3-Methylcatechol mediates anti-fecundity effect by inhibiting estrogen-related receptor-induced glycolytic gene expression in Myzus persicae.

Aphids are a major problem in agriculture, horticulture, and forestry by feeding on leaves and stems, causing discoloration, leaf curling, yellowing, and stunted growth. Although urushiol, a phenolic compound containing a catechol structure, is known for its antioxidant and anticancer properties, using small molecules to control aphids via catechol-mediated mechanisms is poorly understood. In this study, we investigated the effects of 3-methylcatechol (3-MC) on Myzus persicae fecundity. Our results showed that treatment with 3-MC significantly reduced the intrinsic transcriptional activity of the aphid estrogen-related receptor (MpERR), which regulates the expression of glycolytic genes. Additionally, 3-MC treatment suppressed the promoter activity of MpERR-induced rate-limiting enzymes in glycolysis, such as phosphofructokinase and pyruvate kinase, by inhibiting MpERR binding. Finally, 3-MC also suppressed MpERR-induced glycolytic gene expression and reduced the number of offspring produced by viviparous female aphids. Overall, our findings suggest that 3-MC has the potential to be used as a new strategy for managing aphid populations by controlling their offspring production.

Nuclear receptor estrogen-related receptor modulates antimicrobial peptide expression for host innate immunity in Tribolium castaneum.

Antimicrobial peptides (AMPs) are core components of innate immunity to protect insects against microbial infections. Nuclear receptors (NRs) are ligand-dependent transcription factors that can regulate the expression of genes critical for insect development including molting and metamorphosis. However, the role of NRs in host innate immune response to microbial infection remains poorly understood in Tribolium castaneum (T. castaneum). Here, we show that estrogen-related receptor (ERR), an insect ortholog of the mammalian ERR family of NRs, is a novel transcriptional regulator of AMP genes for innate immune response of T. castaneum. Tribolium ERR (TcERR) expression was induced by immune deficiency (IMD)-Relish signaling in response to infection by Escherichia coli (E. coli), a Gram-negative bacterium, as demonstrated in TcIMD-deficient beetles. Interestingly, genome-wide transcriptome analysis of TcERR-deficient old larvae using RNA-sequencing analysis showed that TcERR expression was positively correlated with gene transcription levels of AMPs including attacins, defensins, and coleoptericin. Moreover, chromatin immunoprecipitation analysis revealed that TcERR could directly bind to ERR-response elements on promoters of genes encoding defensin3 and coleoptericin, critical for innate immune response of T. castaneum. Finally, TcERR-deficient old larvae infected with E. coli displayed enhanced bacterial load and significantly less host survival. These findings suggest that TcERR can coordinate transcriptional regulation of AMPs and host innate immune response to bacterial infection.

Nuclear receptor HR3 mediates transcriptional regulation of chitin metabolic genes during molting in Tribolium castaneum.

BACKGROUND: Chitin, a major component of insect cuticles, plays a critical role in insect molting and morphogenesis. Thus, coordination of chitin remodeling during insect development requires tight transcriptional control of the chitin metabolism genes involved in chitin synthesis, assembly and degradation. However, the molecular mechanism underlying transcriptional coordination of chitin metabolism genes during beetle development is not yet completely understood. RESULTS: We cloned the full-length cDNA encoding hormone receptor 3 (TcHR3) from Tribolium castaneum and showed a critical role of TcHR3 in modulating chitin metabolism gene expression during molting. Genome-wide transcriptome analysis of HR3-deficient old larvae using RNA sequencing analysis revealed a positive correlation between TcHR3 and transcription of chitin metabolism genes involved in chitin synthesis and degradation. In addition, HR3 overexpression significantly induced the gene promoter activity of N-acetylglucosaminidase 1 (NAG1) involved in chitin degradation and UDP-N-acetylglucosamine pyrophosphorylase 1 (UAP1) involved in chitin synthesis. Chromatin immunoprecipitation analysis revealed that HR3 could directly bind to HR3-response element of NAG1 and UAP1 promoters. Finally, HR3-deficient late instar larvae and prepupae exhibited defects in larval-larval and larval-pupal molting, respectively, leading to eventual larval death because developing larvae were trapped inside the old cuticle as a result of abnormal chitin metabolism. CONCLUSION: TcHR3 is a transcriptional regulator of chitin metabolic genes for molting of T. castaneum. Controlling the molting system by TcHR3 might be a new management strategy for selective control of red flour beetle infestation. © 2022 Society of Chemical Industry.

