Induction Of Response Genes Research Articles

Identification of systemic nitrogen signaling in foxtail millet (Setaria italica) roots based on split-root system and transcriptome analysis.

The study established split-root system (SRS) in foxtail millet, and identified the molecular regulatory mechanisms and metabolic pathways related to systemic nitrogen signaling based on this system and transcriptome analysis. The growth of crops is primarily constrained by the availability of nitrogen (N), an essential nutrient. Foxtail millet (Setaria italica L.) is a significant orphan crop known for its strong tolerance to barren conditions. Despite this, the signaling pathway of nitrogen in foxtail millet remains largely unexplored. Identifying the candidate genes responsible for nitrogen response in foxtail millet is crucial for enhancing its agricultural productivity. This study utilized the split-root system (SRS) in foxtail millet to uncover genes associated with Systemic Nitrogen Signaling (SNS). Transcriptome analysis of the SRS revealed 2158 differentially expressed genes (DEGs) implicated in SNS, including those involved in cytokinin synthesis, transcription factors, E3 ubiquitin ligase, and ROS metabolism. Silencing of SiIPT5 and SiATL31 genes through RNAi in transgenic plants resulted in reduced SNS response, indicating their role in the nitrogen signaling pathway of foxtail millet. Furthermore, the induction of ROS metabolism-related genes in response to KNO3 of the split-root System (Sp.KNO3) suggests a potential involvement of ROS signaling in the SNS of foxtail millet. Overall, this study sheds light on the molecular regulatory mechanisms and metabolic pathways of foxtail millet in relation to SNS.

The Vacuolar H+-ATPase subunit C is involved in oligogalacturonide (OG) internalization and OG-triggered immunity

In plants, the perception of cell wall fragments initiates signal transduction cascades that activate the immune response. Previous research on early protein dynamics induced by oligogalacturonides (OGs), pectin fragments acting as damage-associated molecular patterns (DAMPs), revealed significant phosphorylation changes in several proteins. Among them, the subunit C of the vacuolar H+-ATPase, known as DE-ETIOLATED 3 (DET3), was selected to elucidate its role in the OG-triggered immune response. The Arabidopsis det3 knockdown mutant exhibited defects in H2O2 accumulation, mitogen-activated protein kinases (MAPKs) activation, and induction of defense marker genes in response to OG treatment. Interestingly, the det3 mutant showed a higher basal resistance to the fungal pathogen Botrytis cinerea that, in turn, was completely reversed by the pre-treatment with OGs. Our results suggest a compromised ability of the det3 mutant to maintain a primed state over time, leading to a weaker defense response when the plant is later exposed to the fungal pathogen. Using fluorescently labelled OGs, we demonstrated that endocytosis of OGs was less efficient in the det3 mutant, implicating DET3 in the internalization process of OGs. This impairment aligns with the observed defect in the priming response in the det3 mutant, underscoring that proper internalization and signaling of OGs are crucial for initiating and maintaining a primed state in plant defense responses.

The CssRS two-component system of Bacillus subtilis contributes to teicoplanin and polymyxin B response.

CssRS is a two-component system that plays a pivotal role in mediating the secretion stress response in Bacillus subtilis. This system upregulates the synthesis of membrane-bound HtrA family proteases that cope with misfolded proteins that accumulate within the cell envelope as a result of overexpression or heat shock. Recent studies have shown the induction of CssRS-regulated genes in response to cell envelope stress. We investigated the induction of the CssRS-regulated htrA promoter in the presence of different cell wall- and membrane-active substances and observed induction of the CssRS-controlled genes by glycopeptides (vancomycin and teicoplanin), polymyxins B and E, certain β-lactams, and detergents. Teicoplanin was shown to elicit remarkably stronger induction than vancomycin and polymyxin B. Teicoplanin and polymyxin B induced the spxO gene expression in a CssRS-dependent fashion, resulting in increased activity of Spx, a master regulator of disulfide stress in Bacillus subtilis. The CssRS signaling pathway and Spx activity were demonstrated to be involved in Bacillus subtilis resistance to teicoplanin and polymyxin B.

