Activation Of Transcription Factors Research Articles

Differential expressions of endoplasmic reticulum stress-associated genes in aortic dissection and their correlation with immune cell infiltration

To explore differentially expressed endoplasmic reticulum stress-associated genes (ERSAGs) in aortic dissection (AD) and their correlations with immune cell infiltration to identify new therapeutic targets for AD. Two AD mRNA expression datasets (GSE190635 and GSE98770) were downloaded from GEO database for analysis of differentially expressed genes between the aorta of AD patients and normal aorta using R software. ERSAGs dataset was downloaded from GeneCards website, and GeneMANIA database was used to analyze the protein-protein interaction network of the differentially expressed ERSAGs and the proteins interacting with these genes. Based on GSE98770 dataset we analyzed the distributions of 22 immune cells within the aortic wall of AD patients using CIBERSORT package of R software. Surgical aortic wall specimens were obtained from 10 AD patients and 10 non-AD patients for detecting AGER mRNA expression using qRTPCR, and the upstream transcriptional factors, miRNAs, and chemicals targeting AGER were analyzed using the TRRUST database and NetworkAnalyst database. Bioinformatic analysis suggested significant differential expression of AGER in AD, which interacted with 20 proteins involved in pattern recognition receptor signaling pathway, positive regulation of DNA-binding transcription factor activity, myeloid leukocyte migration, leukocyte migration, and regulation of the I-κB kinase/NF-κB signaling. In AD, AGER expression level was positively correlated with Treg cell abundance (r=0.59, P < 0.05). The results of qRT-PCR demonstrated significantly lower expression of AGER mRNA in AD than in non-AD patients (1.00±0.30 vs 1.76±0.68, P < 0.05). ROC curve analysis showed that at the cut-off value of 1.335, AGER had an AUC of 0.86 (95% CI: 0.67-1.00, P= 0.0073) for predicting AD. Three transcriptional factors, 3 miRNAs, and 27 chemicals were predicted in the AGER regulatory network. AGER is lowly expressed in the aorta of AD patients and may influence the occurrence of AD through Treg cells.

Decoding the functionality of plant transcription factors.

Transcription factors (TFs) intricately govern cellular processes and responses to external stimuli by modulating gene expression. TFs help plants to balance the trade-off between stress tolerance and growth, thus ensuring their long-term survival in challenging environments. Understanding the factors and mechanisms that define the functionality of plant TFs is of paramount importance for unravelling the intricate regulatory networks governing development, growth, and responses to environmental stimuli in plants. This review provides a comprehensive understanding of these factors and mechanisms defining the activity of TFs. Understanding the dynamic nature of TFs has practical implications for modern molecular breeding programmes, as it provides insights into how to manipulate gene expression to optimize desired traits in crops. Moreover, recent studies also report the functional duality of TFs, highlighting their ability to switch between activation and repression modes; this represents an important mechanism for attuning gene expression. Here we discuss what the possible reasons for the dual nature of TFs are and how this duality instructs the cell fate decision during development, and fine-tunes stress responses in plants, enabling them to adapt to various environmental challenges.

Transcription factors ERα and Sox2 have differing multiphasic DNA- and RNA-binding mechanisms.

Many transcription factors (TFs) have been shown to bind RNA, leading to open questions regarding the mechanism(s) of this RNA binding and its role in regulating TF activities. Here, we use biophysical assays to interrogate the k on, k off, and K d for DNA and RNA binding of two model human TFs, ERα and Sox2. Unexpectedly, we found that both proteins exhibit multiphasic nucleic acid-binding kinetics. We propose that Sox2 RNA and DNA multiphasic binding kinetics can be explained by a conventional model for sequential Sox2 monomer association and dissociation. In contrast, ERα nucleic acid binding exhibited biphasic dissociation paired with novel triphasic association behavior, in which two apparent binding transitions are separated by a 10-20 min "lag" phase depending on protein concentration. We considered several conventional models for the observed kinetic behavior, none of which adequately explained all the ERα nucleic acid-binding data. Instead, simulations with a model incorporating sequential ERα monomer association, ERα nucleic acid complex isomerization, and product "feedback" on isomerization rate recapitulated the general kinetic trends for both ERα DNA and RNA binding. Collectively, our findings reveal that Sox2 and ERα bind RNA and DNA with previously unappreciated multiphasic binding kinetics, and that their reaction mechanisms differ with ERα binding nucleic acids via a novel reaction mechanism.

