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1.
Nucleic Acids Res ; 2020 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-33010171

RESUMO

NBS1 is a critical component of the MRN (MRE11/RAD50/NBS1) complex, which regulates ATM- and ATR-mediated DNA damage response (DDR) pathways. Mutations in NBS1 cause the human genomic instability syndrome Nijmegen Breakage Syndrome (NBS), of which neuronal deficits, including microcephaly and intellectual disability, are classical hallmarks. Given its function in the DDR to ensure proper proliferation and prevent death of replicating cells, NBS1 is essential for life. Here we show that, unexpectedly, Nbs1 deletion is dispensable for postmitotic neurons, but compromises their arborization and migration due to dysregulated Notch signaling. We find that Nbs1 interacts with NICD-RBPJ, the effector of Notch signaling, and inhibits Notch activity. Genetic ablation or pharmaceutical inhibition of Notch signaling rescues the maturation and migration defects of Nbs1-deficient neurons in vitro and in vivo. Upregulation of Notch by Nbs1 deletion is independent of the key DDR downstream effector p53 and inactivation of each MRN component produces a different pattern of Notch activity and distinct neuronal defects. These data indicate that neuronal defects and aberrant Notch activity in Nbs1-deficient cells are unlikely to be a direct consequence of loss of MRN-mediated DDR function. This study discloses a novel function of NBS1 in crosstalk with the Notch pathway in neuron development.

2.
G3 (Bethesda) ; 2020 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-33020192

RESUMO

The flySAM/CRISPRa system has recently emerged as a powerful tool for gain-of-function studies in Drosophila melanogaster This system includes Gal4/UAS-driven dCas9 activators and U6 promoter-controlled sgRNA. Having established dCas9 activators superior to other combinations, to further enhance the efficiency of the targeting activators we systematically optimized the parameters of the sgRNA. Interestingly, the most efficient sgRNAs were found to accumulate in the region from -150bp to -450bp upstream of the transcription start site (TSS), and the activation efficiency showed a strong positive correlation with the GC content of the sgRNA targeting sequence. In addition, the target region is dominant to the GC content, as sgRNAs targeting areas beyond -600bp from the TSS lose efficiency even when containing 75% GC. Surprisingly, when comparing the activities of sgRNAs targeting to either DNA strand, sgRNAs targeting to the non-template strand outperform those complementary to the template strand, both in cells and in vivo In summary, we define criteria for sgRNA design which will greatly facilitate the application of CRISPRa in gain-of-function studies.

3.
J Invest Dermatol ; 2020 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-33058860

RESUMO

Codeine stimulates skin mast cells (MCs) and is therefore used in skin tests and as inducer of experimental itch. MRGPRX2 responds to various drugs, including opioids, to elicit pseudo-allergic reactions, but whether it represents the main opiate receptor of skin MCs remains unknown. By combining a number of approaches, including silencing of MRGPRX2, we now report that MRGPRX2 is indeed the dominant codeine receptor of dermal MCs. Activation by codeine displayed profound subject-variability and correlated with secretion elicited by compound 48/80 (c48/80) or Substance P, but not by FcεRI-aggregation. Degranulation by codeine was attenuated by SCF, while the opposite was found for FcεRI. C48/80 or codeine alone were able to achieve maximum MRGPRX2 activation. MRGPRX2 was rapidly internalized on codeine binding in a ß-arrestin-1-dependent manner. Codeine-triggered ß-arrestin activation was also established by Tango assay. Pre-stimulation with MRGPRX2-agonists (but not C3a or FcεRI-aggregation) resulted in refractoriness to further stimulation by the same or another MRGPRX2 ligand (cross-desensitization). This was duplicated in a cell line (RBL-MRGPRX2). Collectively, codeine degranulates skin MCs via MRGPRX2, at which it acts as a balanced ligand. It has yet to be determined whether codeine-induced refractoriness could be exploited to desensitize MRGPRX2 to prevent severe pseudo-allergic reactions.