Multi-omics analysis provides insight into liver metabolism in yellow catfish (Pelteobagrus fulvidraco) under hypoxic stress

Dissolved oxygen is an environmental factor that affects fish survival. Aquatic environments suffer from an increasing oxygen deficiency resulting from global warming and environmental pollution. In this study, we combined high-throughput transcriptome analysis with non-targeted liquid chromatography-mass spectrometry-based metabolome sequencing to assess the metabolic response to hypoxia in the liver of Pelteobagrus fulvidraco. Healthy fish with body weight of 12.39 ± 1.37 g were subjected to hypoxic conditions at a concentration of 1.02 ± 0.09 mg/L for durations of 6, 24 and 48 h. Hypoxia exposure increased the content of MDA, lactate, glucose and the level of T-AOC, enhanced the activity of T-SOD, CAT, GPx and LDH, and decreased SDH activity. Histological examination revealed that liver tissue was damaged after exposure, and the degree of damage increased with the prolongation of hypoxia time. In the liver transcriptome, 1527 DEGs were identified, of which 240 were co-identified in the three comparison groups. These DEGs showed significant enrichment in pathways associated with oxidation-reduction and metabolic processes. RT- qPCR confirmed transcriptional induction of metabolism-related genes in response to hypoxia. Metabolomic results showed that the SDMs were related to amino acid, carbohydrate, and lipid metabolism. Moreover, a conjoint analysis demonstrated an increase in the activity of several key enzymes, such as phosphoenolpyruvate carboxykinase, phospholipid phosphatase 1a, and betaine-homocysteine methyltransferase. An integrated regulatory network was observed to be involved in glycine, serine, and threonine metabolism; glycolysis/gluconeogenesis; tricarboxylic acid cycle; protein digestion and absorption; and lipid metabolism in response to hypoxia exposure. Overall, the findings indicated that hypoxia can cause oxidative stress in yellow catfish and that their bodies can resist stress by activating the antioxidant defense system and mobilizing multiple metabolic pathways to meet the energy demand. These results will aid in the development of strategies aimed at mitigating the adverse effects of hypoxic exposure on fish.

Impaired reciprocal regulation between SIRT6 and TGF-β signaling in fatty liver.

Dysregulated transforming growth factor-beta (TGF-β) signaling contributes to fibrotic liver disease and hepatocellular cancer (HCC), both of which are associated with fatty liver disease. SIRT6limits fibrosis by inhibiting TGF-β signaling through deacetylating SMAD2 and SMAD3 and limits lipogenesis by inhibiting SREBP1 and SREBP2 activity. Here, we showed that, compared to wild-type mice, high-fat diet-induced fatty liver is worse in TGF-β signaling-deficient mice (SPTBN1+/- ) and the mutant mice had reduced SIRT6 abundance in the liver. Therefore, we hypothesized that altered reciprocal regulation between TGF-β signaling and SIRT6 contributes to these liver pathologies. We found that deficiency in SMAD3 or SPTBN1 reduced SIRT6mRNA and protein abundance and impaired TGF-β induction of SIRT6 transcripts, and that SMAD3 bound to the SIRT6 promoter, suggesting that an SMAD3-SPTBN1 pathway mediated the induction of SIRT6 in response to TGF-β. Overexpression of SIRT6 in HCC cells reduced the expression of TGF-β-induced genes, consistent with the suppressive role of SIRT6 on TGF-β signaling. Manipulation of SIRT6 abundance in HCC cells altered sterol regulatory element-binding protein (SREBP) activity and overexpression of SIRT6 reduced the amount of acetylated SPTBN1 and the abundance of both SMAD3 and SPTBN1. Furthermore, induction of SREBP target genes in response to SIRT6 overexpression was impaired in SPTBN1heterozygous cells. Thus, we identified a regulatory loop between SIRT6 and SPTBN1 that represents a potential mechanism for susceptibility to fatty liver in the presence of dysfunctional TGF-β signaling.

