Gene Activation Research Articles

Multiomics Analyses Reveal that Fatty Acid Metabolism and TCA Cycle Contribute to the Achievement of Functional Cure in Chronic Hepatitis B.

Peg-IFNα is one of the current therapeutic strategies for Hepatitis B virus (HBV) seroclearance. Nevertheless, the underlying mechanisms are not yet adequately understood. The objective of this study was to explore the potential mechanisms using multiomics approach. For the first time, we revealed the transcriptomic, proteomic, and metabolomic characterizations of Peg-IFNα-induced HBsAg seroclearance. We found that Peg-IFNα caused significant changes during the treatment. Patients who achieved HBsAg seroclearance were characterized as having decreased transcriptional activity of genes involved in fatty acid metabolism and the glycolysis/gluconeogenesis pathway, with up-regulated expression of fatty acid degradation-related proteins. Consistently, mitochondrial TCA cycle metabolites, including citric, isocitric, and malic acids, were significantly elevated in patients who achieved HBsAg seroclearance. We also observed up-regulated transcriptional activity of NK cell-mediated cytotoxicity, positive regulation of B cell activation, immunoglobulin production, and T cell receptor complex in functional-cured patients. Conversely, the metabolites associated with unsaturated fatty acid biosynthesis were increased in HBsAg persistent patients, and the transcriptional activity of immunoglobulin production and T cell receptor complex was down-regulated after 48 weeks of Peg-IFNα treatment. Our findings provided valuable resources to better understand the process of HBsAg seroclearance and shed new light on the pathways to facilitate higher functional cure rates for CHB.

Immune Checkpoint Inhibitor Myopathy: The Double-Edged Sword of Cancer Immunotherapy.

Immune checkpoint inhibitor (ICI) therapy has revolutionized the treatment of several malignancies, with improved survival. These monoclonal antibodies target immune checkpoints, including cytotoxic T-lymphocyte-associated protein 4 (ipilimumab and tremelimumab), programmed death 1 (nivolumab, pembrolizumab, cemiplimab, and dostarlimab), programmed death ligand 1 (atezolizumab, avelumab, and durvalumab), and lymphocyte activation gene 3 (relatlimab), and effectively augment the immune response against tumor cells. Releasing the brakes on the immune system has consequences, however, in the form of immune-related adverse events (irAEs), which may affect any organ. Neurologic irAEs represent 1%-3% of all irAEs, with immune-mediated myopathy (ICI myopathy) being the most common manifestation. Recent large patient series and systematic reviews have established the key features and highlighted new insights into ICI myopathy. ICI myopathy is characterized by an acute or subacute onset of oculobulbar and/or proximal limb weakness, with or without associated respiratory insufficiency and myocarditis. Creatine kinase elevation is common. Oculobulbar presentations with or without respiratory failure may be misattributed to neuromuscular junction disorders, particularly because acetylcholine receptor antibodies are present in up to 40% of patients; however, an electrodiagnostic evidence of a defect of neuromuscular transmission is often absent even in patients with severe weakness, highlighting that the myopathic process is the driving force behind these presentations. Muscle histopathology commonly demonstrates a unique signature of multifocal clusters of necrotic and regenerating fibers, differentiating ICI myopathy from other autoimmune myopathies. Transcriptomic analysis has uncovered distinct subgroups within ICI myopathy, revealing varying degrees of type 1 and type 2 interferon pathway activation alongside notable upregulation of the interleukin (IL)-6 pathway in affected muscle tissue. This discovery presents a promising avenue for intervention through the use of therapies that suppress the interferon pathway and target IL-6 or its receptor. Despite clinical improvements with immunomodulatory therapy, with corticosteroids the mainstay of treatment, mortality remains high, particularly in those with associated myocarditis or respiratory failure requiring intubation, where mortality occurs in up to 50%. ICI withdrawal can lead to cancer progression and death, highlighting a need for improved approaches to ICI rechallenge, performed in limited patients with variable success to date.

Chromatin topology dynamics in gene activation.