4.
Food Funct ; 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-33057513

RESUMO

Peptides extracted from Xuanwei ham (XHP) can prevent free radical-induced diseases. The aim of the present study was to isolate and identify bioactive peptides from Xuanwei hams that rescue the oxidative stress damage induced by alcohol in HHL-5 hepatocytes. Alcohol-treated HHL-5 human hepatocytes were utilized as the alcohol-induced hepatocyte damage model to evaluate the effects of XHP on amounts of aminotransferase (ALT), aspartate aminotransferase (AST) and malondialdehyde (MDA). The result showed that XHP could significantly reduce ALT, AST and MDA, the major biomarkers of liver damage. The crude XHP was separated by size exclusion chromatography, followed by the evaluation of respective activities. Then, the most active components were further separated by RP-HPLC, and their activities were evaluated according to the above method. The peptide was identified as a hexapeptide with the sequence of Asn-Pro-Pro-Lys-Phe-Asp (NPPKFD) through LC-MS/MS. Further, the molecular mechanisms by which NPPKFD prevents alcohol-induced oxidative stress damage were revealed. Results showed that the hexapeptide could downregulate CYP2E1 expression, reduce generation of ROS and enhance oxidant defense systems via the activation of NrF2/HO-1 pathway. The findings suggest that Xuanwei ham can be used as a new source of bioactive peptides for protection from alcohol-induced liver damage.

5.
Chem Asian J ; 2020 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-33021084

RESUMO

An electron-assisted strategy was developed to prepare gold nanoparticles (AuNPs) at room temperature. Glow discharge plasma as electron source was successfully used to control the valence state, size, and shape of AuNPs. Stable Au(I) was obtained in 3 min by plasma, and Au(I) was reduced to zero valence with the increase in treatment time. An increase in the amount of Au did not induce an increase in particle size. A narrow size distribution was also achieved. The narrowest size distribution was observed at 9 min at 600 V. AuNPs grew slowly under glow discharge plasma, which slightly changed the mean size of AuNPs. Moreover, the average size of AuNPs was smaller under alkaline conditions. The initial pH of the solution can affect the nucleation and growth of AuNPs and further affect their particle size. Spherical AuNPs, hexagonal AuNPs, rectangular AuNPs, flower-shaped AuNPs, and Au nanorods were easily obtained within 30 min by adding different additives. The hexagonal AuNPs exhibited the largest current response toward caffeine and showed a good linear range (0.1-1000 µM) with a low detection limit (0.064 µM), because their high-energy planes can increase the electron transfer rate and improve electrocatalytic activity.

6.
Int J Neurosci ; : 1-8, 2020 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-33032501

RESUMO

OBJECTIVE: The cerebral ischemia-reperfusion (I/R) model is crucial for the study of cerebral stroke. Chrysophanol (Chry) can protect nerve damage of mice in cerebral ischemia-reperfusion injury. This study aimed at investigating the neuroprotective effects of chrysophanol through mitochondrial autophagy in mice with ischemia-reperfusion injury. MATERIALS AND METHODS: Adult mice were stochastically divided into five groups: sham, I/R (solvent), I/R+Chry (dose, 10.0ml/kg), I/R+Chry (dose, 1.0ml/kg), and I/R+Chry (dose, 0.1ml/kg). The cerebral ischemia-reperfusion model was made in I/R and I/R+Chry groups. The changes in hippocampal formation were observed by hematoxylin and eosin (H&E) staining. The expressions of LC3B-II and LC3B-I protein in hippocampus were demonstrated by western blot (WB). The fluorescence intensities of NIX, LC3B, and mitochondria were detected by immunohistochemistry fluorescent (IF). RESULTS: Comparing with the I/R group, the I/R+Chry groups showed improvements in reducing the damage on the hippocampus, indicated by the reduced ratio of LC3B-II and LC3B-I protein, decreased fluorescence intensity of NIX and LC3B, and increased intensity of mitochondrial fluorescence. CONCLUSION: Our study showed that chrysophanol may regulate mitochondrial autophagy through NIX protein and alleviate the damage of hippocampus through decreasing the level of mitochondrial autophagy.