Hyperthermic intraperitoneal chemotherapy (HIPEC) with carboplatin induces distinct transcriptomic changes in ovarian tumor and normal tissues

ObjectiveTo determine the effect of hyperthermic intraperitoneal chemotherapy (HIPEC) with carboplatin on the transcriptomic profiles of normal and ovarian cancer (OC) tissues. MethodsNormal and tumor samples from four OCs were prospectively collected pre- and immediately post-HIPEC treatment and subjected to RNA-sequencing. Differential gene expression, gene ontology enrichment and pathway analyses were performed. Heat shock protein and immune-response protein expression was assessed using protein arrays and western blotting. ResultsRNA-sequencing revealed 4231 and 322 genes significantly differentially expressed between pre- and post-treatment normal and OC tissues, respectively (both adjusted p-value <0.05). Gene enrichment analyses demonstrated that the most significantly upregulated genes in normal tissues played a role in immune as well as heat shock response (both adjusted p < 0.001). In contrast, HIPEC induced an increased expression of primarily heat shock response and protein folding-related genes in tumor tissues (both adjusted p < 0.001). HIPEC-induced heat shock protein (HSP) expression changes, including in HSP90, HSP40, HSP60, and HSP70, were also observed at the protein level in both normal and tumor tissues. ConclusionsHIPEC with carboplatin resulted in an upregulation of heat shock-related genes in both normal and tumor tissue, with an additional immune response gene induction in normal and protein folding in tumor tissue. The findings of our exploratory study provide evidence to suggest that HIPEC administration may suffice to induce gene expression changes in residual tumor cells and raises a biological basis for the consideration of combinatorial treatments with HSP inhibitors.

TOR complex 2 contributes to regulation of gene expression via inhibiting Gcn5 recruitment to subtelomeric and DNA replication stress genes.

The fission yeast TOR complex 2 (TORC2) is required for gene silencing at subtelomeric regions and for the induction of gene transcription in response to DNA replication stress. Thus, TORC2 affects transcription regulation both negatively and positively. Whether these two TORC2-dependent functions share a common molecular mechanism is currently unknown. Here, we show that Gad8 physically interacts with proteins that regulate transcription, including subunits of the Spt-Ada-Gcn5-acetyltransferase (SAGA) complex and the BET bromodomain protein Bdf2. We demonstrate that in the absence of TORC2, Gcn5, the histone acetyltransferase subunit of SAGA, accumulates at subtelomeric genes and at non-induced promoters of DNA replication genes. Remarkably, the loss of Gcn5 in TORC2 mutant cells restores gene silencing as well as transcriptional induction in response to DNA replication stress. Loss of Bdf2 alleviates excess of Gcn5 binding in TORC2 mutant cells and also rescues the aberrant regulation of transcription in these cells. Furthermore, the loss of either SAGA or Bdf2 suppresses the sensitivity of TORC2 mutant cells to a variety of stresses, including DNA replication, DNA damage, temperature and nutrient stresses. We suggest a role of TORC2 in transcriptional regulation that is critical for gene silencing and gene induction in response to stress and involves the binding of Gcn5 to the chromatin.

IκBα is required for full transcriptional induction of some NFκB-regulated genes in response to TNF in MCF-7 cells