Abstract B026: APOBEC3 promotes squamous differentiation via IL1A/AP-1 signaling

Abstract The APOBEC family of cytidine deaminase enzymes catalyze the conversion of cytosine to uracil in ssDNA and RNA substrates. The resulting C to U transition has been noted in the innate immune response to viral infection, and somatic hypermutation. However, APOBEC family members APOBEC3A (hA3A) and APOBEC3B (hA3B) have been implicated as drivers of the APOBEC3 mutational signatures in numerous cancer types. Notably, approximately 70% of single nucleotide variant mutations in bladder cancer can be attributed to APOBEC3 induced mutagenesis. Studies have implicated various APOBEC3 family members to be drivers of tumorigenesis, disease progression, and therapeutic resistance. To investigate APOBEC3’s role in bladder cancer, we have generated a novel, cre-inducible, pten and p53 knockout, to overexpress mouse Apobec3(mA3) in Upk3a+ cells (UPPA), compared to a model that does not overexpress mA3 (UPP). Apobec3 expression was shown to accelerate bladder tumor formation, indicated by a shorter median time of tumor latency in the UPPA model (43.3 weeks) compared to the UPP model (53.6 weeks). While both tumor models had presence of squamous differentiation, the extensiveness was more profound in the UPPA tumors. Transcriptional analysis of UPP and UPPA tumors, via scRNA sequencing, revealed an enrichment of squamous gene expression in the UPPA tumors. Utilizing mouse syngeneic bladder cancer cell lines and normal bladder urothelial cells, we overexpressed mA3 in a doxycycline inducible manner. Overexpression of mA3 was shown to prevent luminal differentiation. Additionally, mA3 overexpression resulted in the increase of IL-1a production. IL-1a production was sufficient to drive squamous differentiation in BBN963 and normal bladder urothelial cells. Blocking cFOS and IL-1R, individually, prevented expression of squamous genes. These results suggest that DNA damage induced by mA3 is sufficient to drive squamous differentiation via signaling through IL-1a resulting in AP-1 transcription factor activation. We then analyzed bladder cancer samples from The Cancer Genome Atlas and found that hA3A was the only APOBEC family member to be significantly associated with squamous gene expression. Interestingly, hA3A expression was enriched in the basal/squamous consensus subtype. To further interrogate the potential of hA3A being a driver of squamous differentiation in bladder cancer, we have also generated doxycycline inducible expression of hA3A and hA3B in syngeneic bladder cancer cell lines. Targeting IL-1a in various autoimmune diseases has resulted in FDA approval of antibodies targeting IL-1a and IL-1R. Given the clinical utility of targeting IL-1a, it is plausible to target this signaling axis for therapeutic benefit in bladder cancers with ongoing hA3A activity. In aggregate, our work demonstrates the promotion of an aggressive tumor state, driven by APOBEC mutagenesis and implicates hA3A as a possible driver of squamous differentiation in urothelial carcinoma. Citation Format: Andrew S. Truong, Michael S. Sturdivant, Mi Zhou, Wolfgang A. Beckabir, John Raupp, Ujjawal Manocha, Elliot D. Toomer, Ibardo A. Zambrano, Hung-Jui Tan, Marc A. Bjurlin, Angela B. Smith, Tracy L. Rose, Matthew I. Milowsky, Sara E. Wobker, Kathryn H. Gessner, Jeffrey S. Damrauer, William Y. Kim. APOBEC3 promotes squamous differentiation via IL1A/AP-1 signaling [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2024 May 17-20; Charlotte, NC. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(10_Suppl):Abstract nr B026.

Abstract PR002: Sarcomatoid Histological Variants of Canine Muscle-Invasive Bladder Cancer: Transcription Factor Activation Highlights Pathways of Epithelial-Mesenchymal Transformation Similar to Humans

