Up-regulation Of Gene Transcription Research Articles

A dual function of the IDA peptide in regulating cell separation and modulating plant immunity at the molecular level.

The abscission of floral organs and emergence of lateral roots in Arabidopsis is regulated by the peptide ligand inflorescence deficient in abscission (IDA) and the receptor protein kinases HAESA (HAE) and HAESA-like 2 (HSL2). During these cell separation processes, the plant induces defense-associated genes to protect against pathogen invasion. However, the molecular coordination between abscission and immunity has not been thoroughly explored. Here, we show that IDA induces a release of cytosolic calcium ions (Ca2+) and apoplastic production of reactive oxygen species, which are signatures of early defense responses. In addition, we find that IDA promotes late defense responses by the transcriptional upregulation of genes known to be involved in immunity. When comparing the IDA induced early immune responses to known immune responses, such as those elicited by flagellin22 treatment, we observe both similarities and differences. We propose a molecular mechanism by which IDA promotes signatures of an immune response in cells destined for separation to guard them from pathogen attack.

A dual function of the IDA peptide in regulating cell separation and modulating plant immunity at the molecular level

The abscission of floral organs and emergence of lateral roots in Arabidopsis is regulated by the peptide ligand inflorescence deficient in abscission (IDA) and the receptor protein kinases HAESA (HAE) and HAESA-like 2 (HSL2). During these cell separation processes, the plant induces defense-associated genes to protect against pathogen invasion. However, the molecular coordination between abscission and immunity has not been thoroughly explored. Here, we show that IDA induces a release of cytosolic calcium ions (Ca2+) and apoplastic production of reactive oxygen species, which are signatures of early defense responses. In addition, we find that IDA promotes late defense responses by the transcriptional upregulation of genes known to be involved in immunity. When comparing the IDA induced early immune responses to known immune responses, such as those elicited by flagellin22 treatment, we observe both similarities and differences. We propose a molecular mechanism by which IDA promotes signatures of an immune response in cells destined for separation to guard them from pathogen attack.

Evaluation of Cellular Responses of Heterotrophic Escherichia coli Cultured with Autotrophic Chlamydomonas reinhardtii as a Nutrient Source by Analyses Based on Microbiology and Transcriptome.

Heterotrophic microorganism Escherichia coli LS5218 was cultured with flesh green alga Chlamydomonas reinhardtii C-9: NIES-2235 as a nutrient supplier. In order to evaluate the cell response of Escherichia coli with Chlamydomonas reinhardtii, Escherichia coli was evaluated with microbial methods and comprehensive gene transcriptional analyses. Escherichia coli with Chlamydomonas reinhardtii showed a specific growth rate (µmax) of 1.04 ± 0.27, which was similar to that for cells growing in Luria-Bertani medium (µmax = 1.20 ± 0.40 h-1). Furthermore, comparing the cellular responses of Escherichia coli in a green-algae-containing medium with those in the Luria-Bertani medium, transcriptomic analysis showed that Escherichia coli upregulated gene transcription levels related to glycolysis, 5-phospho-d-ribosyl-1-diphosphate, and lipid synthesis; on the other hand, it decreased the levels related to lipid degradation. In particular, the transcription levels were increased by 103.7 times on pgm (p * < 0.05 (p = 0.015)) in glycolysis, and decreased by 0.247 times on fadE (p * < 0.05 (p = 0.041)) in lipolysis. These genes are unique and could regulate the direction of metabolism; these responses possibly indicate carbon source assimilation as a cellular response in Escherichia coli. This paper is the first report to clarify that Escherichia coli, a substance-producing strain, directly uses Chlamydomonas reinhardtii as a nutrient supplier by evaluation of the cellular responses analyzed with microbial methods and transcriptome analysis.