Inflammatory stimuli triggers the degradation of three inhibitory κB (IκB) proteins, allowing for nuclear translocation of nuclear factor-κB (NFκB) for transcriptional induction of its target genes. Of these three, IκBα is a well-known negative feedback regulator that limits the duration of NFκB activity. We sought to determine whether IκBα’s role in enabling or limiting NFκB activation is important for tumor necrosis factor (TNF)-induced gene expression in human breast cancer cells (MCF-7). Contrary to our expectations, many more TNF-response genes showed reduced induction than enhanced induction in IκBα knockdown cells. Mathematical modeling was used to investigate the underlying mechanism. We found that the reduced activation of some NFκB target genes in IκBα-deficient cells could be explained by the incoherent feedforward loop (IFFL) model. In addition, for a subset of genes, prolonged NFκB activity due to loss of negative feedback control did not prolong their transient activation; this implied a multi-state transcription cycle control of gene induction. Genes encoding key inflammation-related transcription factors, such as JUNB and KLF10, were found to be best represented by a model that contained both the IFFL and the transcription cycle motif. Our analysis sheds light on the regulatory strategies that safeguard inflammatory gene expression from overproduction and repositions the function of IκBα not only as a negative feedback regulator of NFκB but also as an enabler of NFκB-regulated stimulus-responsive inflammatory gene expression. This study indicates the complex involvement of IκBα in the inflammatory response to TNF that is induced by radiation therapy in breast cancer.

A novel bivalent chromatin associates with rapid induction of camalexin biosynthesis genes in response to a pathogen signal in Arabidopsis.

Temporal dynamics of gene expression underpin responses to internal and environmental stimuli. In eukaryotes, regulation of gene induction includes changing chromatin states at target genes and recruiting the transcriptional machinery that includes transcription factors. As one of the most potent defense compounds in Arabidopsis thaliana, camalexin can be rapidly induced by bacterial and fungal infections. Though several transcription factors controlling camalexin biosynthesis genes have been characterized, how the rapid activation of genes in this pathway upon a pathogen signal is enabled remains unknown. By combining publicly available epigenomic data with in vivo chromatin modification mapping, we found that camalexin biosynthesis genes are marked with two epigenetic modifications with opposite effects on gene expression, trimethylation of lysine 27 of histone 3 (H3K27me3) (repression) and acetylation of lysine 18 of histone 3 (H3K18ac) (activation), to form a previously uncharacterized type of bivalent chromatin. Mutants with reduced H3K27me3 or H3K18ac suggested that both modifications were required to determine the timing of gene expression and metabolite accumulation at an early stage of the stress response. Our study indicates that the H3K27me3-H3K18ac bivalent chromatin, which we name as kairostat, plays an important role in controlling the timely induction of gene expression upon stress stimuli in plants.

The effect of calcium phosphate biodegradable coatings of titanium implants on cell differentiation and apoptosis in rat bone tissue after experimental fracture.

The effectiveness of bone repair is determined by the balance of proliferative and destructive factors in the fracture union site. It can be enhanced by using various nanostructured materials possessing osteoinductive properties, in particular titanium implants with biodegradable calcium phosphate coatings. The effects of these coatings on the state of stem cells, their differentiation and distribution in the repair zone is unknown. To study the dynamics of proliferation, differentiation, and apoptosis of stem cells after experimental fracture followed by implantation of titanium implants with calcium phosphate coatings. The localization of proliferation (PCNA) and differentiation (CD44 and osteocalcin) factors and apoptotic molecules (MDM2, p53, caspase-3) was studied in a rat femoral fracture model with implant placement. Titanium implant screws with bioactive calcium phosphate and hydroxyapatite coatings formed by plasma electrolytic oxidation were used in the study. Experimental rats were arranged into three groups (15 animals per group): control group; rats implanted with uncoated implants; and rats implanted with coated implants. Control rats were subject to a similar fracture as experimental ones and were allowed to heal conservatively. Rats from all groups were sampled on days 7, 14, and 30 after injury. Low-differentiated PCNA-, osteocalcin-, and CD44-immunopositive cells were localized around the implant in the inner layer of the periosteum, layer of outer circumferential lamellae, and connective tissue lining of haversian canals. The spatial density of cells expressing the above proliferation and differentiation factors, as well as that of MDM2-immunoreactive cells, increased on day 7 and decreased by day 30 after injury. The spatial density of apoptotic cells reached the maximum on day 14 after injury. They were mainly found in the inner layer of the periosteum and outer circumferential lamellae. p53- and caspase-3-positive cells occurred on the surface of the concentric lamellae surrounding haversian canals and under the periosteum. Their spatial density decreased by day 30 after injury. Calcium phosphate coatings stimulate cell proliferation at early stages of fracture restoration and apoptotic cell death at later stages. Coating components may provide positional information guiding the differentiation of mesenchymal stromal cells. A change in the activity of apoptotic factors, osteocalcin, and CD44 is caused by gene induction in response to the diffusion of calcium phosphate compounds from coating to surrounding tissue.