Abstract The histologic heterogeneity of human Muscle Invasive Bladder Cancer (MIBC) is one of the main contributing factors to poor treatment outcomes. Recent reports on MIBC histological variants have highlighted increased levels of transcription factors (TF) associated with epithelial-mesenchymal transformation (EMT) and epigenetic regulation in the sarcomatoid histology variant of MIBC. Dogs with spontaneously developing MIBC have been shown to display similar histological and molecular features, in addition to closely mimicking the clinical behavior of human MIBC. Here, we add to previously reported data on histological variants in canine MIBC by evaluating 21 tumor biopsies for variant histology and performing bulk RNAseq analyses on 3 organoid cell lines derived from one sarcomatoid subvariant and two conventional urothelial carcinoma (UC). MIBC biopsies from 21 dogs were reviewed for variant histology by a human urologic pathologist. Organoids derived from the urine of canine MIBC patients were cultured, preserved in RNA later, and sequenced for mRNA expression to identify TF associated with sarcomatoid variants versus conventional UC. All canine MIBC biopsies were classified as invasive, high-grade UC. Histologic variants were identified as glandular differentiation in 7/21, sarcomatoid differentiation in 3/21, squamous differentiation in 3/21, micropapillary carcinoma in 1/21 MIBC samples and small nested variants in 1/21 samples; with the rest of the tumors classified as conventional UC. Raw RNAseq analyses revealed no clear clustering of expression profiles between the three organoid cell lines, indicating distinct RNA expression profiles between the sarcomatoid variants and conventional UC of canine MIBC. To evaluate differences in the expression of TF targets between the organoid cell lines, pathway analyses and VIPER algorithms were used. Organoids derived from sarcomatoid variant UC showed increased expression of Snail familiy transcription repressor 1 (SNAI1) along with decreased expression of cadherin 1 (CDH1), indicating a signature of EMT, similar to that previously identified in human MIBC organoids with sarcomatoid variant histology. Canine MIBC organoids derived from conventional UC histology samples showed a TF profile associated with proliferation, metastasis, and progression, such as increased Zinc finger E-box binding homeobox 1 (ZEB1), E2F transcription factor 3 (E2F3), and centromere protein F (CENPF), as previously described in humans with high grade, invasive UC. In conclusion, the frequency of some histologic variants in canine MIBC, such as sarcomatoid UC, appear to be higher than reported in human patients. TF analyses of canine MIBC-derived organoid cell lines showed evidence of EMT, similar to the sarcomatoid variants of MIBC in humans. Organoids derived from canine MIBC could potentially serve as a valuable model for evaluating the efficacy of novel treatment modalities for the rare histologic variants of UC. Citation Format: Karin Allenspach, Mohamed Elbadawy, Hannah Nocholson, Christopher Zdyrski, John Cheville, Eugene Douglass, Elizabeth W. Howerth, Lilian Oliveira, Jonathan P. Mochel. Sarcomatoid Histological Variants of Canine Muscle-Invasive Bladder Cancer: Transcription Factor Activation Highlights Pathways of Epithelial-Mesenchymal Transformation Similar to Humans [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2024 May 17-20; Charlotte, NC. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(10_Suppl):Abstract nr PR002.

A novel 12-membered ring non-antibiotic macrolide EM982 attenuates cytokine production by inhibiting IKKβ and IκBα phosphorylation

Antimicrobial resistance poses a serious threat to human health worldwide and its incidence continues to increase owing to the overuse of antibiotics and other factors. Macrolide antibiotics such as erythromycin (EM) have immunomodulatory effects in addition to their antibacterial activity. Long-term, low-dose administration of macrolides has shown clinical benefits in treating non-infectious inflammatory respiratory diseases. However, this practice may also increase the emergence of drug-resistant bacteria. In this study, we synthesized a series of EM derivatives, and screened them for two criteria: (i) lack of antibacterial activity and (ii) ability to suppress tumor necrosis factor-α (TNF-α) production in THP-1cells stimulated with lipopolysaccharide. Among the 37 synthesized derivatives, we identified a novel 12-membered ring macrolide EM982 that lacked antibacterial activity against Staphylococcus aureus and suppressed the production of TNF-α and other cytokines. The effects of EM982 on Toll-like receptor 4 (TLR4) signaling were analyzed using a reporter assay and Western blotting. The reporter assay showed that EM982 suppressed the activation of transcription factors, NF-κB and/or activator protein 1 (AP-1), in HEK293cells expressing human TLR4. Western blotting showed that EM982 inhibited the phosphorylation of both IκB kinase (IKK) β and IκBα, which function upstream of NF-κB, whereas it did not affect the phosphorylation of p38 mitogen-activated protein kinase, extracellular signal-regulated kinase, and c-Jun N-terminal kinase, which act upstream of AP-1. These results suggest that EM982 suppresses cytokine production by inhibiting phosphorylation of IKKβ and IκBα, resulting in the inactivation of NF-κB.