Chromatin topology dynamics in gene activation.

The Resistance to EGFR-TKIs in Non-Small Cell Lung Cancer

Lung cancer has emerged as a highly aggressive malignancy posing a significant global health hazard, with a particular concern for the rising incidence and mortality of gene-altered non-small cell lung cancer (NSCLC). As the prevalence of NSCLC rises, the investigation and selection of tumor treatment techniques are gaining significance. The treatment of individuals with genetic mutations in advanced NSCLC has emphasized the application of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs). Early and subsequent generations of EGFR-TKIs have demonstrated clinical efficacy in NSCLC patients carrying select genetic alterations. Osimertinib, a third-generation EGFR-TKI, has been sanctioned as the standard first-line therapy and is also utilized as a second-line option for individuals who develop the T790M resistance mutation following prior EGFR-TKI treatment. Approximately 10 to 30 percent of patients diagnosed with NSCLC exhibit EGFR mutations. This leads to abnormal activation of cancer genes and inactivation of tumor suppressor genes, resulting in poor prognosis for patients. This paper aims to analyze the development and resistance processes of EGFR inhibitors, thereby enhancing the comprehension of the resistance pathways to EGFR-TKIs.

Cbf11 and Mga2 function together to activate transcription of lipid metabolism genes and promote mitotic fidelity in fission yeast.

Within a eukaryotic cell, both lipid homeostasis and faithful cell cycle progression are meticulously orchestrated. The fission yeast Schizosaccharomyces pombe provides a powerful platform to study the intricate regulatory mechanisms governing these fundamental processes. In S. pombe, the Cbf11 and Mga2 proteins are transcriptional activators of non-sterol lipid metabolism genes, with Cbf11 also known as a cell cycle regulator. Despite sharing a common set of target genes, little was known about their functional relationship. This study reveals that Cbf11 and Mga2 function together in the same regulatory pathway, critical for both lipid metabolism and mitotic fidelity. Deletion of either gene results in a similar array of defects, including slow growth, dysregulated lipid homeostasis, impaired cell cycle progression (cut phenotype), abnormal cell morphology, perturbed transcriptomic and proteomic profiles, and compromised response to the stressors camptothecin and thiabendazole. Remarkably, the double deletion mutant does not exhibit a more severe phenotype compared to the single mutants. In addition, ChIP-nexus analysis reveals that both Cbf11 and Mga2 bind to nearly identical positions within the promoter regions of target genes. Interestingly, Mga2 binding appears to be dependent on the presence of Cbf11 and Cbf11 likely acts as a tether to DNA, while Mga2 is needed to activate the target genes. In addition, the study explores the distribution of Cbf11 and Mga2 homologs across fungi. The presence of both Cbf11 and Mga2 homologs in Basidiomycota contrasts with Ascomycota, which mostly lack Cbf11 but retain Mga2. This suggests an evolutionary rewiring of the regulatory circuitry governing lipid metabolism and mitotic fidelity. In conclusion, this study offers compelling support for Cbf11 and Mga2 functioning jointly to regulate lipid metabolism and mitotic fidelity in fission yeast.

Molecular mechanisms of action of curaxin CBL0137 on breast cancer cells in vitro