Corticosterone Is Essential for Survival Through Frog Metamorphosis.

Thyroid hormone (TH) is required for frog metamorphosis, and corticosterone (CORT) increases TH signaling to accelerate metamorphic progression. However, a requirement for CORT in metamorphosis has been difficult to assess prior to the recent development of gene-editing technologies. We addressed this long-standing question using transcription activator-like effector nuclease (TALEN) gene disruption to knock out proopiomelanocortin (pomc) and disrupt CORT production in Xenopus tropicalis. As expected, mutant tadpoles had a reduced peak of plasma CORT at metamorphosis with correspondingly reduced expression of the CORT-response gene Usher syndrome type-1G (ush1g). Mutants had reduced rates of growth and development and exhibited lower expression levels of 2 TH response genes, Krüppel-like factor 9 (klf9) and TH receptor β (thrb). In response to exogenous TH, mutants had reduced TH response gene induction and slower morphological change. Importantly, death invariably occurred during tail resorption, unless rescued by exogenous CORT and, remarkably, by exogenous TH. The ability of exogenous TH by itself to overcome death in pomc mutants indicates that the CORT-dependent increase in TH signaling may ensure functional organ transformation required for survival through metamorphosis and/or may shorten the nonfeeding metamorphic transition to avoid lethal inanition.

CDK7 Mediates the Beta-Adrenergic Signaling in Thermogenic Brown and White Adipose Tissues.

SummaryCyclin-dependent kinases (CDKs) are emerging regulators of adipose tissue metabolism. Here we aimed to explore the role of CDK7 in thermogenic fat. We found that CDK7 brown adipose tissue (BAT)-specific knockout mice (Cdk7bKO) have decreased BAT mass and impaired β3-adrenergic signaling and develop hypothermia upon cold exposure. We found that loss of CDK7 in BAT disrupts the induction of thermogenic genes in response to cold. However, Cdk7bKO mice do not show systemic metabolic dysfunction. Increased expression of genes of the creatine metabolism compensates for the heat generation in the BAT of Cdk7bKO mice in response to cold. Finally, we show that CDK7 is required for beta 3-adrenergic agonist-induced browning of white adipose tissue (WAT). Indeed, Cdk7 ablation in all adipose tissues (Cdk7aKO) has impaired browning in WAT. Together, our results demonstrate that CDK7 is an important mediator of beta-adrenergic signaling in thermogenic brown and beige fat.

Mechanical Shielding in Plant Nuclei.

In animal single cells in culture, nuclear geometry and stiffness can be affected by mechanical cues, with important consequences for chromatin status and gene expression. This calls for additional investigation into the corresponding physiological relevance in a multicellular context and in different mechanical environments. Using the Arabidopsis root as a model system, and combining morphometry and micro-rheometry, we found that hyperosmotic stress decreases nuclear circularity and size and increases nuclear stiffness in meristematic cells. These changes were accompanied by enhanced expression of touch response genes. The nuclear response to hyperosmotic stress was rescued upon return to iso-osmotic conditions and could even lead to opposite trends upon hypo-osmotic stress. Interestingly, nuclei in a mutant impaired in the functions of the gamma-tubulin complex protein 3 (GCP3) interacting protein (GIP)/MZT1 proteins at the nuclear envelope were almost insensitive to such osmotic changes. The gip1gip2 mutant exhibited constitutive hyperosmotic stress response with stiffer and deformed nuclei, as well as touch response gene induction. The mutant was also resistant to lethal hyperosmoticconditions. Altogether, we unravel a stereotypical geometric, mechanical, and genetic nuclear response to hyperosmotic stress in plants. Our data also suggest that chromatin acts as a gel that stiffens in hyperosmotic conditions and that the nuclear-envelope-associated protein GIPs act as negative regulators of this response.

