• Development and Reproduction
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• v.25 no.3
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• pp.133-143
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• 2021

In contrast to the human lutropin receptor (hLHR) and rat LHR (rLHR), very few naturally occurring mutants in other mammalian species have been identified. The present study aimed to delineate the mechanism of signal transduction by three constitutively activating mutants (designated M410T, L469R, and D590Y) and two inactivating mutants (D383N and Y546F) of the eel LHR, known to be naturally occurring in human LHR transmembrane domains. The mutants were constructed and measured cyclic adenosine monophosphate (cAMP) accumulation via homogeneous time-resolved fluorescence assays in Chinese hamster ovary (CHO)-K1 cells. The activating mutant cells expressing eel LHR-M410T, L469R, and D590Y exhibited a 4.0-, 19.1-, and 7.8-fold increase in basal cAMP response without agonist treatment, respectively. However, inactivating mutant cells expressing D417N and Y558F did not completely impaired signal transduction. Specifically, signal transduction in the cells expressing activating mutant L469R was not occurred with a further ligand stimulation, showing that the maximal response exhibited approximately 53% of those of wild type receptor. Our results suggested that the constitutively activating mutants of the eel LHR consistently occurred without agonist treatment. These results provide important information of LHR function in fish and regulation with regard to mutations of highly conserved amino acids in glycoprotein hormone receptors.

• Genomics & Informatics
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• v.20 no.4
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• pp.45.1-45.7
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• 2022

Food security will be affected by climate change worldwide, particularly in the developing world, where the most important food products originate from plants. Plants are often exposed to environmental stresses that may affect their growth, development, yield, and food quality. Auxin is a hormone that plays a critical role in improving plants' tolerance of environmental conditions. Auxin controls the expression of many stress-responsive genes in plants by interacting with specific cis-regulatory elements called auxin-responsive elements (AuxREs). In this work, we performed an in silico prediction of AuxREs in promoters of five auxin-responsive genes in Zea mays. We applied a data fusion approach based on the combined use of Dempster-Shafer evidence theory and fuzzy sets. Auxin has a direct impact on cell membrane proteins. The short-term auxin response may be represented by the regulation of transmembrane gene expression. The detection of an AuxRE in the promoter of prolyl oligopeptidase (POP) in Z. mays and the 3-fold overexpression of this gene under auxin treatment for 30 min indicated the role of POP in maize auxin response. POP is regulated by auxin to perform stress adaptation. In addition, the detection of two AuxRE TGTCTC motifs in the upstream sequence of the bx1 gene suggests that bx1 can be regulated by auxin. Auxin may also be involved in the regulation of dehydration-responsive element-binding and some members of the protein kinase superfamily.

• Membrane Journal
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• v.33 no.1
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• pp.34-45
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• 2023

Two computational intelligence techniques namely artificial neural networks (ANN) and support vector machine (SVM) are employed to model the permeate flux based on seven input variables including time, transmembrane pressure, rotating velocity, the pore diameter of the membrane, dynamic viscosity, concentration and density of the feed fluid. The best-fit model was selected through the trial-error method and the two statistical parameters including the coefficient of determination (R²) and the average absolute relative deviation (AARD) between the experimental and predicted data. The obtained results reveal that the optimized ANN model can predict the permeate flux with R² = 0.999 and AARD% = 2.245 versus the SVM model with R² = 0.996 and AARD% = 4.09. Thus, the ANN model is found to predict the permeate flux with high accuracy in comparison to the SVM approach.