Introduction. Breast cancer (BC) represents a group of malignant neoplasms with various molecular profiles, which are characterized by aberrations in the mechanisms of epigenetic transcription regulation. One of these disruptions associated with a worse prognosis is the overexpression of the BET protein family, responsible for the interaction of transcription factors with histone-rich acetylated regions. Previously, for the multitargeted epigenetically active agent curaxin CBL0137 (CBL), we have shown the ability to inhibit the expression of BRD2, BRD3, BRD4 proteins in HeLa TI cells and BRD3, BRD4 proteins in BC cells.Aim. To analyze the molecular mechanisms of CBL0137 action on BC cells in vitro, including: 1) assessment of cytotoxicity, 2) analysis of the effect on the cell cycle, 3) assessment of the ability to trigger apoptosis, 4) cause DNA damage, and 5) analyze the effect on the expression of genes involved in proliferation, apoptosis and repair processes.Materials and methods. The cytotoxicity of CBL on BC cells (MCF7, MDA-MB-231, SKBR3) was assessed using the MTT assay. The effect of the agent on the cell cycle and activation of apoptosis was analyzed by flow cytometry. Analysis of DNA damage under the action of CBL was performed using the comet assay. Changes in the expression of genes associated with proliferation, repair and apoptosis were evaluated using real-time polymerase chain reaction.Results. The half maximal inhibitory concentration (IC50) on BC cells under the action of CBL0137 were 1 μM at 72-hour exposure, 14–25 μM at 24-hour. Curaxin CBL (0.5 and 1 μM) caused G2/M cell cycle arrest, and also triggered apoptosis in all cell lines. Under the action of 1 μM CBL, an increase in the degree of DNA damage in MCF7 and SKBR3 cells was recorded, and under the action of both concentrations in MDA-MB-231 cells. The gene expression profile involved in proliferation also corresponded to the arrest in the G2/M phase of the cell cycle. Also, under the action of CBL, the activation of p53-dependent repair genes occurred. An increase in the expression of pro-apoptotic and a decrease in anti-apoptotic genes was shown.Conclusion. Curaxin CBL0137 showed differential effects on molecular processes in BC cells. We have identified a cytostatic effect of the compound, confirmed at various molecular levels. The data obtained indicate the ability of CBL0137 to inhibit tumor processes in BC cells, which makes it a potentially interesting agent for combination therapy.

Differing genetics of saline and cocaine self-administration in the hybrid mouse diversity panel.

To identify genes involved in regulating the behavioral and brain transcriptomic response to the potentially addictive drug cocaine, we performed genome-wide association studies (GWASs) for intravenous self-administration of cocaine or saline (as a control) over 10 days using a panel of inbred and recombinant inbred mice. A linear mixed model increased statistical power for these longitudinal data and identified 145 loci for responding when saline only was delivered, compared to 17 for the corresponding cocaine GWAS. Only one locus overlapped. Transcriptome-wide association studies (TWASs) using RNA-Seq data from the medial frontal cortex and nucleus accumbens identified 5031434O11Rik and Zfp60 as significant for saline self-administration. Two other genes, Myh4 and Npc1 , were nominated based on proximity to loci for multiple endpoints or a cis locus regulating expression. All four genes have previously been implicated in locomotor activity. Our results indicate distinct genetic bases for saline and cocaine self-administration, and suggest some common genes for saline self-administration and locomotor activity.

Comparative liver transcriptome analysis in hamsters infected with food-borne trematodes Opisthorchis felineus, Opisthorchis viverrini, or Clonorchis sinensis.

Epidemiologically important food-borne trematodes Opisthorchis viverrini and Clonorchis sinensis are recognized as biological carcinogens of Group 1A, while Opisthorchis felineus is in Group 3 as noncarcinogenic to humans. Mechanisms of the biological carcinogenesis are still elusive. Some studies highlight chronic inflammation as a key factor and common pathway for cancer initiation and progression. Nonetheless, the chronic inflammation alone does not explain why these three species differ in carcinogenicity. We focused this study on genome-wide landscapes of liver gene expression and activation of cellular pathways in Mesocricetus auratus golden hamsters infected with C. sinensis (South Korea), O. viverrini (Thailand), or O. felineus (Russia) at 1 and 3 months after infection initiation. Liver transcriptomes of golden hamsters (HiSeq Illumina, 2X150 bp) were sequenced at 1 and 3 months postinfection. Data processing was carried out using the following bioinformatic and experimental approaches: analysis of differential gene expression, estimates of proportions of affected liver cell types, liver histopathology, and examination of weighted gene coexpression networks. All infections caused enrichment with inflammatory response signaling pathways, fibrogenesis and cell proliferation, and IL2-STAT5, TNF-NF-κB, TGF-β, Hippo, MAPK, and PI3K-Akt signaling pathways. Nevertheless, species-specific responses to each infection were noted too. We also identified species-specific responses of liver cell types, differentially expressed gene clusters, and cellular pathways associated with structural liver damage (such as periductal fibrosis, epithelial neoplasia, and inflammation). This is the first comparative analysis of gene expression landscapes in the liver of experimental animals infected with O. viverrini, O. felineus, or C. sinensis. The trematodes have species-specific effects on the hepatobiliary system by triggering signaling pathways, thereby leading to differences in the severity of hepatobiliary structural lesions and contributing to the pathogenicity of closely related foodborne trematodes.