Effects of Cinnamon on Cancer Prevention and Progression

Cinnamon has been used medicinally for centuries, but recently research has suggested it may have a role in cancer prevention and potentially treatment. The search for alternative and subjunctive therapies is essential due to the public demand and the increasing cost of healthcare. Here we review the biologically active components of cinnamon and discuss the methods of potential cinnamon activity against cancer including: transcription factor regulation and kinase activity. Nuclear Factor kappa B (NF-\(𝜿\)B) is a stress sensitive transcription factor that regulates transcription of genes involved in tumor progression and is inhibited by cinnamon components. Another way that cinnamon inhibits tumor growth is by suppression of transcription factor activator protein -1 (AP-1) which interacts with genes responsible for apoptosis, metastasis and inflammation. Hypoxia-inducible transcription factor-1 (HIF-1) and vascular endothelial growth factor (VEGF) are involved in angiogenesis, especially in the tumor microenvironment. The HIF-1-VEGF pathway are targeted by cinnamaldehyde, a compound found in cinnamon. Nuclear factor erythroid related factor 2 (Nrf2) is also examined and has been indicated to both potentially prevent cancer as well as induce it; various cinnamon derivatives target Nrf2. A cinnamaldehyde derivative has been implicated in a reduction of the mitogen-activated protein kinases (MAPKs), which are a group of kinases that regulate proliferation. Additionally, cinnamon components have been tied to cancer prevention by positively affecting the gut microbiome and inhibiting inflammation. The review concludes with a discussion of the future research needed and potential risk associated with cinnamon intake.

Unobtrusive Health Hazard

Oxidative stress is a biological mechanism triggered by an imbalance between the biosynthesis and accumulation of reactive oxygen species (ROS) in a biological system. Normally, ROS are produced as byproducts of oxygen metabolism and fulfill various physiological roles such as cell signaling. However, environmental stressors like UV radiation, ionizing radiation, pollutants, heavy metals, and xenobiotics can also lead to increased ROS production. Common types of ROS include superoxide radicals (O2−), hydrogen peroxide (H2O2), hydroxyl radicals (•OH), and singlet oxygen (1O2), which are generated during various metabolic pathways. Key cellular processes such as protein phosphorylation, activation of transcriptional factors, apoptosis, immunity, and differentiation depend on maintaining appropriate levels of ROS within cells. Under conditions of excessive ROS production, which can occur during both normal physiological processes and pathological conditions like inflammation, ischemia, and aging, ROS can damage important cellular components such as proteins, lipids, and nucleic acids. This production is primarily mitochondrial but can also result from enzymatic activities involved in the respiratory chain, prostaglandin synthesis, phagocytosis, and the cytochrome P450 system. Both immune system activation and environmental factors contribute to endogenous and exogenous free radical production. The signs of oxidative stress include fatigue, memory loss, muscle or joint pain, the appearance of wrinkles and grey hair, decreased eyesight, headaches, increased sensitivity to noise, and a heightened susceptibility to infections. To reduce oxidative stress, it is advised to avoid sugar and processed foods while balancing blood sugar levels, prevent infections, incorporate daily stress reduction practices, avoid toxins, and promote the production of antioxidants through diet and herbal supplements that are rich in antioxidants.

Integrated Single-cell Multiomic Analysis of HIV Latency Reversal Reveals Novel Regulators of Viral Reactivation.

Despite the success of antiretroviral therapy, human immunodeficiency virus (HIV) cannot be cured because of a reservoir of latently infected cells that evades therapy. To understand the mechanisms of HIV latency, we employed an integrated single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin with sequencing (scATAC-seq) approach to simultaneously profile the transcriptomic and epigenomic characteristics of ∼ 125,000 latently infected primary CD4+ T cells after reactivation using three different latency reversing agents. Differentially expressed genes and differentially accessible motifs were used to examine transcriptional pathways and transcription factor (TF) activities across the cell population. We identified cellular transcripts and TFs whose expression/activity was correlated with viral reactivation and demonstrated that a machine learning model trained on these data was 75%-79% accurate at predicting viral reactivation. Finally, we validated the role of two candidate HIV-regulating factors, FOXP1 and GATA3, in viral transcription. These data demonstrate the power of integrated multimodal single-cell analysis to uncover novel relationships between host cell factors and HIV latency.

Scalable and unbiased sequence-informed embedding of single-cell ATAC-seq data with CellSpace

Standard scATAC sequencing (scATAC-seq) analysis pipelines represent cells as sparse numeric vectors relative to an atlas of peaks or genomic tiles and consequently ignore genomic sequence information at accessible loci. Here we present CellSpace, an efficient and scalable sequence-informed embedding algorithm for scATAC-seq that learns a mapping of DNA k-mers and cells to the same space, to address this limitation. We show that CellSpace captures meaningful latent structure in scATAC-seq datasets, including cell subpopulations and developmental hierarchies, and can score transcription factor activities in single cells based on proximity to binding motifs embedded in the same space. Importantly, CellSpace implicitly mitigates batch effects arising from multiple samples, donors or assays, even when individual datasets are processed relative to different peak atlases. Thus, CellSpace provides a powerful tool for integrating and interpreting large-scale scATAC-seq compendia.