Two Roles for the Tenebrio molitor Relish in the Regulation of Antimicrobial Peptides and Autophagy-Related Genes in Response to Listeria monocytogenes.

Relish is a key NF-κB transcription factor of the immune-deficiency (Imd) pathway that combats infection by regulating antimicrobial peptides (AMPs). Understanding of the fundamental role of Tenebrio molitor Relish (TmRelish) in controlling of Listeria monocytogenes virulence through the regulation of both AMPs and autophagy-related (ATG) genes is unclear. Here, we show that TmRelish transcripts were highly abundant in the larval fat body and hemocytes compared to the gut upon L. monocytogenes infection. Furthermore, significant mortality was observed in TmRelish-silenced larvae after intracellular insult. To investigate the cause of this lethality, we measured the induction of AMPs and ATG genes in the TmRelish dsRNA-treated T. molitor larvae. The expression of TmTenecin-1, TmTenecin-4, TmColeptericin-1, TmAttacin-2, and TmCecropin-2 were suppressed in the fat body and hemocytes of dsTmRelish-injected larvae during L. monocytogenes infection. In addition, TmRelish knockdown led to a noticeable downregulation of TmATG1 (a serine-threonine protein kinase) in the fat body and hemocytes of young larvae 6 h post-infection (pi). The notable increase of autophagy genes in the early stage of infection (6 h pi), suggesting autophagy response is crucial for Listeria clearance. Taken together, these results suggest that TmRelish plays pivotal roles in not only regulation of AMP genes but also induction of autophagy genes in response to L. monocytogenes challenge in fat body and hemocytes of T. molitor larvae. Furthermore, negative regulation of several AMPs by TmRelish in the fat body, hemocytes, and gut leaves open the possibility of a crosstalk between Toll and Imd pathway.

Induction of AP-1 by YAP/TAZ contributes to cell proliferation and organ growth.

Yes-associated protein (YAP) and its homolog transcriptional coactivator with PDZ-binding motif (TAZ) are key effectors of the Hippo pathway to control cell growth and organ size, of which dysregulation yields to tumorigenesis or hypertrophy. Upon activation, YAP/TAZ translocate into the nucleus and bind to TEAD transcription factors to promote transcriptional programs for proliferation or cell specification. Immediate early genes, represented by AP-1 complex, are rapidly induced and control later-phase transcriptional program to play key roles in tumorigenesis and organ maintenance. Here, we report that YAP/TAZ directly promote FOS transcription that in turn contributes to the biological function of YAP/TAZ. YAP/TAZ bind to the promoter region of FOS to stimulate its transcription. Deletion of YAP/TAZ blocks the induction of immediate early genes in response to mitogenic stimuli. FOS induction contributes to expression of YAP/TAZ downstream target genes. Genetic deletion or chemical inhibition of AP-1 suppresses growth of YAP-driven cancer cells, such as Lats1/2-deficient cancer cells as well as Gαq/11 mutated uveal melanoma. Furthermore, AP-1 inhibition almost completely abrogates the hepatomegaly induced by YAP overexpression. Our findings reveal a feed-forward interplay between immediate early transcription of AP-1 and Hippo pathway function.

The ALK-1/SMAD/ATOH8 axis attenuates hypoxic responses and protects against the development of pulmonary arterial hypertension.