Advancing public understanding of epigenetics: the EPIBOOST project’s video science outreach

ABSTRACT The public’s growing interest in epigenetics, particularly its applications in human health, underscores the significance of epigenetic mechanisms in regulating gene activity and promoting phenotypic diversity without altering DNA sequences. Within the framework of the Extended Evolutionary Synthesis, epigenetics is regarded as a non-genetic contributor to evolution. The EPIBOOST project, “BOOSting excellence in environmental EPIgenetics,” investigates epigenetic changes induced by contaminants in aquatic organisms and aims to enhance public understanding of epigenetics through science outreach videos. A multidisciplinary team comprising Biology, Education, and Design researchers produced the video “What is Epigenetics?” using a design-based research methodology and a design thinking approach. This process involved storyboard design, audio recording, image production, video editing, testing, refinement, and public dissemination. The interactive development of the storyboard and script aimed to efficiently communicate the significance of epigenetics, fostering public understanding of this emerging research area through an effective science communication strategy. This paper focuses on the development process for the first video, which aims to promote public awareness of epigenetics using the AEIOU framework (Awareness, Enjoyment, Interest, Opinions, and Understanding) to elucidate epigenetic concepts such as DNA methylation, gene expression, and phenotype, and their integration in environmental assessment frameworks.

Comprehensive Predictions of Mef2-Mediated Chromatin Loops, Which May Inhibit Ubx Binding by Blocking Low-Affinity Binding Sites

Gene regulation depends on the interaction between chromatin-associated factors, such as transcription factors (TFs), which promote chromatin loops to ensure tight contact between enhancer and promoter regions. So far, positive interactions that lead to gene activation have been the main focus of research, but regulations related to blocking or inhibiting factor binding are also essential to maintaining a defined cellular status. To understand these interactions in greater detail, I investigated the possibility of the muscle differentiation factor Mef2 to prevent early Hox factor binding, leading to the proper timing of regulatory processes and the activation of differentiation events. My investigations relied on a collection of publicly available genome-wide binding data sets of Mef2 and Ubx (as the Hox factor), Capture-C interactions, and ATAC-seq analysis in Mef2 mutant cells. The analysis indicated that Mef2 can form possible chromatin loops to Ubx-bound regions. These regions contain low-affinity Ubx binding sites, and the chromatin architecture is independent of Mef2’s function. High levels of Ubx may disrupt the loops and allow specific Ubx bindings to regulate defined targets. In summary, my investigations highlight that the use of many publicly available data sets enables computational approaches to make robust predictions and, for the first time, suggest a molecular function of Mef2 as a preventer of Hox binding, indicating that it may act as a timer for muscle differentiation.

Oral Delivery of miR146a Conjugated to Cerium Oxide Nanoparticles Improves an Established T Cell-Mediated Experimental Colitis in Mice

