• Molecules and Cells
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• v.41 no.9
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• pp.842-852
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• 2018

Notch signaling is an evolutionarily conserved pathway and involves in the regulation of various cellular and developmental processes. Ligand binding releases the intracellular domain of Notch receptor (NICD), which interacts with DNA-bound CSL [CBF1/Su(H)/Lag-1] to activate transcription of target genes. In the absence of NICD binding, CSL down-regulates target gene expression through the recruitment of various corepressor proteins including SMRT/NCoR (silencing mediator of retinoid and thyroid receptors/nuclear receptor corepressor), SHARP (SMRT/HDAC1-associated repressor protein), and KyoT2. Structural and functional studies revealed the molecular basis of these interactions, in which NICD coactivator and corepressor proteins competitively bind to ${\beta}-trefoil$ domain (BTD) of CSL using a conserved ${\varphi}W{\varphi}P$ motif (${\varphi}$ denotes any hydrophobic residues). To date, there are conflicting ideas regarding the molecular mechanism of SMRT-mediated repression of CSL as to whether CSL-SMRT interaction is direct or indirect (via the bridge factor SHARP). To solve this issue, we mapped the CSL-binding region of SMRT and employed a 'one- plus two-hybrid system' to obtain CSL interaction-defective mutants for this region. We identified the CSL-interaction module of SMRT (CIMS; amino acid 1816-1846) as the molecular determinant of its direct interaction with CSL. Notably, CIMS contains a canonical ${\varphi}W{\varphi}P$ sequence (APIWRP, amino acids 1832-1837) and directly interacts with CSL-BTD in a mode similar to other BTD-binding corepressors. Finally, we showed that CSL-interaction motif, rather than SHARP-interaction motif, of SMRT is involved in transcriptional repression of NICD in a cell-based assay. These results strongly suggest that SMRT participates in CSL-mediated repression via direct binding to CSL.

• Molecules and Cells
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• v.44 no.1
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• pp.1-12
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• 2021

The nuclear receptor peroxisome proliferator-activated receptor γ (PPARγ) is the master transcriptional regulator in adipogenesis. PPARγ forms a heterodimer with another nuclear receptor, retinoid X receptor (RXR), to form an active transcriptional complex, and their transcriptional activity is tightly regulated by the association with either coactivators or corepressors. In this study, we identified T-cell death-associated gene 51 (TDAG51) as a novel corepressor of PPARγ-mediated transcriptional regulation. We showed that TDAG51 expression is abundantly maintained in the early stage of adipogenic differentiation. Forced expression of TDAG51 inhibited adipocyte differentiation in 3T3-L1 cells. We found that TDAG51 physically interacts with PPARγ in a ligand-independent manner. In deletion mutant analyses, large portions of the TDAG51 domains, including the pleckstrin homology-like, glutamine repeat and proline-glutamine repeat domains but not the proline-histidine repeat domain, are involved in the interaction with the region between residues 140 and 506, including the DNA binding domain, hinge, ligand binding domain and activation function-2 domain, in PPARγ. The heterodimer formation of PPARγ-RXRα was competitively inhibited in a ligand-independent manner by TDAG51 binding to PPARγ. Thus, our data suggest that TDAG51, which could determine adipogenic cell fate, acts as a novel negative regulator of PPARγ by blocking RXRα recruitment to the PPARγ-RXRα heterodimer complex in adipogenesis.

• Journal of Ginseng Research
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• v.48 no.1
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• pp.89-97
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• 2024

Background: Ginsenoside F2 (GF2), the protopanaxadiol-type constituent in Panax ginseng, has been reported to attenuate metabolic dysfunction-associated steatotic liver disease (MASLD). However, the mechanism of action is not fully understood. Here, this study investigates the molecular mechanism by which GF2 regulates MASLD progression through liver X receptor (LXR). Methods: To demonstrate the effect of GF2 on LXR activity, computational modeling of protein-ligand binding, Time-resolved fluorescence resonance energy transfer (TR-FRET) assay for LXR cofactor recruitment, and luciferase reporter assay were performed. LXR agonist T0901317 was used for LXR activation in hepatocytes and macrophages. MASLD was induced by high-fat diet (HFD) feeding with or without GF2 administration in WT and LXRα^-/- mice. Results: Computational modeling showed that GF2 had a high affinity with LXRα. LXRE-luciferase reporter assay with amino acid substitution at the predicted ligand binding site revealed that the S264 residue of LXRα was the crucial interaction site of GF2. TR-FRET assay demonstrated that GF2 suppressed LXRα activity by favoring the binding of corepressors to LXRα while inhibiting the accessibility of coactivators. In vitro, GF2 treatments reduced T0901317-induced fat accumulation and pro-inflammatory cytokine expression in hepatocytes and macrophages, respectively. Consistently, GF2 administration ameliorated hepatic steatohepatitis and improved glucose or insulin tolerance in WT but not in LXRα^-/- mice. Conclusion: GF2 alters the binding affinities of LXRα coregulators, thereby interrupting hepatic steatosis and inflammation in macrophages. Therefore, we propose that GF2 might be a potential therapeutic agent for the intervention in patients with MASLD.

• Journal of Life Science
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• v.30 no.2
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• pp.211-220
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• 2020

The activity of CAR can be regulated not only by ligand binding but also by phosphorylation of regulatory factors involved in extracellular signaling pathways, cross-talk interactions with transcription factors, and the recruitment, degradation, and expression of coactivators and corepressors. This regulation of CAR activity can in turn have effects on the control of diverse physiological homeostasis, including xenobiotic and energy metabolism, cellular proliferation, and apoptosis. CAR is phosphorylated by the ERK1/2 signaling pathway, which causes formation of a complex with Hsp-90 and CCRP, leading to its cytoplasmic retention, whereas phenobarbital inhibits ERK1/2, which causes dephosphorylation of the downstream signaling molecules, leading to the recruitment to CAR of the activated RACK-1/PP2A components for the dephosphorylation, nuclear translocation, and the transcriptional activation of CAR. Activated CAR cross-talks with FoxO1 to induce inhibition of its transcriptional activity and with PGC-1α to induce protein degradation by ubiquitination, resulting in the transcriptional suppression of PEPCK and G6Pase involved in gluconeogenesis. Regulation by CAR of lipid synthesis and oxidation is achieved by its functional cross-talks, respectively, with PPARγ through the degradation of PGC-1α to inhibit expression of the lipogenic genes and with PPARα through either the suppression of CPT-1 expression or the interaction with PGC-1α each to induce tissue-specific inhibition or stimulation of β-oxidation. Whereas CAR stimulates cellular proliferation by suppressing p21 expression through the inhibition of FoxO1 transcriptional activity and inducing cyclin D1 expression, it suppresses apoptosis by inhibiting the activities of MKK7 and JNK-1 through the expression of GADD45B. In conclusion, CAR is involved in the maintenance of homeostasis by regulating not only xenobiotic metabolism but also energy metabolism, cellular proliferation, and apoptosis through diverse cross-talk interactions with extracellular signaling pathways and intracellular regulatory factors.