Dysregulated bone morphogenetic protein (BMP) signaling in endothelial cells (ECs) is implicated in vascular diseases such as pulmonary arterial hypertension (PAH). Here, we showed that the transcription factor ATOH8 was a direct target of SMAD1/5 and was induced in a manner dependent on BMP but independent of Notch, another critical signaling pathway in ECs. In zebrafish and mice, inactivation of Atoh8 did not cause an arteriovenous malformation-like phenotype, which may arise because of dysregulated Notch signaling. In contrast, Atoh8-deficient mice exhibited a phenotype mimicking PAH, which included increased pulmonary arterial pressure and right ventricular hypertrophy. Moreover, ATOH8 expression was decreased in PAH patient lungs. We showed that in cells, ATOH8 interacted with hypoxia-inducible factor 2α (HIF-2α) and decreased its abundance, leading to reduced induction of HIF-2α target genes in response to hypoxia. Together, these findings suggest that the BMP receptor type II/ALK-1/SMAD/ATOH8 axis may attenuate hypoxic responses in ECs in the pulmonary circulation and may help prevent the development of PAH.

Epigenetic Regulation of Vascular Smooth Muscle Cells by Histone H3 Lysine 9 Dimethylation Attenuates Target Gene-Induction by Inflammatory Signaling.

Vascular inflammation underlies cardiovascular disease. Vascular smooth muscle cells (VSMCs) upregulate selective genes, including MMPs (matrix metalloproteinases) and proinflammatory cytokines upon local inflammation, which directly contribute to vascular disease and adverse clinical outcome. Identification of factors controlling VSMC responses to inflammation is therefore of considerable therapeutic importance. Here, we determine the role of Histone H3 lysine 9 di-methylation (H3K9me2), a repressive epigenetic mark that is reduced in atherosclerotic lesions, in regulating the VSMC inflammatory response. Approach and Results: We used VSMC-lineage tracing to reveal reduced H3K9me2 levels in VSMCs of arteries after injury and in atherosclerotic lesions compared with control vessels. Intriguingly, chromatin immunoprecipitation showed H3K9me2 enrichment at a subset of inflammation-responsive gene promoters, including MMP3, MMP9, MMP12, and IL6, in mouse and human VSMCs. Inhibition of G9A/GLP (G9A-like protein), the primary enzymes responsible for H3K9me2, significantly potentiated inflammation-induced gene induction in vitro and in vivo without altering NFκB (nuclear factor kappa-light-chain-enhancer of activated B cell) and MAPK (mitogen-activated protein kinase) signaling. Rather, reduced G9A/GLP activity enhanced inflammation-induced binding of transcription factors NFκB-p65 and cJUN to H3K9me2 target gene promoters MMP3 and IL6. Taken together, these results suggest that promoter-associated H3K9me2 directly attenuates the induction of target genes in response to inflammation in human VSMCs. This study implicates H3K9me2 in regulating the proinflammatory VSMC phenotype. Our findings suggest that reduced H3K9me2 in disease enhance binding of NFκB and AP-1 (activator protein-1) transcription factors at specific inflammation-responsive genes to augment proinflammatory stimuli in VSMC. Therefore, H3K9me2-regulation could be targeted clinically to limit expression of MMPs and IL6, which are induced in vascular disease.

Nutrient deprivation and lysosomal stress induce activation of TFEB in retinal pigment epithelial cells

BackgroundInduction of lysosomal function and autophagy is regarded as an adaptive mechanism in response to cellular stress. The transcription factor EB (TFEB) has been identified as a master regulator of lysosomal function and autophagy. TFEB is a member of the microphthalmia family of bHLH-LZ transcription factors that includes other members such as micropthalmia-associated transcription factor (MITF), TFE3, and TFEC. TFEB controls lysosome biogenesis and autophagy by upregulation of a family of genes belonging to the Coordinated Lysosomal Expression and Regulation (CLEAR) network. Here, we investigated the expression of TFEB in cells subjected to nutrient deprivation and lysosomal stress. We studied transcriptional induction of TFEB-regulated genes in response to nutrient deprivation and lysosomal stress in retinal pigment epithelial (RPE) cells. Furthermore, we also investigated the induction of autophagy and lysosomal genes upon overexpression of constitutively active form of TFEB.MethodsExpression of TFEB and MITF protein levels were evaluated in cells subjected to prolonged periods of nutrient deprivation. mRNA levels of the CLEAR network genes was measured by quantitative real time PCR (qRT-PCR) analysis in cells deprived of nutrients, treated with ammonium chloride and upon overexpression of constitutively active TFEB. Immunostaining with LC3 antibody was used to measure autophagy flux. Labeling with lysoTracker dye was used to assess lysosomes.ResultsOur results show that nutrient deprivation increases protein levels of TFEB and MITF in ARPE-19 cells. Nutrient stress induces the expression of lysosomal (LAMP1, CTSD MCOLN1, SGSH) and autophagy (BECN1) genes. Lysosomal stress also increases the expression of lysosomal (ATP6V0A1 and LAMP1) and autophagy (p62 and BECN1) genes. Our results show that overexpression of constitutively active TFEB also induces the expression of CLEAR network genes.ConclusionsCollectively, these observations suggest that nutrient stress induces the protein expression of both MITF and TFEB in ARPE-19 cells. TFEB-regulated transcriptional program plays an important role in adaptive response of cells during both nutrient and lysosomal stress.