Background: Dysregulated inflammation and oxidative stress are strongly implicated in the pathogenesis of inflammatory bowel disease. We have developed a novel therapeutic that targets inflammation and oxidative stress. It is comprised of microRNA-146a (miR146a)-loaded cerium oxide nanoparticles (CNPs) (CNP-miR146a). We hypothesized that oral delivery of CNP-miR146a would reduce colonic inflammation in a mouse model of established, chronic, T cell-mediated colitis. Methods: The stability of CNP-miR146a and mucosal delivery was assessed in vitro with simulated gastrointestinal fluid and in vivo after oral gavage by quantitative real-time RT-PCR. The efficacy of orally administered CNP-miR146a was tested in mice with established colitis using the model of adoptive naïve T-cell transfer in recombinant activating gene 2 knockout (Rag2−/−) mice. Measured outcomes included histopathology; CD45+ immune cell infiltration; oxidative DNA damage (tissue 8-hydroxy-2′-deoxyguanosine; 8-OHdG); expression of IL-6 and TNF mRNA and protein, and flow cytometry analysis of lamina propria Th1 and Th17 cell populations. Results: miR146a expression remained stable in simulated gastric and intestinal conditions. miR146a expression increased in the intestines of mice six hours following oral gavage of CNP-miR146a. Oral delivery of CNP-miR146a in mice with colitis was associated with reduced inflammation and oxidative stress in the proximal and distal colons as evidenced by histopathology scoring, reduced immune cell infiltration, reduced IL-6 and TNF expression, and decreased populations of CD4+Tbet+IFNg+ Th1, CD4+RorgT+IL17+ Th17, as well as pathogenic double positive IFNg+IL17+ T cells. Conclusions: CNP-miR146a represents a novel orally available therapeutic with high potential to advance into clinical trials.

Molecular Study: Nitroreductase Enzyme and Nitroimmidazole Resistance Genes (Nim Genes) Effect on AMR to Metronidazole in Clostridium difficile

Nitroreductase enzymes and Nim genes have been observed to be a factor predisposing to reduced susceptibility to metronidazole in anaerobic bacteria. Therefore, this study was determined the effect of nitroreductase enzymes and nim genes in metronidazole susceptibility in UK C. difficile isolates. Three UK C. difficile ribotypes 027, 001, 106 including metronidazole reduced susceptible (CDRM) and metronidazole susceptible (CDSM) C. difficile strains, and 2 control strains from ribotype 010 (CDRM) and 038 (CDSM) were used. Control strain C. perfringens ATCC 3626 previously identified to be nitroreductase positive, and positive control for nimB which was Bacteroides fragilis strain BF8 was also included in this research. These selected strains were observed for their nitroreductase activity and presence of Nim genes. C. perfringens ATCC 3626 demonstrated nitroreductase activity but was absent in C. difficile control. Three CDRM strains (001 and 106 ribotype) demonstrated nitoreductase activity. None of the CDSM strains analysed showed detectable nitroreductase activity. The nim gene PCR amplification was only observed in the nim positive control strain BF8 (about 400 bp). All samples tested including ribotype 001 027 and 106 CDSM and CDRM strains showed no nimPCR product. In conclusion, despite the nitroreductase result obtained, inconsistencies observed during the assay showed methodology issues, thus using radio labelled metronidazole and monitoring the decline in the radiolabel signal proportional to the level of nitroreduction, maybe a preferred method in correlating nitroreductase activity to the CDRM phenotype. Also, this research observed the absence of nim genes in C. difficile UK ribotypes irrespective of metronidazole susceptibility patterns, depicting the inability of the universal primers for nimA-NimE to amplify a potential nim gene in C. difficile UK ribotypes. The universal primers were previously reported incapable of amplifying nimJ due to a different nucleotide sequence coding for same proteins as the universal primers.

Spatial Transcriptomics, Proteomics, and Epigenomics as Tools in Tissue Engineering and Regenerative Medicine