Distinct microbial characteristics of the robust single-stage coupling system during the conversion from anammox-denitritation to anammox-denitratation patterns

Simultaneous anammox-denitrification is effectively operated in two types, i.e., the anammox-denitritation (SAD pattern) and the anammox-denitratation (PDA pattern). The nitrate derived from inevitable nitrite oxidization likely determines the practical operational pattern of the coupling system, while little information is available regarding the microbial characteristics during the pattern conversion. Here, the single-stage bioreactor coupling anammox with denitrification was operated under conditions with a changed ratio of influent nitrite and nitrate. Results showed that the bioreactor exhibited a robust performance during the conversion from SAD to PDA patterns, corresponding with the total nitrogen removal efficiency ranging from 89.5% to 92.4%. Distinct community structures were observed in two patterns, while functional bacteria including the genera Denitratisoma, Thauera, Candidatus Brocadia, and Ca. Jettenia steadily co-existed. Meanwhile, the high transcription of hydrazine synthase genes demonstrated a stable anammox process, while the up-regulated transcription of nitrite and nitrous oxide reductase genes indicated that the complete denitrification process was enhanced for total nitrogen removal during the PDA pattern. Ecologically, stochastic processes dominantly governed the community assembly in two patterns. The PDA pattern improved the interconnectivity of communities, especially for the cooperative behaviors between dominant denitrifying bacteria and low-abundant species. These findings deepen our understanding of the microbial mechanism underlying the different patterns of the coupling system and potentially expand its engineering application.

AGING MODULATION WITH EXERCISE AND MIMETICS

Abstract Understanding the molecular and cellular processes of the Longevity Pathway AMPK-SIRT1, and its activation by exercise and mimetics, plays a vital role in a series of molecular and cellular processes which stimulates optimal healthy cellular efficiency and functioning, ending with improved exercise tolerance. Knowledge of the role of exercise induced activation of the AMPK-SIRT1 pathway, and altering it towards dysfunctional processes and drivers of aging and disease propagation is important to understand to also understand. Cellular dysfunction unfortunately is the norm. This leads to molecular increase of P53 in a binding state not of DNA repair, but upregulation of gene transcription of NF-kB, which increases inflammatory cytokines, over activation of MTOR, and cellular senescence with secretion of the SASP. All these dysfunctions create feedback loops, which makes the process self propagating, resulting in progressive aging, and disease states. The most optimal state is regulation of the optimal molecular and cellular environment, via exercise or a mimetic stimulator of AMPK-SIRT1. Knowledge of the optimal body state for health, and enhancing this AMPK-SIRT process, is low body fat releasing beneficial adiponectin, and no inflammatory cytokines or adipokines. Training muscle releases positive myokines. Various mimetics can stimulate AMPK-SIRT1, such as Metformin, the mitochondrial derived peptide MOTS-C, AICAR, and other compounds intended for people with medical issues that prevents exercise. Some display a synergistic effect with exercise. Understanding, the benefits of exercise and modulation of aging, by encouraging healthy processes, and reducing the deleterious patterns is important to understand.