Low ambient humidity impairs barrier function and innate resistance against influenza infection

In the temperate regions, seasonal influenza virus outbreaks correlate closely with decreases in humidity. While low ambient humidity is known to enhance viral transmission, its impact on host response to influenza virus infection and disease outcome remains unclear. Here, we showed that housing Mx1 congenic mice in low relative humidity makes mice more susceptible to severe disease following respiratory challenge with influenza A virus. We find that inhalation of dry air impairs mucociliary clearance, innate antiviral defense, and tissue repair. Moreover, disease exacerbated by low relative humidity was ameliorated in caspase-1/11-deficient Mx1 mice, independent of viral burden. Single-cell RNA sequencing revealed that induction of IFN-stimulated genes in response to viral infection was diminished in multiple cell types in the lung of mice housed in low humidity condition. These results indicate that exposure to dry air impairs host defense against influenza infection, reduces tissue repair, and inflicts caspase-dependent disease pathology.

SAT-204 Silencing of Maternal Hepatic Glucocorticoid Receptor Is Essential for Normal Fetal Development

Excessive or chronic stress can lead to a variety of diseases due to aberrant activation of the glucocorticoid receptor (GR), a ligand activated transcription factor. Pregnancy represents a particular window of sensitivity in which excessive stress can have adverse outcomes, particularly on the developing fetus. Models of maternal stress have historically relied on the use of psychosocial stressors or chronic administration of glucocorticoids leading to systemic increases in glucocorticoids, making it near impossible to understand the tissue-specific effects of stress during pregnancy in contributing to maternal and fetal diseases. We have previously demonstrated an antagonistic relationship between reproductive axis activity and hepatic glucocorticoid sensitivity in a mouse model of menopause. However, if stress/sex hormone crosstalk exists in the liver during pregnancy, and if so, what the physiological purpose it serves is completely unknown. We have discovered hepatic stress hormone responsiveness is diminished during pregnancy. Utilizing whole transcriptome array we profiled the maternal hepatic transcriptome and found a substantial overlap between the pregnancy-regulated transcriptome and glucocorticoid regulated transcriptome. Of the overlapping genes and pathways, we observed the direction of regulation to be largely opposite between the pregnancy-regulated transcriptome and stress hormone regulated transcriptome. Furthermore, we observed reduced basal expression of classical GR target genes in the pregnant liver and impaired induction of GR target genes in response to dexamethasone treatment in the liver of pregnant versus virgin mice. The reduced glucocorticoid sensitivity in the liver during pregnancy is due to impaired expression of GR, which is mediated via epigenetic silencing. Provocatively, we found reinstallation of GR to hepatocytes during pregnancy via adeno-associated viral transduction dysregulates genes involved in proliferation, ultimately culminating in impaired pregnancy-induced hepatomegaly. Furthermore. disruption of the maternal hepatic adaptation to pregnancy by GR reinstallation results in impaired in utero growth of the fetus. These data demonstrate the tissue-specific effects of stress hormone signaling in the maternal compartment is regulated in a dynamic fashion to ultimately support the healthy development of embryos.