Comparative transcriptomics analysis reveals defense mechanisms of Manihot esculenta Crantz against Sri Lanka Cassava MosaicVirus

BackgroundCassava mosaic disease (CMD), caused by Sri Lankan cassava mosaic virus (SLCMV) infection, has been identified as a major pernicious disease in Manihot esculenta Crantz (cassava) plantations. It is widespread in Southeast Asia, especially in Thailand, which is one of the main cassava supplier countries. With the aim of restricting the spread of SLCMV, we explored the gene expression of a tolerant cassava cultivar vs. a susceptible cassava cultivar from the perspective of transcriptional regulation and the mechanisms underlying plant immunity and adaptation.ResultsTranscriptomic analysis of SLCMV-infected tolerant (Kasetsart 50 [KU 50]) and susceptible (Rayong 11 [R 11]) cultivars at three infection stages—that is, at 21 days post-inoculation (dpi) (early/asymptomatic), 32 dpi (middle/recovery), and 67 dpi (late infection/late recovery)—identified 55,699 expressed genes. Differentially expressed genes (DEGs) between SLCMV-infected KU 50 and R 11 cultivars at (i) 21 dpi to 32 dpi (the early to middle stage), and (ii) 32 dpi to 67 dpi (the middle stage to late stage) were then identified and validated by real-time quantitative PCR (RT-qPCR). DEGs among different infection stages represent genes that respond to and regulate the viral infection during specific stages. The transcriptomic comparison between the tolerant and susceptible cultivars highlighted the role of gene expression regulation in tolerant and susceptible phenotypes.ConclusionsThis study identified genes involved in epigenetic modification, transcription and transcription factor activities, plant defense and oxidative stress response, gene expression, hormone- and metabolite-related pathways, and translation and translational initiation activities, particularly in KU 50 which represented the tolerant cultivar in this study.

Abstract 3064: Distinct Regulatory Programs Govern Smooth Muscle Cell Fate In Atherosclerosis

Introduction: During atherosclerosis, some vascular smooth muscle cells (SMCs) undergo phenotypic modulation, de-differentiating and adopting a fibroblast-like (FMC) or a chondrocyte/osteoblast-like phenotype (CMC). Although FMCs and CMCs appear to occur in succession in single-cell RNAseq data, they occupy distinct locations within the lesion and are associated with protective and deleterious plaque features, respectively. Hypothesis: Smooth muscle cell transitions to FMC and CMC occur via discrete transcriptional pathways during atherosclerosis. Methods: We performed single-cell RNA-sequencing (scRNAseq) and assay for transposase-accessible chromatin with sequencing (ATAC-seq) on lineage-traced vascular SMCs isolated from the aortic root of ApoE mice (n=3). Our analysis included pseudotime and RNA velocity analyses of the scRNAseq data, which independently identified a subgroup of Vcam+Cd74+ modulated SMCs as a potential cell state from which FMCs and CMCs appear to originate, each following distinct trajectories. We explored lineage-specific gene expression using the TradeSeq R package and conducted Ingenuity Pathway Analysis on differentially expressed genes to identify relevant upstream transcription factors, which were assayed for lineage-specific motif accessibility. Results: Our analysis showed profound changes in gene expression between the two trajectories as well as differential activation and chromatin accessibility of specific transcription factors at the point where FMCs diverge from CMCs. Within the CMC lineage, we noted heightened activation and chromatin accessibility of transcription factors associated with cellular stress and inflammatory responses, including Tp53, Foxo4, and Irf1. In stark contrast, these factors exhibited diminished activity within the FMC lineage during the same pseudotime interval. Furthermore, coronary artery disease GWAS transcription factors Twist1 and Tcf21 displayed differential motif accessibility over the FMC and CMC trajectories. Conclusions: Our study provides valuable insights into the regulatory mechanisms governing SMC phenotypic transitions in atherosclerosis, and further underscores the importance of GWAS genes in influencing these transitions.

Transcription factors as molecular switches regulating plant responses to drought stress.