THU117 Lipocalin-2 As A Mediator Of Negative Affect After Sepsis

Abstract Disclosure: S.E. Yang: None. C. Johnston: None. K. Laborc: None. S. Kounelis-Wuillaume: None. W. Schwieterman: None. A.R. Hill: None. J.L. Spencer-Segal: None. Sepsis is characterized by a life-threatening response to severe infection including systemic inflammation. Many sepsis survivors suffer long-term from a post-sepsis syndrome including prominent neuropsychiatric symptoms, but the mechanisms underlying these symptoms remain unknown. In mice, cecal ligation and puncture (CLP) is used to induce sepsis, which is then treated with antibiotics and fluids. Although mice successfully recover within a week, we observe long-lasting anxiety-like behaviors in C57BL/6J mice of both sexes up to 3 weeks post-CLP as shown by decreased time spent in the open areas of an elevated zero maze compared to SHAM mice (52.24±24.60 vs. 74.32±20.25 seconds; p&amp;lt;0.05; n=12 CLP, n=18 SHAM), reminiscent of the symptoms seen in human sepsis survivors. To elucidate the mechanisms underlying anxiety-like behavior in CLP survivors, we performed bulk RNA-sequencing in hippocampal tissues 3 weeks after SHAM or CLP surgeries. We found that lipocalin-2 (LCN2), a key component of the innate immune response, was one of the most upregulated gene transcripts in the hippocampus after CLP (24.3-fold increase relative to SHAM). We also observed a significant increase in systemic LCN2 protein levels in plasma samples 3 weeks after CLP compared to SHAM (1246±1902.62 vs. 77.53±59.09 ng/mL; p&amp;lt;0.001; n=30 CLP, n=39 SHAM). To identify the cellular source of lcn2 mRNA upregulation, we performed RNAscope in hippocampal tissues and found that lcn2 mRNA expression is similar between SHAM and CLP mice in CA1 hippocampal neurons, and highly abundant in TEK+ endothelial cells after CLP (0.43±0.18 vs. 0.25±0.05 relative optical density; p&amp;lt;0.05; n=8 CLP, n=20 SHAM) suggesting that endothelial cells are the major source of LCN2 upregulation after CLP. To investigate the role of LCN2 as a hormonal mediator of negative affect in sepsis, we administered recombinant LCN2 (rLCN2) once a day for 7 days via subcutaneous injections in both male and female mice and assessed affect and memory using the open field test, the elevated zero maze and the novel objection recognition test. While vehicle and rLCN2-injected female mice showed no significant differences in behavior, rLCN2-injected male mice exhibited anxiety-like behaviors as indicated by a significant decrease in the time spent in the center of an open field compared to vehicle-injected controls (48.25±18.62 vs. 65.87±14.76 seconds; p&amp;lt;0.05; n=9 per group). This difference was not caused by locomotion deficits as the mean total distance travelled was similar between groups. Overall, our results demonstrate that LCN2 is a hormone produced in excess by endothelial cells in sepsis survivors, and that chronic rLCN2 administration was sufficient to induce anxiety-like behaviors in male mice but not female mice. Taken together, our data suggest that LCN2 is a novel endocrine mediator of post-sepsis syndrome. Presentation: Thursday, June 15, 2023

Neuroprotection by upregulation of the major histocompatibility complex class I (MHC I) in SOD1G93A mice.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that progressively affects motoneurons, causing muscle atrophy and evolving to death. Astrocytes inhibit the expression of MHC-I by neurons, contributing to a degenerative outcome. The present study verified the influence of interferon β (IFN β) treatment, a proinflammatory cytokine that upregulates MHC-I expression, in SOD1G93A transgenic mice. For that, 17 days old presymptomatic female mice were subjected to subcutaneous application of IFN β (250, 1,000, and 10,000 IU) every other day for 20 days. Rotarod motor test, clinical score, and body weight assessment were conducted every third day throughout the treatment period. No significant intergroup variations were observed in such parameters during the pre-symptomatic phase. All mice were then euthanized, and the spinal cords collected for comparative analysis of motoneuron survival, reactive gliosis, synapse coverage, microglia morphology classification, cytokine analysis by flow cytometry, and RT-qPCR quantification of gene transcripts. Additionally, mice underwent Rotarod motor assessment, weight monitoring, and neurological scoring. The results show that IFN β treatment led to an increase in the expression of MHC-I, which, even at the lowest dose (250 IU), resulted in a significant increase in neuronal survival in the ALS presymptomatic period which lasted until the onset of the disease. The treatment also influenced synaptic preservation by decreasing excitatory inputs and upregulating the expression of AMPA receptors by astrocytes. Microglial reactivity quantified by the integrated density of pixels did not decrease with treatment but showed a less activated morphology, coupled with polarization to an M1 profile. Disease progression upregulated gene transcripts for pro- and anti-inflammatory cytokines, and IFN β treatment significantly decreased mRNA expression for IL4. Overall, the present results demonstrate that a low dosage of IFN β shows therapeutic potential by increasing MHC-I expression, resulting in neuroprotection and immunomodulation.