Spatial transcriptomics, proteomics, and epigenomics are innovative technologies which offer an unparalleled resolution and wealth of data in understanding and the interpretation of cellular functions and interactions. These techniques allow researchers to investigate gene and protein expressions at an individual cell level, revealing cellular heterogeneity within, for instance, bioengineered tissues and classifying novel and rare cell populations that could be essential for the function of the tissues and in disease processes. It is possible to analyze thousands of cells simultaneously, which gives thorough insights into the transcriptomic view of complex tissues. Spatial transcriptomics combines gene expressions with spatial information, conserving tissue architecture and making the mapping of gene activity across different tissue regions possible. Despite recent advancements in these technologies, they face certain limitations. Single-cell transcriptomics can suffer from technical noise and dropout events, leading to incomplete data. Its applicability has been limited by the complexity of data integration and interpretation, although better resolution and tools for the interpretation of data are developing fast. Spatial proteomics and spatial epigenomics provide data on the distribution of proteins and the gene regulatory aspects in tissues, respectively. The disadvantages of these approaches include rather costly and time-consuming analyses. Nevertheless, combining these techniques promises a more comprehensive understanding of cell function and tissue organization, which can be predicted to be useful in achieving better knowledge of cell guidance in tissue-engineered constructs and a higher quality of tissue technology products.

Green synthesized silicon dioxide nanoparticles (SiO2NPs) ameliorated the cadmium toxicity in melon by regulating antioxidant enzymes activity and stress-related genes expression

Green synthesized nanoparticles (NPs) are an eco-friendly and cost-effective approach to reduce heavy metal stress in plants. Among heavy metals, cadmium (Cd) possesses higher toxicity to the crops and ultimately reduces their growth and yield. The current study aims to evaluate the effectiveness of green synthesized SiO2NPs to reduce toxic effects of Cd in melon (Cucumis melo) by regulating physiological parameters, enhancing antioxidant enzyme activity, and modulating stress-related gene expression. The SiO2NPs were synthesized using Artemisia annua plant extract having spherical shape and size within the range of 40-70 nm and characterized using advanced spectroscopic and analytical techniques. The application of SiO2NPs (75mg/L) significantly improved physiological parameters such as shoot length (SL), root length (RL), leaf fresh weight (LFW), root fresh weight (RFW), leaf dry weight (LDW) and root dry weight (RDW) by 14%, 20%, 15%, 16%, 14%, and 28%, respectively, compared to Cd-stressed plants. Photosynthetic pigments (chlorophyll and carotenoids) showed a notable increase of 15% and 40%, respectively. Furthermore, the activities of antioxidant enzymes such as SOD, POD, CAT, and APX were enhanced by 28.67%, 35.45%, 32.07%, and 42.75%, respectively. In addition, applying SiO2NPs increased the concentration of macronutrients N, P, and K by 33%, 40%, and 37%, respectively, compared to Cd-stressed plants. Moreover, SiO2NPs upregulated the expression of several stress-related genes and reduced Cd accumulation in shoots and roots. This study reveals that green synthesized SiO2NPs effectively reduced the Cd toxicity in melon by improving morphological and physiological parameters, enhancing antioxidant enzyme activity, and regulating the expression of stress-related genes. These findings suggest that green synthesized SiO2NPs could play a crucial role in sustainable agriculture by protecting crops from heavy metal stress.

Synthesis and biological assessment of chalcone and pyrazoline derivatives as novel inhibitor for ELF3-MED23 interaction.

HER2 overexpression significantly contributes to the aggressive nature and recurrent patterns observed in various solid tumors, notably gastric cancers. Trastuzumab, HER2-targeting monoclonal antibody drug, has shown considerable clinical success; however, readily emerging drug resistance emphasizes the pressing need for improved interventions in HER2-overexpressing cancers. To address this, we proposed targeting the protein-protein interaction (PPI) between ELF3 and MED23 as an alternative therapeutic approach to trastuzumab. In this study, we synthesized a total of 26 compounds consisting of 10 chalcones, 7 pyrazoline acetyl, and 9 pyrazoline propionyl derivatives, and evaluated their biological activity as potential ELF3-MED23 PPI inhibitors. Upon systematic analysis, candidate compound 10 was selected due to its potency in downregulating reporter gene activity of ERBB2 promoter confirmed by SEAP activity and its effect on HER2 protein and mRNA levels. Compound 10 effectively disrupted the binding interface between the ELF3 TAD domain and the 391-582 amino acid region of MED23, resulting in successful inhibition of the ELF3-MED23 PPI. This intervention led to a substantial reduction in HER2 levels and its downstream signals in the HER2-positive gastric cancer cell line. Subsequently, compound 10 induced significant apoptosis and anti-proliferative effects, demonstrating superior in vitro and in vivo anticancer activity overall. We found that the anticancer activity of compound 10 was not only restricted to trastuzumab-sensitive cases, but was also valid for trastuzumab-refractory clones. This suggests its potential as a viable therapeutic option for trastuzumab-resistant gastric cancers. In summary, compound 10 could be a novel alternative therapeutic strategy for HER2-overexpressing cancers, overcoming the limitations of trastuzumab.