Plants often experience abiotic stress, which severely affects their growth. With the advent of global warming, drought stress has become a pivotal factor affecting crop yield and quality. Increasing numbers of studies have focused on elucidating the molecular mechanisms underlying plant responses to drought stress. As molecular switches, transcription factors (TFs) are key participants in drought-resistance regulatory networks in crops. TFs regulate the transcription of downstream genes and are regulated by various upstream regulatory factors. Therefore, understanding the mechanisms of action of TFs in regulating drought stress can help enhance the adaptive capacity of crops under drought conditions. In this review, we summarize the structural characteristics of several common TFs, their multiple drought-response pathways, and recently employed research strategies. We describe the application of new technologies such as analysis of stress granule dynamics and function, multi-omics data, gene editing, and molecular crosstalk between TFs in drought resistance. This review aims to familiarize readers with the regulatory network of TFs in drought resistance and to provide a reference for examining the molecular mechanisms of drought resistance in plants and improving agronomic traits.

Machine learning identifies activation of RUNX/AP-1 as drivers of mesenchymal and fibrotic regulatory programs in gastric cancer.

Gastric cancer (GC) is the fifth most common cancer worldwide and is a heterogeneous disease. Among GC subtypes, the mesenchymal phenotype (Mes-like) is more invasive than the epithelial phenotype (Epi-like). Although gene expression of the epithelial-to-mesenchymal transition (EMT) has been studied, the regulatory landscape shaping this process is not fully understood. Here we use ATAC-seq and RNA-seq data from a compendium of GC cell lines and primary tumors to detect drivers of regulatory state changes and their transcriptional responses. Using the ATAC-seq data, we developed a machine learning approach to determine the transcription factors (TFs) regulating the subtypes of GC. We identified TFs driving the mesenchymal (RUNX2, ZEB1, SNAI2, AP-1 dimer) and the epithelial (GATA4, GATA6, KLF5, HNF4A, FOXA2, GRHL2) states in GC. We identified DNA copy number alterations associated with dysregulation of these TFs, specifically deletion of GATA4 and amplification of MAPK9 Comparisons with bulk and single-cell RNA-seq data sets identified activation toward fibroblast-like epigenomic and expression signatures in Mes-like GC. The activation of this mesenchymal fibrotic program is associated with differentially accessible DNA cis-regulatory elements flanking upregulated mesenchymal genes. These findings establish a map of TF activity in GC and highlight the role of copy number driven alterations in shaping epigenomic regulatory programs as potential drivers of GC heterogeneity and progression.

Elevation of miR-150-5p in Brain Microvascular Endothelial Cells as a Result of Prenatal Alcohol Exposure Is Mediated by Transcriptional Mechanisms

Fetal Alcohol Spectrum Disorders (FASD) encompass a variety of conditions impacting development in children exposed to alcohol in utero. Vascular alterations such as defective angiogenesis, blood brain barrier permeability, vessel reactivity, and blood flow have been found in various animal models of prenatal alcohol exposure (PAE). The underlying mechanisms of these vascular defects are not fully understood. Our lab previously showed that miR-150-5p was upregulated in brain microvascular endothelial cells (BMVECs) isolated from embryonic mouse cortices following moderate PAE. We identified downstream vascular targets of miR-150-5p in BMVECs and demonstrated that miR-150-5p inhibition could counter alcohol-mediated effects on BMVECS and the cortical microvasculature in vitro and in vivo, respectively. Here, we investigate upstream mechanisms that contribute to miR-150-5p elevation in BMVECs following PAE. We hypothesized that multiple mechanisms, including pathways at the transcriptional level, contribute to increased miR-150-5p abundance in BMVECs following ethanol (EtOH) exposure. Overall transcriptional activity at the miR-150 promoter was increased in BMVECs with EtOH treatment, and pri-miR-150, the primary transcript for miR-150-5p, was elevated in EtOH-treated BMVECs. Additionally, miR-150 promoter methylation was decreased in PAE cortices compared to saccharine-exposed controls (SAC). Current work aims to confirm the role of DNA methylation in transcriptional regulation of the miR-150 promoter in BMVECs and characterize DNA methyltransferase activity in BMVECs during PAE. We also assessed transcription factor binding at the miR-150 promoter and found that miR-150 promoter activation by several transcription factors was blunted with EtOH treatment. Current work involves the investigation of global transcription factor activation in BMVECs in PAE versus SAC. In summary, our research sheds light on multiple mechanisms of miR-150-5p regulation that may contribute to brain microvascular defects as a result of PAE. American Physiological Society Porter Physiology Development Fellowship University of New Mexico Health School of Medicine Department of Cell Biology and Physiology. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Molecular docking, dynamics, and preclinical experimental studies identify Morin hydrate as a potent PPAR-γ and NRF-2 transcription factor agonist that mitigates colon inflammation

