Thyroid Hormone Receptor alpha-1 Directly Controls Transcription of the beta-Catenin Gene in Intestinal Epithelial Cells

Thyroid hormones, T3 and T4, are known regulators of intestine development. The best characterized example is the remodeling of the gastrointestinal tract during amphibian metamorphosis. Thyroid hormones act via nuclear receptors, the TRs, which are T3-dependent transcription factors. We previously showed that intestinal epithelial cell proliferation is controlled by thyroid hormones and the TRalpha gene. To analyze the mechanisms responsible, we studied the expression of genes belonging to and/or activated by the Wnt/beta-catenin pathway, a major actor in the control of physiological and pathological epithelial proliferation in the intestine. We show that T3-TR1 controls the transcription of the beta-catenin gene in an epithelial cell-autonomous way. This is parallel to positive regulation of proliferation-controlling genes such as type D cyclins and c-myc, known targets of the Wnt/-beta-catenin. In addition, we show that the regulation of the beta-catenin gene is direct, as TR binds in vitro and in chromatin in vivo to a specific thyroid hormone-responsive element present in intron 1 of this gene. This is the first report concerning in vivo transcriptional control of the beta-catenin gene. As Wnt/beta-catenin plays a crucial role in intestinal tumorigenesis, our observations open a new perspective on the study of TRs as potential tumor inducers

A point mutation in the AF-2 domain of thyroid hormone receptor alpha1 expressed after CRE mediated recombination partially recapitulates hypothyroidism.

Thyroid hormones act directly on transcription by binding to TRα1, TRβ1, TRβ2 nuclear receptors, regulating many aspects of post-natal development and homeostasis. To precisely analyze the implication of the widely expressed TRα1 isoform in this pleiotropic action, we have generated transgenic mice with a point mutation in the TRα1 coding sequence, which is expressed only after CRE/loxP mediated DNA recombination. The amino-acid change prevents interaction between TRα1 and histone acetyltransferase coactivators and the release of corepressors. Early expression of this dominant-negative receptor deeply affects post-natal development and adult homeostasis, recapitulating many aspects of congenital and adult hypothyroidism, except in tissues and cells where TRβ1 and TRβ2 are predominantly expressed. Both respective abundance and intrinsic properties of TRα1 and TRβ1/2 seems to govern specificity of action

Single and Synergistic Effects of Cannabidiol and Δ-9-Tetrahydrocannabinol on Zebrafish Models of Neuro-Hyperactivity

In this study, we aimed to investigate the effect of the two main active cannabinoids extracted from cannabis: Δ-9-tetrahydrocannabinol (THC) and cannabidiol (CBD) on two distinct behavioral models of induced neuro-hyperactivity. We have taken advantage of two previously developed zebrafish models of neuro-hyperactivity: a chemically induced pentylenetetrazole model and a genetic model caused by loss-of-function mutations in the GABA receptor subunit alpha 1 (GABRA1−/−). Both CBD and THC have a significant effect on the behavioral changes induced by both models. Importantly, we have also shown that when applied together at different ratios of THC to CBD (1:1, 1:5, and 1:10), there was a synergistic effect at a ratio of 1:1. This was particularly important for the genetically induced neuro-hyperactivity as it brought the concentrations of THC and CBD required to oppose the induced behavioral changes to levels that had much less of an effect on baseline larval behavior. The results of this study help to validate the ability of THC and CBD to oppose neuro-hyperactivity linked to seizure modalities. Additionally, it appears that individually, each cannabinoid may be more effective against the chemically induced model than against the GABRA1−/− transgenic model. However, when applied together, the concentration of each compound required to oppose the GABRA1−/− light-induced activity was lowered. This is of particular interest since the use of cannabinoids as therapeutics can be dampened by their side-effect profile. Reducing the level of each cannabinoid required may help to prevent off target effects that lead to side effects. Additionally, this study provides a validation of the complimentary nature of the two zebrafish models and sets a platform for future work with cannabinoids, particularly in the context of neuro-hyperactivity disorders such as epilepsy

The transforming acidic coiled coil (TACC1) protein modulates the transcriptional activity of the nuclear receptors TR and RAR

Background: The transcriptional activity of Nuclear hormone Receptors (NRs) is regulated by interaction with coactivator or corepressor proteins. Many of these cofactors have been shown to have a misregulated expression or to show a subcellular mislocalization in cancer cell lines or primary tumors. Therefore they can be factors involved in the process of oncogenesis. Results: We describe a novel NR coregulator, TACC1, which belongs to the Transforming Acidic Coiled Coil (TACC) family. The interaction of TACC1 with Thyroid Hormone Receptors (TR) and several other NRs has been shown in a yeast two-hybrid screen and confirmed by GST pulldown, colocalization and co-immunoprecipitation experiments. TACC1 interacts preferentially with unliganded NRs. In F9 cells, endogenous TACC1 localized in the chromatin-enriched fraction of the nucleus and interacted with Retinoid Acid Receptors (RAR alpha) in the nucleus. TACC1 depletion in the cell led to decreased RAR alpha and TR alpha ligand-dependent transcriptional activity and to delocalization of TR from the nucleus to the cytoplasm. Conclusions: From these experimental studies we propose that TACC1 might be a scaffold protein building up a transcriptional complex around the NRs we studied. This function of TACC1 might account for its involvement in several forms of tumour development

Thyroid hormone receptor {beta} (TR{beta}) and liver X receptor (LXR) regulate carbohydrate response element binding protein (ChREBP) expression in a tissue selective manner.