• Biomolecules & Therapeutics
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• v.30 no.2
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• pp.191-202
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• 2022

Tetrazoles were designed and synthesized as potential inhibitors of triple monoamine neurotransmitters (dopamine, norepinephrine, serotonin) reuptake based on the functional and docking simulation of compound 6 which were performed in a previous study. The compound structure consisted of a tetrazole-linker (n)-piperidine/piperazine-spacer (m)-phenyl ring, with tetrazole attached to two phenyl rings (R1 and R2). Altering the carbon number in the linker (n) from 3 to 4 and in the spacer (m) from 0 to 1 increased the potency of serotonin reuptake inhibition. Depending on the nature of piperidine/piperazine, the substituents at R1 and R2 exerted various effects in determining their inhibitory effects on monoamine reuptake. Docking study showed that the selectivity of tetrazole for different transporters was determined based on multiple interactions with various residues on transporters, including hydrophobic residues on transmembrane domains 1, 3, 6, and 8. Co-expression of dopamine transporter, which lowers dopamine concentration in the biophase by uptaking dopamine into the cells, inhibited the dopamine-induced endoctytosis of dopamine D₂ receptor. When tested for compound 40 and 56, compound 40 which has more potent inhibitory activity on dopamine reuptake more strongly disinhibited the inhibitory activity of dopamine transporter on the endocytosis of dopamine D₂ receptor. Overall, we identified candidate inhibitors of triple monoamine neurotransmitter reuptake and provided a theoretical background for identifying such neurotransmitter modifiers for developing novel therapeutic agents of various neuropsychiatric disorders.

• Biomolecules & Therapeutics
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• v.32 no.2
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• pp.205-213
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• 2024

Hydroxychavicol, a primary active phenolic compound of betel leaves, previously inhibited bone loss in vivo by stimulating osteogenesis. However, the effect of hydroxychavicol on bone remodeling induced by osteoclasts is unknown. In this study, the anti-osteoclastogenic effects of hydroxychavicol and its mechanism were investigated in receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclasts. Hydroxychavicol reduced the number of tartrate resistance acid phosphatase (TRAP)-positive multinucleated, F-actin ring formation and bone-resorbing activity of osteoclasts differentiated from RAW264.7 cells in a concentration-dependent manner. Furthermore, hydroxychavicol decreased the expression of osteoclast-specific genes, including cathepsin K, MMP-9, and dendritic cell-specific transmembrane protein (DC-STAMP). For mechanistic studies, hydroxychavicol suppressed RANKL-induced expression of major transcription factors, including the nuclear factor of activated T-cells 1 (NFATc1), c-Fos, and c-Jun. At the early stage of osteoclast differentiation, hydroxychavicol blocked the phosphorylation of NF-κB subunits (p65 and Iκβα). This blockade led to the decrease of nuclear translocation of p65 induced by RANKL. In addition, the anti-osteoclastogenic effect of hydroxychavicol was confirmed by the inhibition of TRAP-positive multinucleated differentiation from human peripheral mononuclear cells (PBMCs). In conclusion, hydroxychavicol inhibits osteoclastogenesis by abrogating RANKL-induced NFATc1 expression by suppressing the NF-κB signaling pathway in vitro.

• The Korean Journal of Physiology and Pharmacology
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• v.28 no.2
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• pp.113-120
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• 2024

Solute carrier 40A1 (SLC40A1) encodes ferroportin, which is the only known transmembrane protein that exports elemental iron from mammalian cells and is essential for iron homeostasis. Mutations in SLC40A1 are associated with iron-overload disorders. In addition to ferroportin diseases, SLC40A1 expression is downregulated in various cancer types. Despite the clinical significance of the SLC40A1 transporter, only a few studies have investigated genetic variants in SLC40A1. The present study was performed to identify genetic variations in the SLC40A1 promoter and functionally characterize each variant using in vitro assays. We investigated four haplotypes and five variants in the SLC40A1 promoter. We observed that haplotype 3 (H3) had significantly lower promoter activity than H1, whereas the activity of H4 was significantly higher than that of H1. Luciferase activity of H2 was comparable to that of H1. In addition, four variants of SLC40A1, c.-1355G>C, c.-662C>T, c.-98G>C, and c.-8C>G, showed significantly increased luciferase activity compared to the wild type (WT), whereas c.-750G>A showed significantly decreased luciferase activity compared to the WT. Three transcription factors, cAMP response element-binding protein-1 (CREB-1), chicken ovalbumin upstream promoter transcription factor 1, and hepatic leukemia factor (HLF), were predicted to bind to the promoter regions of SLC40A1 near c.-662C>T, c.-98G>C, and c.-8C>G, respectively. Among these, CREB1 and HLF bound more strongly to the variant sequences than to the WT and functioned as activators of SLC40A1 transcription. Collectively, our findings indicate that the two SLC40A1 promoter haplotypes affect SLC40A1 transcription, which is regulated by CREB-1 and HLF.