7.
Nat Commun ; 11(1): 5076, 2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-33033264

RESUMO

Proper threat-reward decision-making is critical to animal survival. Emerging evidence indicates that the motor system may participate in decision-making but the neural circuit and molecular bases for these functions are little known. We found in C. elegans that GABAergic motor neurons (D-MNs) bias toward the reward behavior in threat-reward decision-making by retrogradely inhibiting a pair of premotor command interneurons, AVA, that control cholinergic motor neurons in the avoidance neural circuit. This function of D-MNs is mediated by a specific ionotropic GABA receptor (UNC-49) in AVA, and depends on electrical coupling between the two AVA interneurons. Our results suggest that AVA are hub neurons where sensory inputs from threat and reward sensory modalities and motor information from D-MNs are integrated. This study demonstrates at single-neuron resolution how motor neurons may help shape threat-reward choice behaviors through interacting with other neurons.

8.
Chem Eng J ; : 127240, 2020 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-33052192

RESUMO

Antibacterial agents with enzyme-like properties and bacteria-binding ability have provided an alternative method to efficiently disinfect drug-resistance microorganism. Herein, a Fe3O4@MoS2-Ag nanozyme with defect-rich rough surface was constructed by a simple hydrothermal method and in-situ photodeposition of Ag nanoparticles. The nanozyme exhibited good antibacterial performance against E. coli (~69.4%) by the generated ROS and released Ag+, while the nanozyme could further achieve an excellent synergistic disinfection (~100%) by combining with the near-infrared photothermal property of Fe3O4@MoS2-Ag. The antibacterial mechanism study showed that the antibacterial process was determined by the collaborative work of peroxidase-like activity, photothermal effect and leakage of Ag+. The defect-rich rough surface of MoS2 layers facilitated the capture of bacteria, which enhanced the accurate and rapid attack of •OH and Ag+ to the membrane of E. coli with the assistance of local hyperthermia. This method showed broad-spectrum antibacterial performance against Gram-negative bacteria, Gram-positive bacteria, drug-resistant bacteria and fungal bacteria. Meanwhile, the magnetism of Fe3O4 was used to recycle the nanozyme. This work showed great potential of engineered nanozymes for efficient disinfection treatment.

9.
Biomaterials ; 264: 120453, 2020 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-33069138

RESUMO

Thiol capped gold nanoparticles with small size, high dispersity, and broad light absorption covering ultraviolet (UV) to near infrared (NIR) region have been developed for catalysis, fluorescence imaging and photodynamic therapy (PDT). The constitution of the metal core in such nanoparticles can strongly influence the luminescence, catalysis, and stability properties. However, to date, a corresponding investigation of the influence of the metallic core on the generation of reaction oxygen species (ROS) and its therapeutic efficiency towards tumor cells remains to be lacking. Herein, we fabricated bimetallic nanoparticles by introducing bismuth into captopril capped gold nanoparticles. Surprisingly, the introduction of the Bi was found enhance the photothermal effect of the nanoparticles to a great extent, and the variation trends for the thermal effect, ROS generation rate, and tumor cell inhibition effect were found to disparate with the changes in the Au and Bi composition. The origin of the photothermal effect was deduced through density functional theory calculations based on microscopic construction. Combined with the intrinsic photodynamic effect, the bimetallic nanoparticles showed an outstanding tumor cell inhibition effect. Furthermore, due to the excellent CT imaging property, our designed nanoparticles provide the exciting possibility to realize CT imaging guided and light-mediated tumor therapy.

10.
Artigo em Inglês | MEDLINE | ID: mdl-33040537

RESUMO

The vulcanization of rubber is a chemical process to improve the mechanical properties by cross-linking unsaturated polymer chains. Zinc oxide (ZnO) acts as an activator, boosting the rubbers' sulfur vulcanization. Maintaining the level of ZnO content in the rubber compounds as low as possible is desirable, not only for economic reasons but also to reduce the environmental footprint of the process. In this contribution, octylamine (OA) capped ZnO nanoparticles (5 nm diameter), prepared through a thermal decomposition method, were demonstrated to be efficient activators for the sulfur vulcanization of natural rubber, enabling the reduction of the required amount of ZnO as compared to commercial systems. The effect of different ZnO activators (OA capped ZnO/commercial indirect process ZnO) on the curing characteristics, cross-linking densities, and mechanical performance, as well as the thermal behavior of rubber compounds, were investigated. Compared to the commercial indirect process ZnO, OA capped ZnO nanoparticles not only effectively enhanced the curing efficiency of natural rubber but also improved the mechanical performance of the composites after vulcanization. This was interpreted as, by applying the OA capped ZnO nanoparticles, the ZnO levels in rubber compounding were significantly reduced under the industrial vulcanization condition (151 °C, 30 min).