Combined impacts of microplastics and cadmium on the liver function, immune response, and intestinal microbiota of crucian carp (Carassius carassius)

Microplastics (MPs) and the heavy metal cadmium (Cd) have attracted global attention for their toxicological interactions in aquatic organisms. The purpose of this investigation was evaluating the effect of MPs (1 mg L−1) and Cd (5 mg L−1) on the liver function, immune response of crucian carp (Carassius carassius) after 96 h exposure, and intestinal microbiota after 21 days, respectively. Co-exposure to MPs and Cd significantly enhanced MP accumulation in the liver of the crucian carp compared to the accumulation with exposure to MPs alone. Co-exposure to MPs and Cd triggered notable histopathological alterations accompanied by increased hepatic cell necrosis and inflammation, and was associated with higher aspartate aminotransferase and alanine aminotransferase levels, lower superoxide dismutase and catalase activity levels, but higher malondialdehyde content and total antioxidant capacity in the liver. Moreover, the combined treatment of MPs and Cd led to the up-regulated transcription of genes related to immune response, such as interleukin 8 (il-8), il-10, il-1β, tumor necrosis factor-α, and heat shock protein 70, both in the liver and spleen. Co-exposure to MPs and Cd reduced the variety and abundance of the intestinal microbiota in the crucian carp. Our research indicates that the combined exposure to MPs and Cd may exert synergistic toxic effects on crucian carp, which could impede the sustainable growth of the aquaculture industry and pose potential risks to food safety.

Transcriptome analysis reveals the essential role of NK2 homeobox 1/thyroid transcription factor 1 (NKX2-1/TTF-1) in gastric adenocarcinoma of fundic-gland type.

Gastric adenocarcinoma of fundic-gland type (GA-FG) is a gastric malignancy with little relation to Helicobacter pylori. Clinical characteristics of GA-FG have been established, but molecular mechanisms leading to tumorigenesis have not yet been elucidated. We subjected three GA-FG tumors-normal mucosa pairs to microarray analysis. Network analysis was performed for the top 30 up-regulated gene transcripts, followed by immunohistochemical staining to confirm the gene expression analysis results. AGS and NUGC4 cells were transfected with the gene-encoding NK2 homeobox 1/thyroid transcription factor 1 (NKX2-1/TTF-1) to evaluate transcriptional changes in its target genes. Comprehensive gene expression analysis identified 1410 up-regulated and 1395 down-regulated gene probes with ≥ two-fold difference in expression. Among the top 30 up-regulated genes in GA-FG, we identified transcription factor NKX2-1/TTF-1, a master regulator of lung/thyroid differentiation, together with surfactant protein B (SFTPB), SFTPC, and secretoglobin family 3A member 2(SCGB3A2), which are regulated by NKX2-1/TTF-1. Immunohistochemical analysis of 16 GA-FG specimens demonstrated significantly higher NKX2-1/TTF-1 and SFTPB levels, as compared to that in adjacent normal mucosa (P < 0.05), while SCGB3A2 levels did not differ (P = 0.341). Transduction of NKX2-1/TTF-1 into AGS and NUGC4 cells induced transactivation of SFTPB and SFTPC, indicating that NKX2-1/TTF-1 can function as normally in gastric cells as it can in the lung cells. Our first transcriptome analysis of GA-FG indicates significant expression of NKX2-1/TTF1 in GA-FG. Immunohistochemistry and cell biology show ectopic expression and normal transactivation ability of NKX2-1/TTF-1, suggesting that it plays an essential role in GA-FG development.

