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Expression of nicotinic acetylcholine receptor subunit α9 in type Ⅱ vesti-bular hair cells of rats

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Acta Pharmacologica Sinica 2006年第11期27卷 1509-1514页

作者： Wei-jia KONG~(2,4,5)Hua-mao CHENG~(2,4)Paul van CAUWENBERGE~3 2 Department of Otorhinolaryngology,Union Hospital of Tongji Medical College,Huazhong Science and Technology University,Wuhan 430030,China 3 Department of Otorhinolaryngology,Ghent University Hospital,De Pintelaan 185,9000 Gent,Belgium Department of Otorhinolaryngology Union Hospital of Tongji Medical College Huazhong Science and Technology University Wuhan China Department of Otorhinolaryngology Ghent University Hospital Gent Belgium

Aim:To explore the cell specific existence of α9 AChR in the vestibular type Ⅱ haircells(VHC Ⅱ)of ***:To detect the expression of α9 AChR messengerRNA(mRNA)in the vestibular endorgans and single VHC Ⅱ of rats by using thereverse transcription polymerase chain reaction(RT-PCR)technique and the singlecell RT-PCR technique,***:It was shown that α9 AChR mRNAwas detected in the vestibular *** using single-cell RT-PCR,mRNAencoding α9 AChR was also detected in the VHC Ⅱ of the *** analysisof the PCR products confirmed identity to corresponding cDNA sequence in thepredicted ***:We established a method which could effectivelydetect the cell specific expression of mRNA in an individual *** dataconfirm that α9 AChR mRNA is expressed in the VHC Ⅱ of rats and indicates thatα9 AChR may function as a mediator of efferent cholinergic signaling in mamma-lian VHC.

关键词： hair cells vestibule rats nicotinic acetylcholine receptor single cell reverse transcriptase polymerase chain reaction

Influence of Y151 F mutation in loop B on the agonist potency in insect nicotinic acetylcholine receptor

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Insect Science 2009年第5期16卷 365-369页

作者： Feng Song Yi-Xi Zhang Xiang-Mei Yao Ze-Wen Liu Key Laboratory of Monitoring and Management of Plant Disease and Insects Ministry of Agriculture Nanjing Agricultural UniversityNanjing China

nicotinic acetylcholine receptors （nAChRs） are ligand-gated ion channels, which mediate fast cholinergic synaptic transmission in insect and vertebrate nervous systems. The nAChR agonist-binding site is present at the interface of adjacent subunits and is formed by loops A-C present in α subunits together with loops D-F present in either non-α subunits or homomer-forming α subunits. Although Y151 in loop B has been identified as important in agonist binding, various residues at the 151-site are found among vertebrate and invertebrate nAChR α subunits, such as F 151. In Xenopus oocytes expressing Nlα1 or Nlα1^Y151F plus rat β2, Y151F mutation was found to significantly change the rate of receptor desensitization and altered the pharmacological properties of acetylcholine, but not imidacloprid, including the decrease Of Imax, the increase of EC50 （the concentration causing 50% of the maximum response） and the fast time-constant of decay （τf）. By comparisons of residue structure, the hydroxyl group in the side chain of Y151 was thought to be important in the interaction between Nlα1/β2 nAChRs and acetylcholine, and the phenyl group to be important between Nlα1/β2 nAChRs and imidacloprid.