11.
J Hazard Mater ; 404(Pt A): 124083, 2020 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-33011634

RESUMO

Boscalid is a persistent fungicide that is frequently detected in surface waters and may be neurotoxic to aquatic organisms. Herein, we evaluated the effects of environmentally relevant boscalid concentrations to zebrafish to explore its potentially neurotoxic mechanisms of effect. Behavioral responses (swimming, phototaxis, and predation), histopathology, transcriptomics, biochemical parameter analysis and gene expression of larval and adult zebrafish following boscalid treatment were assessed. We found that boscalid significantly inhibited the locomotor ability and phototactic response of larvae after an 8-d exposure, and altered the locomotor activity, predation trajectories and ability in adults after a 21-d exposure. It was noted that predation rates of zebrafish were significantly decreased by 30% and 100% after exposure to 0.1 and 1.0 mg/L boscalid, respectively. Adverse alterations in the cell differentiation of eyes and brain injury were also observed in both larvae and adults following boscalid exposure. The expression of genes related to neurodevelopment, neurotransmission, eye development, and visual function, in conjunction with RNA-Seq results, indicated that boscalid may impair visual phototransduction and nervous system processes in larval zebrafish. Conclusively, boscalid exposure may affect the neurobehavioral response of zebrafish by impairing proper visual and nervous system function.

12.
Spectrochim Acta A Mol Biomol Spectrosc ; 246: 118960, 2020 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-33017795

RESUMO

Drug-induced liver injury (DILI) is a prevalent liver disease and the leading cause for acute liver failure (ALF) worldwide. Screening of DILI in patients is central to ensure drug safety and improve therapy efficiency. Mounting evidences revealed that peroxynitrite (ONOO-) is involved in the DILI process and can be a potential biomarker for DILI. Thus far, there are few two-photon fluorescence probes for ONOO- that can accomplish this challenging task in DILI liver tissues. Hereby, a peroxynitrite activatable two-photon fluorescence probe BN-PN for the imaging of ONOO- in mice liver was elaborately constructed. The probe specifically reacted with peroxynitrite to furnish 140-fold fluorescence increase in vitro, which elucidated a high sensitivity for ONOO-. Thus, subtle changes of ONOO- levels in live cells can be sensitively imaged with this probe by two-photon microscopy. The probe also denoted the overproduction of ONOO- in APAP-induced liver injury, and proved that administration with NAC can effectively alleviate DILI and reduce ONOO- production in mouse liver. Further, the probe demonstrated the rapid rise of ONOO- level in the liver of DILI mice administrated with alcohol. This work disclosed the rational construction of a two-photon fluorescence probe-based DILI screening method, which would help the estimation of drug safety and new drug development.

13.
J Integr Neurosci ; 19(3): 413-420, 2020 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-33070519

RESUMO

Electrical stimulation in the brain is an emerging therapy for treating a wide range of neurological disorders. Although electrical pulses are commonly used in the clinic, other electrical waveforms such as sinusoidal-waves have been investigated to improve the therapeutic efficacy, to reduce the risk of tissue damage induced by stimulation, and to decrease the consumption of electrical energy. However, the effects of sinusoidal stimulation on neuronal activity are still unclear. In the present study, we investigated the neuronal responses to the stimulation of 50-Hz sinusoidal-waves applied on the afferent fibers of the neurons in the hippocampal CA1 region of Sprague-Dawley rat in vivo. Results show that the stimulation increased the firing rate of both pyramidal neurons and interneurons in the downstream region of stimulation. Also, the stimulation eliminated the original theta rhythms (2-5 Hz) in the single-unit activity of the two types of neurons and entrained these neurons to fire at the stimulation rhythm. These results provide new clues for the mechanisms of brain stimulation to suppress the pathological rhythms in the neuronal activity, and for the application of sinusoidal waveforms in brain stimulation therapy.