Introduction: Peroxisome proliferator-activated receptor gamma (PPAR-γ) plays a vital role in regulating intestine inflammation. PPAR-γ is highly expressed in the adipose tissue and the colon. The nuclear erythroid 2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein1 (Keap1) system protects cells from oxidative stress and inflammation. Therefore, PPAR-γ and Nrf2 ligands offer a drug-targeting system that can be exploited to treat inflammatory bowel diseases (IBDs), especially ulcerative colitis. IBD prevalence is on the rise globally. Current mainstream therapies for IBD offer protection but with many side effects; therefore, up to 30% of IBD patients turn to alternate therapy using bioactive phytochemicals. Morin hydrate, a naturally occurring bioflavonoid in many fruits, stems, and leaves of the Moraceae family plant, possesses potent anti-inflammatory properties. The docking studies reveal that it is a potent activator of the PPAR-γ and Nrf2 transcription factors. Therefore, in this study, we investigated the role of morin hydrate modulating colon epithelial PPAR-γ transcription in colon inflammation. Aim: Therefore, this study aims to investigate morin hydrate’s anti-inflammatory properties using both in vivo and in vitro models of colon inflammation. Methodology: C57BL/6J black mice were administered 2% dextran sodium sulfate (DSS) morin hydrate administered at concentrations of 25 &amp; 50mg/kg body for in vivo studies. The DAI, colon length, MPO, and histology changes were determined. Proinflammatory cytokines (IL-1β, IL-6, TNF-α, &amp; IL17A) were measured using ELISA and mRNA using real-time PCR. PPAR-γ and Nrf2 expression and phosphoNF-κB/NF-κB expression were estimated. HT-29 cells were cultured in DMEM media and were stimulated using TNF-α (1ng/ml). Different concentrations of morin hydrate were treated to HT-29 cells to determine a safe dose range for further in vitro experiments by estimating cell toxicity assay. CXCL-1, IL-8, and mRNA levels were determined using real-time PCR after morin hydrate co-treatment. Morin hydrate effect on PPAR-γ and Nrf2 promoter activity was determined using full-length promoter transfection studies. Results: Morin treatment significantly ( p&lt;0.01) decreased DAI, MPO level, and restored colon length dose-dependently in the treated group. Histological scoring for colonic inflammation was significantly ( p&lt;0.001) improved upon morin treatment. Morin treatment also inhibited the proinflammatory cytokines (IL-1β, IL-6, TNF-α, &amp; IL17A) significantly ( p&lt;0.01) both at protein and mRNA levels. Morin significantly inhibited COX-2 and iNOS both at protein and mRNA levels and also decreased tissue nitrite levels. Morin also activated PPAR-γ protein expression NRF2 nuclear translocation and inhibited NF-κB expression in DSS-administered mice. Morin significantly decreased antioxidant surrogate markers superoxide dismutase (SOD) and catalase activity. Morin significantly ( p&lt;0.01) decreased proinflammatory cytokine (CXCL-1 &amp; IL-8) mRNA expression when HT-29 cells were challenged with TNF-α. Morin also increased PPAR-γ and Nrf2 promoter activity. Conclusion: Morin hydrate is a potent PPAR-γ and Nrf2 transcription factor activator that mitigates colon inflammation through its anti-inflammatory and antioxidant activities. This is supported from UAE University grant, fund number 12R006 and 12R169. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Genetic dependencies associated with transcription factor activities in human cancer cell lines

Transcription factors (TFs) are important mediators of aberrant transcriptional programs in cancer cells. In this study, we focus on TF activity (TFa) as a biomarker for cell-line-selective anti-proliferative effects, in that high TFa predicts sensitivity to loss of function of a given gene (i.e., genetic dependencies [GDs]). Our linear-regression-based framework identifies 3,047 pan-cancer and 3,952 cancer-type-specific candidate TFa-GD associations from cell line data, which are then cross-examined for impact on survival in patient cohorts. One of the most prominent biomarkers is TEAD1 activity, whose associations with its predicted GDs are validated through experimental evidence as proof of concept. Overall, these TFa-GD associations represent an attractive resource for identifying innovative, biomarker-driven hypotheses for drug discovery programs in oncology.

Emerging role of MYB transcription factors in cancer drug resistance.