关键词： acetylcholine agonist potency imidacloprid nicotinic acetylcholine receptor Nilaparvata lugens Xenopus oocytes

Nanoscale interactions between the nicotinic acetylcholine receptor and cholesterol

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BIOCELL 2021年第6期45卷 1479-1484页

作者： FRANCISCO J.BARRANTES Laboratory of Molecular Neurobiology Institute of Biomedical Research(BIOMED)UCA-CONICETFaculty of Medical SciencesBuenos AiresArgentina

Cholesterol is a major lipid in biological *** not only plays a structural role but also modulates a wide range of functional properties of neurotransmitter and hormone receptors and ion *** membraneembedded segments of the paradigm neurotransmitter receptor for acetylcholine(nAChR)contain linear sequences of amino acids with the capacity to recognize *** cholesterol consensus domains have been designated as“CARC”and its mirror sequence“CRAC”.CARC preferentially occurs in the exoplasmic-facing membrane leaflet,and CRAC,in the cytoplasmic-facing *** motifs are highly conserved among ion-channel and neurotransmitter receptor proteins in vertebrate nervous systems,where they recognize cholesterol,and in prokaryotic homologues in bacteria,where they recognize *** phylogenetically conserved trait is an indication that the hopanoids in some bacteria and cholesterol in eukaryotes subserve analogous functions,probably contributing to the stability of membrane-embedded protein *** studies from our laboratory using superresolution optical microscopy(“nanoscopy”)have disclosed other interrelated functional and structural properties exerted by cholesterol on the *** neutral lipid content at the cell surface influences both the macromolecular organization of the receptor and its translational mobility(diffusion)in the plane of the membrane.

关键词： Cholesterol Pentameric ligand-gated ion channel nicotinic acetylcholine receptor Membrane proteins Evolution Nanoscopy Cholesterol-recognition domains

Possible implications of dysregulated nicotinic acetylcholine receptor diffusion and nanocluster formation in myasthenia gravis

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Neural Regeneration Research 2021年第2期16卷 242-246页

作者： Francisco J.Barrantes Biomedical Research Institute(BIOMED)UCA-CONICET Buenos AiresArgentina

Myasthenia gravis is a rare and invalidating disease affecting the neuromuscular junction of voluntary *** classical form of this autoimmune disease is characterized by the presence of antibodies against the most abundant protein in the neuromuscular junction,the nicotinic acetylcholine *** variants of the disease involve autoimmune attack of non-receptor scaffolding proteins or enzymes essential for building or maintaining the integrity of this peripheral *** review summarizes the participation of the above proteins in building the neuromuscular junction and the destruction of this cholinergic synapse by autoimmune aggression in myasthenia *** review also covers the application of a powerful biophysical technique,superresolution optical microscopy,to image the nicotinic receptor in live cells and follow its motional *** hypothesis is entertained that anomalous nanocluster formation by antibody crosslinking may lead to accelerated endocytic internalization and elevated turnover of the receptor,as observed in myasthenia gravis.

关键词： agrin autoimmune diseases muscle end-plate muscle specific kinase MuSK myasthenia gravis nanoscopy neuromuscular junction nicotinic acetylcholine receptor rapsyn superresolution microscopy

Computational Determination of the Binding Mode of α-Conotoxin to nicotinic acetylcholine receptor

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Journal of Ocean University of China 2016年第6期15卷 1027-1033页

作者： TABASSUM Nargis YU Rilei JIANG Tao Key Laboratory of Marine Drugs of Chinese Ministry of Education School of Medicine and PharmacyOcean University of China Qingdao 266003 P. R. China

Abstract Conotoxins belong to the large families of disulfide-rich peptide toxins from cone snail venom, and can act on a broad spectrum of ion channels and receptors. They are classified into different subtypes based on their targets. The a-conotoxins selectively inhibit the current of the nicotinic acetylcholine receptors. Because of their unique selectivity towards distinct nAChR subtypes, a-conotoxins become valuable tools in nAChR study. In addition to the X-ray structures of a-conotoxins in complex with acetyleholine-binding protein, a homolog of the nAChR ligand-binding domain, the high-resolution crystal structures of the extracellular domain of the al and a9 subunits are also obtained. Such structures not only revealed the details of the configuration of nAChR, but also provided higher sequence identity templates for modeling the binding modes of a-conotoxins to nAChR. This mini-review summarizes recent modeling studies for the determination of the binding modes of a-conotoxins to nAChR. As there are not crystal structures of the nAChR in complex with conotoxins, computational modeling in combination of mutagenesis data is expected to reveal the molecular recognition mechanisms that govern the interactions between a-conotoxins and nAChR at molecular level. An accurate determination of the binding modes of a-conotoxins on AChRs allows rational design of a-conotoxin analogues with improved potency or selectivity to nAChRs.