Spatial distribution and time-course of polyphenol accumulation in grape berry (Vitis labruscana cv. ‘Kyoho’)

The spatial distribution and time-course evolution of polyphenol compounds in response to exogenous light and temperature in grape berry (Vitis labruscana cv. ‘Kyoho’) were examined. Results demonstrated that the polyphenol biosynthesis was induced via the up-regulated gene transcription levels during berry ripening. The polyphenol biosynthesis was differentially observed in skins, stems, leaves and seeds, and the more polyphenols were accumulated in seeds; and the polyphenols were significantly distributed in bottom berries of grape clusters. Additionally, the lower temperature (4 ℃) accelerated the polyphenol biosynthesis by 1.3 times after storage of 28 days, whereas the light exposure slightly inhibited the polyphenol biosynthesis of berry during storage at 25 ℃. In conclusion, the data presented in this study shed more light on deeper understanding of the spatial accumulation and time-course evolution of polyphenols in grape berry.

Structure, dynamics, and function of SrnR, a transcription factor for nickel-dependent gene expression.

Streptomyces griseus, a bacterium producing antibacterial drugs and featuring possible application in phytoremediation, expresses two metal-dependent superoxide dismutase (SOD) enzymes, containing either Fe(II) or Ni(II) in their active site. In particular, the alternative expression of the two proteins occurs in a metal-dependent mode, with the Fe(II)-enzyme gene (sodF) repressed at high intracellular Ni(II) concentrations by a two-component system (TCS). This complex involves two proteins, namely SgSrnR and SgSrnQ, which represent the transcriptional regulator and the Ni(II) sensor of the system, respectively. SgSrnR belongs to the ArsR/SmtB family of metal-dependent transcription factors; in the apo-form and in the absence of SgSrnQ, it can bind the DNA operator of sodF, upregulating gene transcription. According to a recently proposed hypothesis, Ni(II) binding to SgSrnQ would promote its interaction with SgSrnR, causing the release of the complex from DNA and the consequent downregulation of the sodF expression. SgSrnQ is predicted to be highly disordered, thus the understanding, at the molecular level, of how the SgSrnR/SgSrnQ TCS specifically responds to Ni(II) requires the knowledge of the structural, dynamic, and functional features of SgSrnR. These were investigated synergistically in this work using X-ray crystallography, nuclear magnetic resonance (NMR) spectroscopy, atomistic molecular dynamics calculations, isothermal titration calorimetry, and in silico molecular docking. The results reveal that the homodimeric apo-SgSrnR binds to its operator in a two-step process that involves the more rigid globular portion of the protein and leaves its largely disordered regions available to possibly interact with the disordered SgSrnQ in a Ni-dependent process.

Protein Elicitor EsxA Induces Resistance to Seedling Blight and PR Genes Differential Transcription in Rice

Protein elicitors can induce plant systemic resistance to pathogens. In an earlier study, we cloned an EsxA gene from the plant growth-promoting rhizobacterium Paenibacillus terrae NK3-4 and expressed it in Pichia pastoris. In addition to being important for the pathogenicity of animal pathogens, EsxA can also induce an immune response in animals. While, we found the exogenously expressed EsxA has the activity of elicitor, which can trigger hypersensitive response and reactive oxygen species burst in leaves as well as enhanced rice plant growth. The effects of EsxA on seedling blight (Fusarium oxysporum) resistance and gene transcription, including pathogenesis-related (PR) genes in rice were evaluated. The germination rate was 95.0% for seeds treated with EsxA and then inoculated with F. oxysporum, which was 2.8-times higher than that of F. oxysporum-infected control seeds that were not treated with EsxA (Con). The buds and roots of EsxA-treated seedlings were 2.4- and 15.9-times longer than those of Con seedlings. The plants and roots of seedlings dipped in an EsxA solution and then inoculated with F. oxysporum were longer than those of the Con seedlings. Theplant length, number of total roots, and number of white roots were respectively 23.2%, 1.74-times, and 7.42-times greater for the seedlings sprayed with EsxA and then inoculated with F. oxysporum than for the Con seedlings. The EsxA induction efficiency (spray treatment) on seedling blight resistance was 60.9%. The transcriptome analysis revealed 1137 and 239 rice genes with EsxA-induced up-regulated and down-regulated transcription levels, respectively. At 48 h after the EsxA treatment, the transcription of 611 and 160 genes was up-regulated and down-regulated, respectively, compared with the transcription levels for the untreated control at the same time-point. Many disease resistance-related PR genes had up-regulated transcription levels. The qPCR data were consistent with the transcriptome sequencing results. EsxA triggered rice ISR to seedling blight and gene differential transcription, including the up-regulated transcription of rice PR genes. These findings may be relevant for the use of EsxA as a protein elicitor to control plant diseases.