Decades ago, the viral myeloblastosis oncogene v-myb was identified as a gene responsible for the development of avian leukemia. However, the relevance of MYB proteins for human cancer diseases, in particular for solid tumors, remained basically unrecognized for a very long time. The human family of MYB transcription factors comprises MYB (c-MYB), MYBL2 (b-MYB), and MYBL1 (a-MYB), which are overexpressed in several cancers and are associated with cancer progression and resistance to anticancer drugs. In addition to overexpression, the presence of activated MYB-fusion proteins as tumor drivers was described in certain cancers. The identification of anticancer drug resistance mediated by MYB proteins and their underlying mechanisms are of great importance in understanding failures of current therapies and establishing new and more efficient therapy regimens. In addition, new drug candidates targeting MYB transcription factor activity and signaling have emerged as a promising class of potential anticancer therapeutics that could tackle MYB-dependent drug-resistant cancers in a more selective way. This review describes the correlation of MYB transcription factors with the formation and persistence of cancer resistance to various approved and investigational anticancer drugs.

Understanding and computational design of genetic circuits of metabolic networks.

The expression of metabolic proteins is controlled by genetic circuits, matching metabolic demands and changing environmental conditions. Ideally, this regulation brings about a competitive level of metabolic fitness. Understanding how cells can achieve a robust (close-to-optimal) functioning of metabolism by appropriate control of gene expression aids synthetic biology by providing design criteria of synthetic circuits for biotechnological purposes. It also extends our understanding of the designs of genetic circuitry found in nature such as metabolite control of transcription factor activity, promoter architectures and transcription factor dependencies, and operon composition (in bacteria). Here, we review, explain and illustrate an approach that allows for the inference and design of genetic circuitry that steers metabolic networks to achieve a maximal flux per unit invested protein across dynamic conditions. We discuss how this approach and its understanding can be used to rationalize Escherichia coli's strategy to regulate the expression of its ribosomes and infer the design of circuitry controlling gene expression of amino-acid biosynthesis enzymes. The inferred regulation indeed resembles E. coli's circuits, suggesting that these have evolved to maximize amino-acid production fluxes per unit invested protein. We end by an outlook of the use of this approach in metabolic engineering applications.

Single-cell insights: pioneering an integrated atlas of chromatin accessibility and transcriptomic landscapes in diabetic cardiomyopathy

BackgroundDiabetic cardiomyopathy (DCM) poses a growing health threat, elevating heart failure risk in diabetic individuals. Understanding DCM is crucial, with fibroblasts and endothelial cells playing pivotal roles in driving myocardial fibrosis and contributing to cardiac dysfunction. Advances in Multimodal single-cell profiling, such as scRNA-seq and scATAC-seq, provide deeper insights into DCM’s unique cell states and molecular landscape for targeted therapeutic interventions.MethodsSingle-cell RNA and ATAC data from 10x Multiome libraries were processed using Cell Ranger ARC v2.0.1. Gene expression and ATAC data underwent Seurat and Signac filtration. Differential gene expression and accessible chromatin regions were identified. Transcription factor activity was estimated with chromVAR, and Cis-coaccessibility networks were calculated using Cicero. Coaccessibility connections were compared to the GeneHancer database. Gene Ontology analysis, biological process scoring, cell-cell communication analysis, and gene-motif correlation was performed to reveal intricate molecular changes. Immunofluorescent staining utilized various antibodies on paraffin-embedded tissues to verify the findings.ResultsThis study integrated scRNA-seq and scATAC-seq data obtained from hearts of WT and DCM mice, elucidating molecular changes at the single-cell level throughout the diabetic cardiomyopathy progression. Robust and accurate clustering analysis of the integrated data revealed altered cell proportions, showcasing decreased endothelial cells and macrophages, coupled with increased fibroblasts and myocardial cells in the DCM group, indicating enhanced fibrosis and endothelial damage. Chromatin accessibility analysis unveiled unique patterns in cell types, with heightened transcriptional activity in myocardial cells. Subpopulation analysis highlighted distinct changes in cardiomyocytes and fibroblasts, emphasizing pathways related to fatty acid metabolism and cardiac contraction. Fibroblast-centered communication analysis identified interactions with endothelial cells, implicating VEGF receptors. Endothelial cell subpopulations exhibited altered gene expressions, emphasizing contraction and growth-related pathways. Candidate regulators, including Tcf21, Arnt, Stat5a, and Stat5b, were identified, suggesting their pivotal roles in DCM development. Immunofluorescence staining validated marker genes of cell subpopulations, confirming PDK4, PPARγ and Tpm1 as markers for metabolic pattern-altered cardiomyocytes, activated fibroblasts and endothelial cells with compromised proliferation.ConclusionOur integrated scRNA-seq and scATAC-seq analysis unveils intricate cell states and molecular alterations in diabetic cardiomyopathy. Identified cell type-specific changes, transcription factors, and marker genes offer valuable insights. The study sheds light on potential therapeutic targets for DCM.