Melatonin Regulates Iron Homeostasis by Inducing Hepcidin Expression in Hepatocytes.

The pineal hormone, melatonin, plays important roles in circadian rhythms and energy metabolism. The hepatic peptide hormone, hepcidin, regulates iron homeostasis by triggering the degradation of ferroportin (FPN), the protein that transfers cellular iron to the blood. However, the role of melatonin in the transcriptional regulation of hepcidin is largely unknown. Here, we showed that melatonin upregulates hepcidin gene expression by enhancing the melatonin receptor 1 (MT1)-mediated c-Jun N-terminal kinase (JNK) activation in hepatocytes. Interestingly, hepcidin gene expression was increased during the dark cycle in the liver of mice, whereas serum iron levels decreased following hepcidin expression. In addition, melatonin significantly induced hepcidin gene expression and secretion, as well as the subsequent FPN degradation in hepatocytes, which resulted in cellular iron accumulation. Melatonin-induced hepcidin expression was significantly decreased by the melatonin receptor antagonist, luzindole, and by the knockdown of MT1. Moreover, melatonin activated JNK signaling and upregulated hepcidin expression, both of which were significantly decreased by SP600125, a specific JNK inhibitor. Chromatin immunoprecipitation analysis showed that luzindole significantly blocked melatonin-induced c-Jun binding to the hepcidin promoter. Finally, melatonin induced hepcidin expression and secretion by activating the JNK-c-Jun pathway in mice, which were reversed by the luzindole treatment. These findings reveal a previously unrecognized role of melatonin in the circadian regulation of hepcidin expression and iron homeostasis.

Nuclear receptor HR96 up-regulates cytochrome P450 for insecticide detoxification in Tribolium castaneum.

BACKGROUND: Red flour beetle, Tribolium castaneum (T. castaneum), is a major agricultural pest that causes significant damage to stored grains and products. Although hormone receptor 96 (HR96) is known to be the single ortholog corresponding to mammalian constitutive androstane receptor and pregnane X receptor, the structural features of Tribolium HR96 (TcHR96) and its role in insecticide-mediated transcription control of cytochrome P450 enzyme genes in T. castaneum have not been elucidated yet. RESULTS: We cloned full-length complementary DNA encoding TcHR96 and revealed the role of TcHR96 in transcriptional control of cytochrome P450 enzyme genes. Interestingly, genome-wide transcriptome analysis of HR96-deficient beetles using RNA sequencing showed a positive correlation between TcHR96 and gene transcription of metabolizing enzymes involved in phase I detoxification processes. Moreover, TcHR96 overexpression significantly increased the promoter activity of genes encoding phase I P450 enzymes such as CYP4Q4, CYP4G7, CYP4BR3, and CYP345A1. Chromatin immunoprecipitation analysis showed that TcHR96 could directly bind to the promoter of gene encoding CYP345A1, an enzyme for metabolizing insecticides in T. castaneum. Furthermore, imidacloprid, a neonicotinoid insecticide, significantly increased gene expression of phase I P450 enzymes in old larvae of T. castaneum, which were reversed by TcHR96 knockdown. Finally, TcHR96 knockdown significantly decreased the resistance of old larvae to imidacloprid concomitant with reduction of imidacloprid-mediated phase I P450 enzyme gene expression. CONCLUSION: TcHR96 plays a major role in transcriptional control of P450 enzyme for imidacloprid detoxification. Controlling TcHR96 might facilitate the regulation of insecticide tolerance in T. castaneum, thus providing a promising new strategy to manage pest beetle populations. © 2021 Society of Chemical Industry.

Epigallocatechin-3-Gallate Suppresses BMP-6-Mediated SMAD1/5/8 Transactivation of Hepcidin Gene by Inducing SMILE in Hepatocytes.

