The activation mechanism of the aryl hydrocarbon receptor (AhR) by molecular chaperone HSP90

The aryl hydrocarbon receptor is a member of the nuclear receptor superfamily that associates with the molecular chaperone HSP90 in the cytoplasm. The activation mechanism of the AhR is not yet fully understood. It has been proposed that after binding of ligands such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 3methylcholanthrene (3-MC), or β-naphthoflavone (β-NF), the AhR dissociates from HSP90 and translocates to the nucleus. It has also been hypothesized that the AhR translocates to the nucleus and forms a complex with HSP90 and other co-chaperones. There are a few reports about the direct association or dissociation of AhR and HSP90 due to difficulties in purifying AhR. We constructed and purified the PAS domain from AhR. Binding of the AhR-PAS domain to β-NF affinity resin suggested that it possesses ligand-binding affinity. We demonstrated that the AhR-PAS domain binds to HSP90 and the association is not affected by ligand binding. The ligand 17-DMAG inhibited binding of HSP90 to GST-PAS. In an immunoprecipitation assay, HSP90 was co-immunoprecipitated with AhR both in the presence or absence of ligand. Endogenous AhR decreased in the cytoplasm and increased in the nucleus of HeLa cells 15. min after treatment with ligand. These results suggested that the ligand-bound AhR is translocated to nucleus while in complex with HSP90.We used an in situ proximity ligation assay to confirm whether AhR was translocated to the nucleus alone or together with HSP90. HSP90 was co-localized with AhR after the nuclear translocation. It has been suggested that the ligand-bound AhR was translocated to the nucleus with HSP90. Activated AhR acts as a transcription factor, as shown by the transcription induction of the gene CYP1A1 8. h after treatment with β-NF

Development and preliminary evaluation of a tele-rehabilitation exercise system using computer-generated animation

OBJECTIVES: To evaluate the safety and acceptability of a newly developed tele-rehabilitation exercise system using computer-generated animation. METHODS: The participants comprised a convenience sample of 38 diverse individuals in Experiment 1 (15 healthy young people, 16 healthy older people, 5 patients with stroke, and 2 patients with respiratory disease) and 18 healthy older individuals in Experiment 2. Experiment 1 assessed safety in terms of cardiopulmonary vascular aspects and risk of fall, and Experiment 2 assessed treatment acceptability via a subjective evaluation. All participants completed the same exercise program. The safety assessment was conducted using heart rate (HR) and saturation of percutaneous oxygen (SpO2), measured before and after exercise. In addition, the occurrence of falls was assessed. For the acceptability assessment, the participants answered five questions (three-point Likert scale) after the exercise program. RESULTS: The safety assessment indicated that HR and SpO2 changed from 70.5±10.2 beats per minute and 97.8±1.3% before exercise to 87.6±13.6 beats per minute and 98.2±0.9% after exercise, respectively. In addition, all participants completed the exercises without experiencing any falls. In the acceptability assessment, the score reflecting continuation desire was the highest of the five items examined (2.71±0.46). In contrast, the adequacy of exercise intensity had the lowest score (1.29±0.57). CONCLUSIONS: The present system was confirmed to be safe, and the participants were motivated to continue the exercises. Future developments should incorporate a function to enable participants and medical staff to adjust exercise intensity according to individual physical function

Discovery of a Macropinocytosis‐Inducing Peptide Potentiated by Medium‐Mediated Intramolecular Disulfide Formation

Macropinocytosis is among ubiquitous cellular uptake mechanisms of peptide-based intracellular delivery. Due to its capability of engulfing large macromolecules, macropinocytosis shows promise as a route for the intracellular uptake of biomacromolecules and nanoparticles. We previously reported SN21, a peptide derived from the N-terminus of stromal cell-derived growth factor 1α (SDF-1α), as a potent macropinocytosis inducer. In this work, we obtained the 8-residue analog P4A bearing higher macropinocytosis induction ability. P4A contains vital cysteine residues in its sequence, which immediately reacts with cystine in culture medium to convert into its oxidized forms, including the intramolecularly oxidized form (oxP4A) as the dominant and active species. The conjugate of oxP4A with membrane lytic peptide LK15 delivered bioactive proteins into cells; notably, this peptide delivered functional proteins fused with a negatively charged protein tag at a significantly reduced amount (up to nanomolar range) without compromising the delivery efficiency and the cellular activities of delivered proteins

Discovery of a Macropinocytosis‐Inducing Peptide Potentiated by Medium‐Mediated Intramolecular Disulfide Formation

Macropinocytosis is among ubiquitous cellular uptake mechanisms of peptide-based intracellular delivery. Due to its capability of engulfing large macromolecules, macropinocytosis shows promise as a route for the intracellular uptake of biomacromolecules and nanoparticles. We previously reported SN21, a peptide derived from the N-terminus of stromal cell-derived growth factor 1α (SDF-1α), as a potent macropinocytosis inducer. In this work, we obtained the 8-residue analog P4A bearing higher macropinocytosis induction ability. P4A contains vital cysteine residues in its sequence, which immediately reacts with cystine in culture medium to convert into its oxidized forms, including the intramolecularly oxidized form (oxP4A) as the dominant and active species. The conjugate of oxP4A with membrane lytic peptide LK15 delivered bioactive proteins into cells; notably, this peptide delivered functional proteins fused with a negatively charged protein tag at a significantly reduced amount (up to nanomolar range) without compromising the delivery efficiency and the cellular activities of delivered proteins

The activation mechanism of the aryl hydrocarbon receptor (AhR) by molecular chaperone HSP90

The aryl hydrocarbon receptor is a member of the nuclear receptor superfamily that associates with the molecular chaperone HSP90 in the cytoplasm. The activation mechanism of the AhR is not yet fully understood. It has been proposed that after binding of ligands such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 3methylcholanthrene (3-MC), or β-naphthoflavone (β-NF), the AhR dissociates from HSP90 and translocates to the nucleus. It has also been hypothesized that the AhR translocates to the nucleus and forms a complex with HSP90 and other co-chaperones. There are a few reports about the direct association or dissociation of AhR and HSP90 due to difficulties in purifying AhR. We constructed and purified the PAS domain from AhR. Binding of the AhR-PAS domain to β-NF affinity resin suggested that it possesses ligand-binding affinity. We demonstrated that the AhR-PAS domain binds to HSP90 and the association is not affected by ligand binding. The ligand 17-DMAG inhibited binding of HSP90 to GST-PAS. In an immunoprecipitation assay, HSP90 was co-immunoprecipitated with AhR both in the presence or absence of ligand. Endogenous AhR decreased in the cytoplasm and increased in the nucleus of HeLa cells 15 min after treatment with ligand. These results suggested that the ligand-bound AhR is translocated to nucleus while in complex with HSP90. We used an in situ proximity ligation assay to confirm whether AhR was translocated to the nucleus alone or together with HSP90. HSP90 was co-localized with AhR after the nuclear translocation. It has been suggested that the ligand-bound AhR was translocated to the nucleus with HSP90. Activated AhR acts as a transcription factor, as shown by the transcription induction of the gene CYP1A1 8 h after treatment with β-NF