Synthesis and biological assessment of chalcone and pyrazoline derivatives as novel inhibitor for ELF3-MED23 interaction

HER2 overexpression significantly contributes to the aggressive nature and recurrent patterns observed in various solid tumors, notably gastric cancers. Trastuzumab, HER2-targeting monoclonal antibody drug, has shown considerable clinical success; however, readily emerging drug resistance emphasizes the pressing need for improved interventions in HER2-overexpressing cancers. To address this, we proposed targeting the protein-protein interaction (PPI) between ELF3 and MED23 as an alternative therapeutic approach to trastuzumab. In this study, we synthesized a total of 26 compounds consisting of 10 chalcones, 7 pyrazoline acetyl, and 9 pyrazoline propionyl derivatives, and evaluated their biological activity as potential ELF3-MED23 PPI inhibitors. Upon systematic analysis, candidate compound 10 was selected due to its potency in downregulating reporter gene activity of ERBB2 promoter confirmed by SEAP activity and its effect on HER2 protein and mRNA levels. Compound 10 effectively disrupted the binding interface between the ELF3 TAD domain and the 391–582 amino acid region of MED23, resulting in successful inhibition of the ELF3-MED23 PPI. This intervention led to a substantial reduction in HER2 levels and its downstream signals in the HER2-positive gastric cancer cell line. Subsequently, compound 10 induced significant apoptosis and anti-proliferative effects, demonstrating superior in vitro and in vivo anticancer activity overall. We found that the anticancer activity of compound 10 was not only restricted to trastuzumab-sensitive cases, but was also valid for trastuzumab-refractory clones. This suggests its potential as a viable therapeutic option for trastuzumab-resistant gastric cancers. In summary, compound 10 could be a novel alternative therapeutic strategy for HER2-overexpressing cancers, overcoming the limitations of trastuzumab.

Remodeling of cytoskeleton, chromatin, and gene expression during mechanical rejuvenation of aged human dermal fibroblasts.

Aging is associated with a progressive decline in cellular function. To reset the aged cellular phenotype, various reprogramming approaches, including mechanical routes, have been explored. However, the epigenetic mechanisms underlying cellular rejuvenation are poorly understood. Here, we studied the cytoskeletal, genome-wide chromatin and transcriptional changes in young, aged, and mechanically rejuvenated fibroblasts using immunofluorescence, RNA sequencing, and Hi-C experiments. The mechanically rejuvenated aged fibroblasts, that had partially reset their transcription to a younger cell state, showed a local reorganization of the interchromosomal contacts and lamina-associated domains. Interestingly, the observed chromatin reorganization correlated with the transcriptional changes. Immunofluorescence experiments in the rejuvenated state confirmed increased actomyosin contractility like younger fibroblasts. In addition, the rejuvenated contractile properties were maintained over multiple cell passages. Overall, our results give an overview of how changes in the cytoskeleton, chromatin, and gene activity are connected to aging and rejuvenation.

Arrayed CRISPR libraries for the genome-wide activation, deletion and silencing of human protein-coding genes.