RNA Activation-A Novel Approach to Therapeutically Upregulate Gene Transcription.

RNA activation (RNAa) is a mechanism whereby RNA oligos complementary to genomic sequences around the promoter region of genes increase the transcription output of their target gene. Small activating RNA (saRNA) mediate RNAa through interaction with protein co-factors to facilitate RNA polymerase II activity and nucleosome remodeling. As saRNA are small, versatile and safe, they represent a new class of therapeutics that can rescue the downregulation of critical genes in disease settings. This review highlights our current understanding of saRNA biology and describes various examples of how saRNA are successfully used to treat various oncological, neurological and monogenic diseases. MTL-CEBPA, a first-in-class compound that reverses CEBPA downregulation in oncogenic processes using CEBPA-51 saRNA has entered clinical trial for the treatment of hepatocellular carcinoma (HCC). Preclinical models demonstrate that MTL-CEBPA reverses the immunosuppressive effects of myeloid cells and allows for the synergistic enhancement of other anticancer drugs. Encouraging results led to the initiation of a clinical trial combining MTL-CEBPA with a PD-1 inhibitor for treatment of solid tumors.

BMAA, Methylmercury, and Mechanisms of Neurodegeneration in Dolphins: A Natural Model of Toxin Exposure.

Dolphins are well-regarded sentinels for toxin exposure and can bioaccumulate a cyanotoxin called β-N-methylamino-l-alanine (BMAA) that has been linked to human neurodegenerative disease. The same dolphins also possessed hallmarks of Alzheimer’s disease (AD), suggesting a possible association between toxin exposure and neuropathology. However, the mechanisms of neurodegeneration in dolphins and the impact cyanotoxins have on these processes are unknown. Here, we evaluate BMAA exposure by investigating transcription signatures using PCR for dolphin genes homologous to those implicated in AD and related dementias: APP, PSEN1, PSEN2, MAPT, GRN, TARDBP, and C9orf72. Immunohistochemistry and Sevier Münger silver staining were used to validate neuropathology. Methylmercury (MeHg), a synergistic neurotoxicant with BMAA, was also measured using PT-GC-AFS. We report that dolphins have up to a three-fold increase in gene transcription related to Aβ+ plaques, neurofibrillary tangles, neuritic plaques, and TDP-43+ intracytoplasmic inclusions. The upregulation of gene transcription in our dolphin cohort paralleled increasing BMAA concentration. In addition, dolphins with BMAA exposures equivalent to those reported in AD patients displayed up to a 14-fold increase in AD-type neuropathology. MeHg was detected (0.16–0.41 μg/g) and toxicity associated with exposure was also observed in the brain. These results demonstrate that dolphins develop neuropathology associated with AD and exposure to BMAA and MeHg may augment these processes.

CCCH protein-PvCCCH69 acted as a repressor for leaf senescence through suppressing ABA-signaling pathway

CCCH is a subfamily of zinc finger proteins involved in plant growth, development, and stresses response. The function of CCCH in regulating leaf senescence, especially its roles in abscisic acid (ABA)-mediated leaf senescence is largely unknown. The objective of this study was to determine functions and mechanisms of CCCH gene in regulating leaf senescence in switchgrass (Panicum virgatum). A CCCH gene, PvCCCH69 (PvC3H69), was cloned from switchgrass. Overexpressing PvC3H69 in rice suppressed both natural senescence with leaf aging and dark-induced leaf senescence. Endogenous ABA content, ABA biosynthesis genes (NCED3, NCED5, and AAO3), and ABA signaling-related genes (SnRKs, ABI5, and ABF2/3/4) exhibited significantly lower levels in senescencing leaves of PvC3H69-OE plants than those in WT plants. PvC3H69-suppression of leaf senescence was associated with transcriptional upregulation of genes mainly involved in the light-dependent process of photosynthesis, including light-harvesting complex proteins, PSI proteins, and PSII proteins and downregulation of ABA biosynthesis and signaling genes and senescence-associated genes. PvC3H69 could act as a repressor for leaf senescence via upregulating photosynthetic proteins and repressing ABA synthesis and ABA signaling pathways.