Hepcidin, a major regulator of systemic iron homeostasis, is mainly induced in hepatocytes by activating bone morphogenetic protein 6 (BMP-6) signaling in response to changes in the iron status. Small heterodimer partner-interacting leucine zipper protein (SMILE), a polyphenol-inducible transcriptional co-repressor, regulates hepatic gluconeogenesis and lipogenesis. Here, we examine the epigallocatechin-3-gallate (EGCG) effect on BMP-6-mediated SMAD1/5/8 transactivation of the hepcidin gene. EGCG treatment significantly decreased BMP-6-induced hepcidin gene expression and secretion in hepatocytes, which, in turn, abated ferroportin degradation. SMILE overexpression significantly decreased BMP receptor-induced hepcidin promoter activity. SMILE overexpression also significantly suppressed BMP-6-mediated induction of hepcidin mRNA and its secretion in HepG2 and AML12 cells. EGCG treatment inhibited BMP-6-mediated hepcidin gene expression and secretion, which were significantly reversed by SMILE knockdown in hepatocytes. Interestingly, SMILE physically interacted with SMAD1 in the nucleus and significantly blocked DNA binding of the SMAD complex to the BMP-response element on the hepcidin gene promoter. Taken together, these findings suggest that SMILE is a novel transcriptional repressor of BMP-6-mediated hepcidin gene expression, thus contributing to the control of iron homeostasis.

Orphan Nuclear Receptor ERRγ Is a Transcriptional Regulator of CB1 Receptor-Mediated TFR2 Gene Expression in Hepatocytes.

Orphan nuclear receptor estrogen-related receptor γ (ERRγ) is an important transcription factor modulating gene transcription involved in endocrine control of liver metabolism. Transferrin receptor 2 (TFR2), a carrier protein for transferrin, is involved in hepatic iron overload in alcoholic liver disease (ALD). However, TFR2 gene transcriptional regulation in hepatocytes remains largely unknown. In this study, we described a detailed molecular mechanism of hepatic TFR2 gene expression involving ERRγ in response to an endocannabinoid 2-arachidonoylglycerol (2-AG). Treatment with 2-AG and arachidonyl-2'-chloroethylamide, a selective cannabinoid receptor type 1 (CB1) receptor agonist, increased ERRγ and TFR2 expression in hepatocytes. Overexpression of ERRγ was sufficient to induce TFR2 expression in both human and mouse hepatocytes. In addition, ERRγ knockdown significantly decreased 2-AG or alcohol-mediated TFR2 gene expression in cultured hepatocytes and mouse livers. Finally, deletion and mutation analysis of the TFR2 gene promoter demonstrated that ERRγ directly modulated TFR2 gene transcription via binding to an ERR-response element. This was further confirmed by chromatin immunoprecipitation assay. Taken together, these results reveal a previously unrecognized role of ERRγ in the transcriptional regulation of TFR2 gene expression in response to alcohol.

Aphid estrogen-related receptor controls glycolytic gene expression and fecundity.

Aphids, the major insect pests of agricultural crops, reproduce sexually and asexually depending upon environmental factors such as the photoperiod and temperature. Nuclear receptors, a unique family of ligand-dependent transcription factors, control insect development and growth including morphogenesis, molting, and metamorphosis. However, the structural features and biological functions of the aphid estrogen-related receptor (ERR) are largely unknown. Here, we cloned full-length cDNA encoding the ERR in the green peach aphid, Myzus persicae, (Sulzer) (Hemiptera: Aphididae) (MpERR) and demonstrated that the MpERR modulated glycolytic gene expression and aphid fecundity. The phylogenetic analysis revealed that the MpERR originated in a unique evolutionary lineage distinct from those of hemipteran insects. Moreover, the AF-2 domain of the MpERR conferred nuclear localization and transcriptional activity. The overexpression of the MpERR significantly upregulated the gene expression of rate-limiting enzymes involved in glycolysis such as phosphofructokinase and pyruvate kinase by directly binding to ERR-response elements in their promoters. Moreover, ERR-deficient viviparous female aphids showed decreased glycolytic gene expression and produced fewer offspring. These results suggest that the aphid ERR plays a pivotal role in glycolytic transcriptional control and fecundity.

Iron-Oxide Nanocluster Labeling of Clostridium novyi-NT Spores for MR Imaging-Monitored Locoregional Delivery to Liver Tumors in Rat and Rabbit Models.

