peptide in the N-terminal, and the ninth position of the signal peptide in most DUF1216 is cysteine. Synteny analysis revealed that segmental duplication is a major mechanism for expanding of DUF1216 genes in Brassica oleracea, Brassica juncea, Brassica napus, Lepidium meyneii, and Brassica carinata, while in Arabidopsis thaliana and Capsella rubella, tandem duplication plays a major role in the expansion of the DUF1216 gene family. The analysis of Ka/Ks (non-synonymous substitution rate/synonymous substitution rate) ratios for DUF1216 paralogous indicated that most of gene pairs underwent purifying selection. DUF1216 genes displayed a specifically high expression in reproductive tissues in most Brassicaceae species, while its expression in Brassica juncea was specifically high in root. Our studies offered new insights into the phylogenetic relationships, gene structures and expressional patterns of DUF1216 members in Brassicaceae, which provides a foundation for future functional analysis.

Brief research report: ETS-1 blockade increases ICAM-1 expression in activated human retinal endothelial cells.

Intercellular adhesion molecule 1 (ICAM-1) is a central cell adhesion molecule for retinal transendothelial migration of the leukocytes in non-infectious posterior uveitis. Inhibiting ICAM1 gene transcription reduces induction of ICAM-1 in inflamed retinal endothelium. Based on published literature implicating transcription factor ETS-1 as an activator of ICAM1 gene transcription, we investigated the effect of ETS-1 blockade on ICAM-1 levels in cytokine-stimulated human retinal endothelial cells. We first examined ICAM1 and ETS1 transcript expression in human retinal endothelial cells exposed to tumor necrosis factor-alpha (TNF-α) or interleukin-1beta (IL-1β). ICAM1 and ETS1 transcripts were increased in parallel in primary human retinal endothelial cell isolates (n = 5) after a 4-hour stimulation with TNF-α or IL-1β (p ≤ 0.012 and ≤ 0.032, respectively). We then assessed the effect of ETS-1 blockade by small interfering (si)RNA on cellular ICAM1 transcript and membrane-bound ICAM-1 protein. ETS1 transcript was reduced by greater than 90% in cytokine-stimulated and non-stimulated human retinal endothelial cell monolayers following a 48-hour treatment with two ETS-1-targeted siRNA, in comparison to negative control non-targeted siRNA (p ≤ 0.0002). The ETS-1 blockade did not reduce ICAM1 transcript expression nor levels of membrane-bound ICAM-1 protein, rather it increased both for a majority of siRNA-treatment and cytokine-stimulation conditions (p ≤ 0.018 and ≤ 0.004, respectively). These unexpected findings indicate that ETS-1 blockade increases ICAM-1 transcript and protein levels in human retinal endothelial cells. Thus ETS-1-targeting would be expected to promote rather than inhibit retinal transendothelial migration of leukocytes in non-infectious posterior uveitis.

Bioinformatic Analysis of Metabolomic Data: From Raw Spectra to Biological Insight

Metabolites are at the end of the gene–transcript–protein–metabolism cascade. As such, metabolomics is the omics approach that offers the most direct correlation with phenotype. This allows, where genomics, transcriptomics and proteomics fail to explain a trait, metabolomics to possibly provide an answer. Complex phenotypes, which are determined by the influence of multiple small-effect alleles, are an example of these situations. Consequently, the interest in metabolomics has increased exponentially in recent years. As a newer discipline, metabolomic bioinformatic analysis pipelines are not as standardized as in the other omics approaches. In this review, we synthesized the different steps that need to be carried out to obtain biological insight from annotated metabolite abundance raw data. These steps were grouped into three different modules: preprocessing, statistical analysis, and metabolic pathway enrichment. We included within each one of them the different state-of-the-art procedures and tools that can be used depending on the characteristics of the study, providing details about each method’s characteristics and the issues the reader might encounter. Finally, we introduce genome-scale metabolic modeling as a tool for obtaining pseudo-metabolomic data in situations where their acquisition is difficult, enabling the analysis of the resulting data with the modules of the described workflow.

An integrated approach to predict activators of NRF2 - the transcription factor for oxidative stress response

A variety of environmental and physiological conditions can cause oxidative stress that damage cellular components such as DNA, proteins and lipids. Oxidative stress is implicated in many human diseases including cancer, cardiovascular diseases, neurological diseases, inflammatory diseases, and aging. The nuclear factor erythroid 2–related factor 2 (NRF2) is a transcriptional factor that plays a key role in the cellular antioxidant defense system as it regulates transcription of antioxidant proteins and detoxifying enzymes. There is an urgent need to identify novel compounds that activate NRF2 and enhance antioxidant defense. We collected data from the high-throughput screening of NRF2 activators and identified molecular fragments (structural alerts) associated with the activation of NRF2. We also developed ten classification models using different types of molecular descriptors and machine learning techniques. Two approaches were used to establish the applicability domain of developed models: the structure-based approach and the distance to model approach. The best performing model that used message passing neural network (MPNN) technique showed accuracy of 87 % for the test set of chemicals within the distance to model of 0.3. The integrative approach using a combination of generated structural alerts and MPNN model was used to screen approved drugs collected in the DrugBank to identify potential NRF2 activators. Out of 2393 screened chemicals 138 compounds were predicted as NRF2 activators by both approaches. Analysis of these compounds showed that some drugs were already known activators of NRF2 while others are potentially novel activators.