Thyroid hormone- (TR) and Liver X- (LXR)receptors are transcription factors involved in lipogenesis. Both receptors recognize the same consensus DNA response element in vitro. It was previously shown that their signalling pathways interact in the control of cholesterol elimination in the liver. In the present study ChREBP, a major transcription factor controlling the activation of glucose-induced lipogenesis in liver, is characterized as a direct target of thyroid hormones(TH) in liver and white adipose tissue(WAT), the two main lipogenic tissues in mice. Using genetic and molecular approaches ChREBP is shown to be specifically regulated by TRbeta, but not by TRalpha in vivo even in WAT where both TR isoforms are expressed. However this isotype specificity is not found in vitro. This TRbeta specific regulation correlates with the loss of TH-induced lipogenesis in TRbeta-/- mice. Fasting/refeeding experiments show that TRbeta is not required for the activation of ChREBP expression particularly marked in WAT following refeeding. However TH can stimulate ChREBP expression in WAT even under fasting conditions suggesting completely independent pathways. Since ChREBP has been described as an LXR target, the interaction of LXR and TRbeta in ChREBP regulation was assayed both in vitro and in vivo. Each receptor recognizes a different response element on the ChREBP promoter, located only eight base pairs apart.There is a crosstalk between LXR and TRbeta signalling on the ChREBP promoter in liver but not in WAT where LXR does not regulate ChREBP expression. The molecular basis for this crosstalk has been determined in in vitro systems

The Oct4 homologue PouV and Nanog regulate pluripotency in chicken embryonic stem cells

International audienceEmbryonic stem cells ( ESC) have been isolated from pregastrulation mammalian embryos. The maintenance of their pluripotency and ability to self- renew has been shown to be governed by the transcription factors Oct4 ( Pou5f1) and Nanog. Oct4 appears to control cell- fate decisions of ESC in vitro and the choice between embryonic and trophectoderm cell fates in vivo. In nonmammalian vertebrates, the existence and functions of these factors are still under debate, although the identification of the zebrafish pou2 ( spg; pou5f1) and Xenopus Pou91 ( XlPou91) genes, which have important roles in maintaining uncommitted putative stem cell populations during early development, has suggested that these factors have common functions in all vertebrates. Using chicken ESC ( cESC), which display similar properties of pluripotency and long- term self- renewal to mammalian ESC, we demonstrated the existence of an avian homologue of Oct4 that we call chicken PouV ( cPouV). We established that cPouV and the chicken Nanog gene are required for the maintenance of pluripotency and self- renewal of cESC. These findings show that the mechanisms by which Oct4 and Nanog regulate pluripotency and self- renewal are not exclusive to mammal

Identification of curcumin analogues with anti-seizure potential in vivo using chemical and genetic zebrafish larva seizure models

Seizures are the outward manifestation of abnormally excessive or synchronous brain activity. While seizures can be somewhat symptomatically managed with anti-epileptic drugs (AEDs), many patients are still refractory to the currently available AEDs. As a result, there is a need to identify new molecules with anti-seizure properties. Curcumin is the principle curcuminoid of Curcuma longa, or colloquially turmeric, and has been experimentally proven to have anti-convulsive properties, but its poor bioavailability has dampened further therapeutic interest. Hence, this study aimed to ask if structural analogues of curcumin with an adequate bioavailability could have an anti-seizure effect in vivo. To do so, we tested these analogues following a multipronged approach combining the use of several zebrafish seizure models (chemically-induced and genetic) and complementary assays (behavioural and brain activity). Overall, from the 68 analogues tested, we found 15 different derivatives that were able to significantly decrease the behavioural hyperactivity induced by pentylenetetrazol. Of those, only a few showed an effect on the hyperactivity phenotype of two genetic models of brain seizures that are the gabra1 and gabrg2 knockouts. Two analogues, CA 80(1) and CA 74(1), were able to significantly alleviate brain seizures of gabrg2-mutant larvae. As a result, these analogues are good candidates as novel anti-seizure agents

Glycine decarboxylase deficiency-induced motor dysfunction in zebrafish is rescued by counterbalancing glycine synaptic level

Glycine encephalopathy (GE), or nonketotic hyperglycinemia (NKH), is a rare recessive genetic disease caused by defective glycine cleavage and characterized by increased accumulation of glycine in all tissues. Here, based on new case reports of GLDC loss-of-function mutations in GE patients, we aimed to generate a zebrafish model of severe GE in order to unravel the molecular mechanism of the disease. Using CRISPR/Cas9, we knocked out the gldc gene and showed that gldc–/– fish recapitulate GE on a molecular level and present a motor phenotype reminiscent of severe GE symptoms. The molecular characterization of gldc–/– mutants showed a broad metabolic disturbance affecting amino acids and neurotransmitters other than glycine, with lactic acidosis at stages preceding death. Although a transient imbalance was found in cell proliferation in the brain of gldc–/– zebrafish, the main brain networks were not affected, thus suggesting that GE pathogenicity is mainly due to metabolic defects. We confirmed that the gldc–/– hypotonic phenotype is due to NMDA and glycine receptor overactivation, and demonstrated that gldc–/– larvae depict exacerbated hyperglycinemia at these synapses. Remarkably, we were able to rescue the motor dysfunction of gldc–/– larvae by counterbalancing pharmacologically or genetically the level of glycine at the synapse