• Korean Circulation Journal
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• v.53 no.4
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• pp.239-250
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• 2023

Background and Objectives: Brugada syndrome (BrS) is an inherited arrhythmia syndrome that presents as sudden cardiac death (SCD) without structural heart disease. One of the mechanisms of SCD has been suggested to be related to the uneven dispersion of transient outward potassium current (I_to) channels between the epicardium and endocardium, thus inducing ventricular tachyarrhythmia. Artemisinin is widely used as an antimalarial drug. Its antiarrhythmic effect, which includes suppression of I_to channels, has been previously reported. We investigated the effect of artemisinin on the suppression of electrocardiographic manifestations in a canine experimental model of BrS. Methods: Transmural pseudo-electrocardiograms and epicardial/endocardial transmembrane action potentials (APs) were recorded from coronary-perfused canine right ventricular wedge preparations (n=8). To mimic the BrS phenotypes, acetylcholine (3 μM), calcium channel blocker verapamil (1 μM), and I_to agonist NS5806 (6-10 μM) were used. Artemisinin (100-150 μM) was then perfused to ameliorate the ventricular tachyarrhythmia in the BrS models. Results: The provocation agents induced prominent J waves in all the models on the pseudo-electrocardiograms. The epicardial AP dome was attenuated. Ventricular tachyarrhythmia was induced in six out of 8 preparations. Artemisinin suppressed ventricular tachyarrhythmia in all 6 of these preparations and recovered the AP dome of the right ventricular epicardium in all preparations (n=8). J wave areas and epicardial notch indexes were also significantly decreased after artemisinin perfusion. Conclusions: Our findings suggest that artemisinin has an antiarrhythmic effect on wedge preparation models of BrS. It might work by inhibition of potassium channels including I_to channels, subsequently suppressing ventricular tachycardia/ventricular fibrillation.

• Animal Bioscience
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• v.37 no.8
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• pp.1367-1376
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• 2024

Objective: Parathyroid hormone like hormone (PTHLH), as an essential factor for bone growth, is involved in a variety of physiological processes. The aim of this study was to explore the role of PTHLH gene in the growth of antlers. Methods: The coding sequence (CDS) of PTHLH gene cDNA was obtained by cloning in sika deer (Cervus nippon), and the bioinformatics was analyzed. The quantitative real-time polymerase chain reaction (qRT-PCR) was used to analyze the differences expression of PTHLH mRNA in different tissues of the antler tip at different growth periods (early period, EP; middle period, MP; late period, LP). Results: The CDS of PTHLH gene was 534 bp in length and encoded 177 amino acids. Predictive analysis results revealed that the PTHLH protein was a hydrophilic protein without transmembrane structure, with its secondary structure consisting mainly of random coil. The PTHLH protein of sika deer had the identity of 98.31%, 96.82%, 96.05%, and 94.92% with Cervus canadensis, Bos mutus, Oryx dammah and Budorcas taxicolor, which were highly conserved among the artiodactyls. The qRT-PCR results showed that PTHLH mRNA had a unique spatio-temporal expression pattern in antlers. In the dermis, precartilage, and cartilage tissues, the expression of PTHLH mRNA was extremely significantly higher in MP than in EP, LP (p<0.01). In the mesenchyme tissue, the expression of PTHLH mRNA in MP was significantly higher than that of EP (p<0.05), but extremely significantly lower than that of LP (p<0.01). The expression of PTHLH mRNA in antler tip tissues at all growth periods had approximately the same trend, that is, from distal to basal, it was first downregulated from the dermis to the mesenchyme and then continuously up-regulated to the cartilage tissue. Conclusion: PTHLH gene may promote the rapid growth of antler mainly through its extensive regulatory effect on the antler tip tissue.