An In Vitro Model of Avian Skin Reveals Evolutionarily Conserved Transcriptional Regulation of Epidermal Barrier Formation

The function of the skin as a barrier against a dry environment evolved in a common ancestor of terrestrial vertebrates such as mammals and birds. However, it is unknown which elements of the genetic program of skin barrier formation are evolutionarily ancient and conserved. In this study, we determined the transcriptomes of chicken keratinocytes (KCs) grown in monolayer culture and in an organotypic model of avian skin. The differentiation-associated changes in global gene expression were compared with previously published transcriptome changes of human KCs cultured under equivalent conditions. We found that specific keratins and genes of the epidermal differentiation complex were upregulated during the differentiation of both chicken and human KCs. Likewise, the transcriptional upregulation of genes that control the synthesis and transport of lipids, anti-inflammatory cytokines of the IL-1 family, protease inhibitors, and other regulators of tissue homeostasis was conserved in the KCs of both species. However, some avian KC differentiation-associated transcripts lack homologs in mammals and vice versa, indicating a genetic basis for taxon-specific skin features. The results of this study reveal an evolutionarily ancient program in which dynamic gene transcription controls the metabolism and transport of lipids as well as other core processes during terrestrial skin barrier formation.

Glycogen Synthase Kinase 3 Beta Regulates the Human Aryl Hydrocarbon Receptor Cellular Content and Activity.

The aryl hydrocarbon receptor (AHR) is a cytosolic receptor which is involved in diverse cellular events in humans. The most well-characterized function of AHR is its ability to upregulate gene transcription after exposure to its ligands, such as environmental toxicants, dietary antioxidants, drugs, and endogenous ligands. The cellular content of AHR is partly controlled by its degradation via the ubiquitin–proteasome system and the lysosome-dependent autophagy. We used human cervical cancer (HeLa) cells to investigate how AHR undergoes protein degradation and how its activity is modulated. Since the glycogen synthase kinase 3 beta (GSK3β)-mediated phosphorylation can trigger protein degradation and substrates of GSK3β contain stretches of serine/threonine residues which can be found in AHR, we examined whether degradation and activity of AHR can be controlled by GSK3β. We observed that AHR undergoes the GSK3β-dependent, LC3-mediated lysosomal degradation without ligand treatment. The AHR can be phosphorylated in a GSK3β-dependent manner at three putative sites (S436/S440/S444, S689/S693/T697, and S723/S727/T731), which leads to lysosomal degradation of the AHR protein. Inhibition of the GSK3β activity suppresses the ligand-activated transcription of an AHR target gene in HeLa, human liver cancer (Hep3B), and human breast cancer (MCF-7) cells. Collectively, our findings support that phosphorylation of AHR by GSK3β is essential for the optimal activation of its target gene transcription and this phosphorylation may partake as an “off” switch by subjecting the receptor to lysosomal degradation.