Arrayed CRISPR libraries extend the scope of gene-perturbation screens to non-selectable cell phenotypes. However, library generation requires assembling thousands of vectors expressing single-guide RNAs (sgRNAs). Here, by leveraging massively parallel plasmid-cloning methodology, we show that arrayed libraries can be constructed for the genome-wide ablation (19,936 plasmids) of human protein-coding genes and for their activation and epigenetic silencing (22,442 plasmids), with each plasmid encoding an array of four non-overlapping sgRNAs designed to tolerate most human DNA polymorphisms. The quadruple-sgRNA libraries yielded high perturbation efficacies in deletion (75-99%) and silencing (76-92%) experiments and substantial fold changes in activation experiments. Moreover, an arrayed activation screen of 1,634 human transcription factors uncovered 11 novel regulators of the cellular prion protein PrPC, screening with a pooled version of the ablation library led to the identification of 5 novel modifiers of autophagy that otherwise went undetected, and 'post-pooling' individually produced lentiviruses eliminated template-switching artefacts and enhanced the performance of pooled screens for epigenetic silencing. Quadruple-sgRNA arrayed libraries are a powerful and versatile resource for targeted genome-wide perturbations.

Identification of potential biomarkers from amino acid transporter in the activation of hepatic stellate cells via bioinformatics.

The etiopathogenesis of hepatic stellate cells (HSC) activation has yet to be completely comprehended, and there has been broad concern about the interplay between amino acid transporter and cell proliferation. This study proposed exploring the molecular mechanism from amino acid transport-related genes in HSC activation by bioinformatic methods, seeking to identify the potentially crucial biomarkers. GSE68000, the mRNA expression profile dataset of activated HSC, was applied as the training dataset, and GSE67664 as the validation dataset. Differently expressed amino acid transport-related genes (DEAATGs), GO, DO, and KEGG analyses were utilized. We applied the protein-protein interaction analysis and machine learning of LASSO and random forests to identify the target genes. Moreover, single-gene GESA was executed to investigate the potential functions of target genes via the KEGG pathway terms. Then, a ceRNA network and a drug-gene interaction network were constructed. Ultimately, correlation analysis was explored between target genes and collagen alpha I (COL1A), alpha-smooth muscle actin (α-SMA), and immune checkpoints. We identified 15 DEAATGs, whose enrichment analyses indicated that they were primarily enriched in the transport and metabolic process of amino acids. Moreover, two target genes (SLC7A5 and SLC1A5) were recognized from the PPI network and machine learning, confirmed through the validation dataset. Then single-gene GESA analysis revealed that SLC7A5 and SLC1A5 had a significant positive correlation to ECM-receptor interaction, cell cycle, and TGF-β signaling pathway and negative association with retinol metabolism conversely. Furthermore, the mRNA expression of target genes was closely correlated with the COL1A and α-SMA, as well as immune checkpoints. Additionally, 12 potential therapeutic drugs were in the drug-gene interaction network, and the ceRNA network was constructed and visualized. SLC7A5 and SLC1A5, with their relevant molecules, could be potentially vital biomarkers for the activation of HSC.

Olikomycin A–A Novel Calcium‐Dependent Lipopeptide with Antibiotic Activity Against Multidrug‐Resistant Bacteria

AbstractResearch into new antibiotics is becoming increasingly important as antibiotic resistance increases worldwide. The genus Streptomyces in particular is able to produce a wide range of antimicrobial products due to the large number of biosynthetic gene clusters (BGCs) in its genome. However, not all BGCs are expressed under laboratory conditions. In this work, deletion of the gene wblA, encoding a global regulator of natural product biosynthesis and morphogenesis in Streptomyces, led to the production of a novel natural product, olikomycin A, in Streptomyces ghanaensis ATCC 14672. Complete structure elucidation revealed that olikomycin A belongs to a class of calcium‐dependent antibiotics known as non‐ribosomal peptide synthetase (NRPS)‐encoded acidic lipopeptides. These compounds exhibit remarkable antimicrobial activity in the presence of calcium. Insights into olikomycin A biosynthesis were provided by whole genome sequencing and gene inactivation studies, while bioactivity assays showed strong inhibition of the growth of multidrug‐resistant Gram‐positive pathogens via disrupting cell membrane integrity. Olikomycin A shows an antibiotic profile similar to that of daptomycin, which is already in clinical use.