• Membrane and Water Treatment
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• v.15 no.2
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• pp.51-57
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• 2024

The removal of phosphorus, especially phosphate-form phosphorus, is necessary in wastewater treatment. Biofouling induced by the quorum sensing mechanism is also a major problem in membrane bioreactor (MBR), which reduces membrane flux. This study introduces lanthanum-doped quorum quenching (QQ) beads into MBR, confirming their inhibitory effect on biofouling due to Rhodococcus sp. BH4 and their capacity for phosphorus removal through lanthanum adsorption. A batch test was conducted to access the phosphate adsorption of lanthanum-QQ (La-QQ) beads and lab-scale MBR to verify the effect of inhibition. The study aimed to identify distinctions among the MBR, QQ MBR, and La-QQ MBR. In the batch test, the phosphate removal rate increased as the volume of beads increased, while the unit volume removal rate of phosphate decreased. In the lab-scale MBR, the phosphate removal rates were below 20% in the control MBR and QQ MBR, whereas the La-QQ MBR achieved a phosphate removal rate of 74%. There was not much difference between the ammonia and total organic carbon (TOC) removal rates. Regarding the change in transmembrane pressure(TMP), 3.7 days were taken for the control MBR to reach critical pressure. In contrast, the QQ-MBR took 9.8 days, and the La-QQ MBR took 6.1 days, which confirms the delay in biofouling. It is expected that La-QQ can be used within MBR to design a more stable MBR process that regulates biofouling and enhances phosphate removal.

• Molecules and Cells
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• v.27 no.5
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• pp.591-599
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• 2009

Nicotinic acetylcholine receptors (nAChRs) play important roles in nervous system functions and are involved in a variety of diseases. We previously demonstrated that ginsenosides, the active ingredients of Panax ginseng, inhibit subsets of nAChR channel currents, but not ${\alpha}7$, expressed in Xenopus laevis oocytes. Mutation of the highly conserved Leu247 to Thr247 in the transmembrane domain 2 (TM2) channel pore region of ${\alpha}7$ nAChR induces alterations in channel gating properties and converts ${\alpha}7$ nAChR antagonists into agonists. In the present study, we assessed how point mutations in the Leu247 residue leading to various amino acids affect 20(S)-ginsenoside $Rg_3$ ($Rg_3$) activity against the ${\alpha}7$ nAChR. Mutation of L247 to L247A, L247D, L247E, L247I, L247S, and L247T, but not L247K, rendered mutant receptors sensitive to $Rg_3$. We further characterized $Rg_3$ regulation of L247T receptors. We found that $Rg_3$ inhibition of mutant ${\alpha}7$ nAChR channel currents was reversible and concentration-dependent. $Rg_3$ inhibition was strongly voltage-dependent and noncompetitive manner. These results indicate that the interaction between $Rg_3$ and mutant receptors might differ from its interaction with the wild-type receptor. To identify differences in $Rg_3$ interactions between wild-type and L247T receptors, we utilized docked modeling. This modeling revealed that $Rg_3$ forms hydrogen bonds with amino acids, such as Ser240 of subunit I and Thr244 of subunit II and V at the channel pore, whereas $Rg_3$ localizes at the interface of the two wild-type receptor subunits. These results indicate that mutation of Leu247 to Thr247 induces conformational changes in the wild-type receptor and provides a binding pocket for $Rg_3$ at the channel pore.