14.
Cell Adh Migr ; 14(1): 153-164, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32881638

RESUMO

FNDC4 is an anti-inflammatory factor that alters the activation state of macrophages; it is used to treat colitis in mice. However, its role in muscle formation and mechanism of function remains unknown. We found that FNDC4 promotes the bovine MDSCs migration and differentiation. Furthermore, we reported that it interacts with integrin ß1 (ITGß1). FAK, mediated by ITGß1, regulates cell migration. Our results found FNDC4 to influence the expression of p-FAK, p-paxillin, and vinculin. Then, overexpressed or added FNDC4 protein could not influence migration and differentiation any more when the activated form of FAK was reduced. Therefore, we concluded that FNDC4 promotes the differentiation and migration of bovine MDSCs via the FAK, mediated by the ITGß1 receptor.

15.
ISME J ; 2020 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-32887943

RESUMO

Ambient conditions shape microbiome responses to both short- and long-duration environment changes through processes including physiological acclimation, compositional shifts, and evolution. Thus, we predict that microbial communities inhabiting locations with larger diel, episodic, and annual variability in temperature and pH should be less sensitive to shifts in these climate-change factors. To test this hypothesis, we compared responses of surface ocean microbes from more variable (nearshore) and more constant (offshore) sites to short-term factorial warming (+3 °C) and/or acidification (pH -0.3). In all cases, warming alone significantly altered microbial community composition, while acidification had a minor influence. Compared with nearshore microbes, warmed offshore microbiomes exhibited larger changes in community composition, phylotype abundances, respiration rates, and metatranscriptomes, suggesting increased sensitivity of microbes from the less-variable environment. Moreover, while warming increased respiration rates, offshore metatranscriptomes yielded evidence of thermal stress responses in protein synthesis, heat shock proteins, and regulation. Future oceans with warmer waters may enhance overall metabolic and biogeochemical rates, but they will host altered microbial communities, especially in relatively thermally stable regions of the oceans.

16.
J Med Syst ; 44(10): 182, 2020 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-32885290

RESUMO

Physiological signals can contain abundant personalized information and indicate health status and disease deterioration. However, in current medical practice, clinicians working in the general wards are usually lack of plentiful means and tools to continuously monitor the physiological signals of the inpatients. To address this problem, we here presented a medical-grade wireless monitoring system based on wearable and artificial intelligence technology. The system consists of a multi-sensor wearable device, database servers and user interfaces. It can monitor physiological signals such as electrocardiography and respiration and transmit data wirelessly. We highly integrated the system with the existing hospital information system and explored a set of processes of physiological signal acquisition, storage, analysis, and combination with electronic health records. Multi-scale information extracted from physiological signals and related to the deterioration or abnormality of patients could be shown on the user interfaces, while a variety of reports could be provided daily based on time-series signal processing technology and machine learning to make more information accessible to clinicians. Apart from an initial attempt to implement the system in a realistic clinical environment, we also conducted a preliminary validation of the core processes in the workflow. The heart rate veracity validation of 22 patient volunteers showed that the system had a great consistency with ECG Holter, and bias for heart rate was 0.04 (95% confidence interval: -7.34 to 7.42) beats per minute. The Bland-Altman analysis showed that 98.52% of the points were located between Mean ± 1.96SD. This system has been deployed in the general wards of the Hyperbaric Oxygen Department and Respiratory Medicine Department and has collected more than 1000 cases from the clinic. The whole system will continue to be updated based on clinical feedback. It has been demonstrated that this system can provide reliable physiological monitoring for patients in general wards and has the potential to generate more personalized pathophysiological information related to disease diagnosis and treatment from the continuously monitored physiological data.

17.
Artigo em Inglês | MEDLINE | ID: mdl-32959146

RESUMO

Aspergillus oryzae lipase (AOL) is a potential biocatalyst for industrial application. In this study, a mutant lipase AOL-3F38N/V230R was screened through two rounds of directed evolution, resulting in a fourfold increase in lipase activity, and threefold in catalytic efficiency (kcat/Km), while maintaining its excellent stereoselectivity. AOL-3F38N/V230R enzyme activity was maximum at pH 7.5 and also at 40 °C. And compared with wild-type AOL-3, AOL-3F38N/V230R preferentially hydrolyzed the fatty acid ethyl ester carbon chain length from C4 to C6-C10. In the same catalytic reaction conditions, the conversion of (R,S)-ethyl-2-(4-hydroxyphenoxy) propanoate ((R,S)-EHPP) by AOL-3F38N/V230R can be increased 169.7% compared to the original enzyme. The e.e.s of (R,S)-EHPP achieved 99.4% and conversion about 50.2% with E value being 829.0. Therefore, AOL-3F38N/V230R was a potential biocatalyst for obtaining key chiral compounds for aryloxyphenoxy propionate (APP) herbicides.