Iron supplementation regulates the progression of high fat diet induced obesity and hepatic steatosis via mitochondrial signaling pathways

Disruption of iron metabolism is closely related to metabolic diseases. Iron deficiency is frequently associated with obesity and hepatic steatosis. However, the effects of iron supplementation on obesity and energy metabolism remain unclear. Here we show that a high-fat diet supplemented with iron reduces body weight gain and hepatic lipid accumulation in mice. Iron supplementation was found to reduce mitochondrial morphological abnormalities and upregulate gene transcription involved in mitochondrial function and beta oxidation in the liver and skeletal muscle. In both these tissues, iron supplementation increased the expression of genes involved in heme or iron–sulfur (Fe–S) cluster synthesis. Heme and Fe–S cluster, which are iron prosthetic groups contained in electron transport chain complex subunits, are essential for mitochondrial respiration. The findings of this study demonstrated that iron regulates mitochondrial signaling pathways—gene transcription of mitochondrial component molecules synthesis and their energy metabolism. Overall, the study elucidates the molecular basis underlying the relationship between iron supplementation and obesity and hepatic steatosis progression, and the role of iron as a signaling molecule.

Extra virgin olive oil improved body weight and insulin sensitivity in high fat diet-induced obese LDLr\u2212/\u2212.Leiden mice without attenuation of steatohepatitis

Dietary fatty acids play a role in the pathogenesis of obesity-associated non-alcoholic fatty liver disease (NAFLD), which is associated with insulin resistance (IR). Fatty acid composition is critical for IR and subsequent NAFLD development. Extra-virgin olive oil (EVOO) is the main source of monounsaturated fatty acids (MUFA) in Mediterranean diets. This study examined whether EVOO-containing high fat diets may prevent diet-induced NAFLD using Ldlr−/−. Leiden mice. In female Ldlr−/−.Leiden mice, the effects of the following high fat diets (HFDs) were examined: a lard-based HFD (HFD-L); an EVOO-based HFD (HFD-EVOO); a phenolic compounds-rich EVOO HFD (HFD-OL). We studied changes in body weight (BW), lipid profile, transaminases, glucose homeostasis, liver pathology and transcriptome. Both EVOO diets reduced body weight (BW) and improved insulin sensitivity. The EVOOs did not improve transaminase values and increased LDL-cholesterol and liver collagen content. EVOOs and HFD-L groups had comparable liver steatosis. The profibrotic effects were substantiated by an up-regulation of gene transcripts related to glutathione metabolism, chemokine signaling and NF-kappa-B activation and down-regulation of genes relevant for fatty acid metabolism. Collectivelly, EVOO intake improved weight gain and insulin sensitivity but not liver inflammation and fibrosis, which was supported by changes in hepatic genes expression.

Unraveling the Molecular Nexus between GPCRs, ERS, and EMT.

G protein-coupled receptors (GPCRs) represent a large family of transmembrane proteins that transduce an external stimulus into a variety of cellular responses. They play a critical role in various pathological conditions in humans, including cancer, by regulating a number of key processes involved in tumor formation and progression. The epithelial-mesenchymal transition (EMT) is a fundamental process in promoting cancer cell invasion and tumor dissemination leading to metastasis, an often intractable state of the disease. Uncontrolled proliferation and persistent metabolism of cancer cells also induce oxidative stress, hypoxia, and depletion of growth factors and nutrients. These disturbances lead to the accumulation of misfolded proteins in the endoplasmic reticulum (ER) and induce a cellular condition called ER stress (ERS) which is counteracted by activation of the unfolded protein response (UPR). Many GPCRs modulate ERS and UPR signaling via ERS sensors, IRE1α, PERK, and ATF6, to support cancer cell survival and inhibit cell death. By regulating downstream signaling pathways such as NF-κB, MAPK/ERK, PI3K/AKT, TGF-β, and Wnt/β-catenin, GPCRs also upregulate mesenchymal transcription factors including Snail, ZEB, and Twist superfamilies which regulate cell polarity, cytoskeleton remodeling, migration, and invasion. Likewise, ERS-induced UPR upregulates gene transcription and expression of proteins related to EMT enhancing tumor aggressiveness. Though GPCRs are attractive therapeutic targets in cancer biology, much less is known about their roles in regulating ERS and EMT. Here, we will discuss the interplay in GPCR-ERS linked to the EMT process of cancer cells, with a particular focus on oncogenes and molecular signaling pathways.