PURPOSE: To label Clostridium novyi-NT spores (C. novyi-NT) with iron oxide nanoclusters and track distribution of bacteria during magnetic resonance (MR) imaging-monitored locoregional delivery to liver tumors using intratumoral injection or intra-arterial transcatheter infusion. MATERIALS AND METHODS: Vegetative state C. novyi-NT were labeled with iron oxide particles followed by induction of sporulation. Labeling was confirmed with fluorescence microscopy and transmission electron microscopy (TEM). T2 and T2* relaxation times for magnetic clusters and magnetic microspheres were determined using 7T and 1.5T MR imaging scanners. In vitro assays compared labeled bacteria viability and oncolytic potential to unlabeled controls. Labeled spores were either directly injected into N1-S1 rodent liver tumors (n = 24) or selectively infused via the hepatic artery in rabbits with VX2 liver tumors (n = 3). Hematoxylin-eosin, Prussian blue, and gram staining were performed. Statistical comparison methods included paired t-test and ANOVA. RESULTS: Both fluorescence microscopy and TEM studies confirmed presence of iron oxide labels within the bacterial spores. Phantom studies demonstrated that the synthesized nanoclusters produce R2 relaxivities comparable to clinical agents. Labeling had no significant impact on overall growth or oncolytic properties (P >.05). Tumor signal-to-noise ratio (SNR) decreased significantly following intratumoral injection and intra-arterial infusion of labeled spores (P <.05). Prussian blue and gram staining confirmed spore delivery. CONCLUSIONS: C. novyi-NT spores can be internally labeled with iron oxide nanoparticles to visualize distribution with MR imaging during locoregional bacteriolytic therapy involving direct injection or intra-arterial transcatheter infusion.

Enhanced topical delivery of small hydrophilic or lipophilic active agents and epidermal growth factor by fractional radiofrequency microporation.

PURPOSE: To evaluate the ability of a novel radiofrequency (RF) microporation technology based on ablation of the skin barrier to enhance topical delivery of active ingredients METHODS: The influence of RF fluence and the molecular size of the absorbent on the permeation enhancement was confirmed by in vitro skin permeation study using Franz diffusion cells. The improved skin rejuvenation effects, such as depigmentation and anti-wrinkle effects, by enhanced topical delivery of α-bisabolol and epidermal growth factor (EGF) through the RF microchannels were investigated in photo-damaged skin. RESULTS: The cumulative amounts of active ingredients through the RF microporated skin were significantly increased. Topically applied α-bisabolol after RF microporation induced rapid onset of skin whitening and significantly increased the ΔL-value of UVB-induced hyperpigmented melanin hairless mouse skin. In addition, wrinkle formation after topical application of EGF with RF microporation was significantly reduced and prevented after 12 weeks, and all parameters involving wrinkles in a replica analysis were similar to those in the negative control. CONCLUSIONS: RF microporation enhances the topical delivery of active ingredients with high molecular weight or of small hydrophilic or lipophilic molecules. Thus, this technology can effectively improve photo-induced hyperpigmentation and wrinkle formation by enhancing topical delivery of active agents.

Fabrication of pseudo-ceramide-based lipid microparticles for recovery of skin barrier function.

The recovery of skin barrier functions was investigated with pseudo-ceramide-based lipid microparticles. The microparticles were prepared by using a fluid bed technique where lipid components (a pseudo-ceramide, cholesterol and a fatty acid) were coated on a sugar seed, and a polymer was subsequently coated on the lipid microparticles. The microparticles contained large amount of pseudo-ceramide, and the pseudo-ceramide was in the form of lamellar structures mixed with other lipid components. In addition, the microparticles were stably dispersed in aqueous media or emulsion systems without any disruption of the microparticles' structures, thereby supplying sufficient amount of the pseudo-ceramide to skins for improving skin barrier functions such as preventing water loss. Such a role of the microparticles was proven by evaluating in vivo the efficacy of the lipid microparticles in reducing a trans-epidermal water loss (TEWL) of impaired murine skins. As a result, the novel pseudo-ceramide-based lipid microparticles for barrier recovery may potentially be applied in the field of dermatology, cosmetics and pharmaceuticals.