18.
J Int Med Res ; 48(9): 300060520957180, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32954890

RESUMO

OBJECTIVE: Low serum free triiodothyronine (FT3) levels are associated with the occurrence of coronary heart disease and with the prognosis of cardiovascular diseases. This study aimed to investigate the relationship between FT3 levels and risk stratification in Chinese Han patients with acute coronary syndrome (ACS) receiving percutaneous coronary intervention (PCI) treatment. METHODS: Plasma FT3 levels and other parameters were measured in 191 patients with ACS who received PCI. The risk of adverse cardiovascular events was assessed using the Age, Creatinine, and Ejection Fraction (ACEF) score. RESULTS: FT3 levels were significantly lower in the high-risk group than in the medium- and low-risk groups. Serum FT3 levels were negatively linearly correlated with the ACEF score (r = -0.590). Stepwise regression analysis showed a negative correlation between FT3 levels and the risk of adverse cardiovascular events as measured by the ACEF score (standardized ß = -0.261). CONCLUSION: Serum FT3 levels are negatively related to risk stratification in patients with ACS. Serum FT3 levels may be used as a potential predictor for adverse outcomes of patients with ACS undergoing PCI.

19.
Anal Chem ; 92(19): 13305-13312, 2020 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-32907322

RESUMO

A malignant tumor remains one of the leading causes of deaths across the world. Thus, diagnosis of tumor development with noninvasive visualizing methods is significant for tumor therapy. Herein, an activatable two-photon NIR fluorescent probe DHQ-Rd-PN for in vivo imaging of peroxynitrite in a tumor was elaborately designed. The probe demonstrated an increased NIR emission in response to peroxynitrite in vitro, which ensured that the probe detects ONOO- in cell and in vivo. Cellular imaging results disclosed that the probe was competent to detect adscititious ONOO- level change in HeLa cells, as well as endogenous ONOO- concentration in lipopolysaccharides (LPS) and IFN-γ-stimulated RAW 264.7 cells. Additionally, zebrafish in vivo imaging revealed that the probe accumulated in the pancreas and was lightened up by the addition of ONOO-. Remarkably, the probe can be harnessed to image an ONOO- production profile in xenograft 4T1 tumor mice by both one-photon and two-photon in vivo fluorescence imaging. Benefiting with the two-photon excitable properties and NIR emissive properties, the probe can be used for noninvasive in vivo imaging of ONOO- in the onset and development of tumors for the first time. This work provided a noninvasive and efficient detection method for ONOO- in a tumor, which would find more applications in tumor diagnosis and therapies.

20.
Proc Natl Acad Sci U S A ; 117(40): 25128-25137, 2020 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-32958651

RESUMO

Melatonin (Mel) promotes sleep through G protein-coupled receptors. However, the downstream molecular target(s) is unknown. We identified the Caenorhabditis elegans BK channel SLO-1 as a molecular target of the Mel receptor PCDR-1-. Knockout of pcdr-1, slo-1, or homt-1 (a gene required for Mel synthesis) causes substantially increased neurotransmitter release and shortened sleep duration, and these effects are nonadditive in double knockouts. Exogenous Mel inhibits neurotransmitter release and promotes sleep in wild-type (WT) but not pcdr-1 and slo-1 mutants. In a heterologous expression system, Mel activates the human BK channel (hSlo1) in a membrane-delimited manner in the presence of the Mel receptor MT1 but not MT2 A peptide acting to release free Gßγ also activates hSlo1 in a MT1-dependent and membrane-delimited manner, whereas a Gßλ inhibitor abolishes the stimulating effect of Mel. Our results suggest that Mel promotes sleep by activating the BK channel through a specific Mel receptor and Gßλ.

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