关键词： nicotinic acetylcholine receptor a-conotoxin acetylcholine binding protein docking homology modeling moleculardynamics simulation mutational energy

Limitations of RNAi of e6 nicotinic acetylcholine receptor subunits for assessing the in vivo sensitivity to spinosad

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Insect Science 2013年第1期20卷 101-108页

作者： Frank D. Rinkevich Jeffrey G. Scott Department of Entomology Comstock Hall Cornell University Ithaca New York USA

Spinosad is a widely used insecticide that exerts its toxic effect primarily through interactions with the nicotinic acetylcholine receptor. The a6 nicotinic acetyl- choline receptor subunit is involved in spinosad toxicity as demonstrated by the high levels of resistance observed in strains lacking a6. RNAi was performed against the Da6 nicotinic acetylcholine receptor subunit in Drosophila melanogaster using the Gal4-UAS system to examine if RNAi would yield results similar to those of a6 null mutants. These Da6-deficient flies were subject to spinosad contact bioassays to evaluate the role of the Da6 nicotinic acetylcholine receptor subunit on spinosad sensitivity. The expression of Da6 was reduced 60%-75% as verified by quantitative polymerase chain reaction. However, there was no change in spinosad sensitivity in D. melanogaster. We repeated RNAi experiments in Tribolium castaneum using injection of dsRNA for Tcasct6. RNAi of Tcasa6 did not result in changes in spinosad sensitivity, similar to results obtained with D. melanogaster. The lack of change in spinosad sensitivity in both D. melanogaster and T. castaneum using two routes of dsRNA administration shows that RNAi may not provide adequate conditions to study the role of nicotinic acetylcholine receptor subunits on insecticide sensitivity due to the inability to completely eliminate expression of the a6 subunit in both species. Potential causes for the lack of change in spinosad sensitivity are discussed.

关键词： bioassay insecticide target site nicotinic acetylcholine receptor RNAi,spinosad

Cloning and Sequence of nicotinic acetylcholine receptor α Subunit from Chilo suppressalis

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Zoological Research 2002年第1期23卷 7-13页

作者：韩招久韩召军南京农业大学农业部病虫监测与治理重点开放实验室江苏南京210095

nicotinic acetylcholine receptors (nAChRs) play a significant role in excitatory synaptic transmission in insects and are the target for chloronicotinyl and nereistoxin *** recent years,Chilo suppressalis,an economically important pest of rice,developed high resistance against monosultap,a nereistoxin insecticide acting on *** order to reveal the hypothesized target insensitive mechanism,studies on the molecular property of nAChR from Chilo suppressalis are *** this study,the full length cDNA of nAChR α subunit from this pest was cloned by *** analysis shows that it is a novel nAChR α subunit,which was named as Cs α 1(Genbank accession ***418987).It contains 1?997?bp nucleotides and involves an open reading frame (ORF) encoding a mature protein of 509 amino acids excluding a signal peptide of 24 amino *** deduced amino acid sequence was 52%-94% identical to the reported insect nAChR genes.

关键词： Chilo suppressalis Gene cloning nicotinic acetylcholine receptor α subunit

Neuronal nicotinic acetylcholine receptors serve as sensitive targets that mediate β-amyloid neurotoxicity

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Acta Pharmacologica Sinica 2006年第10期27卷 1277-1286页

作者： Qiang LIU Jie WU Division of Neurology,Barrow Neurological Institute,St Joseph’s Hospital and Medical Center,Phoenix,Arizona 85013,USA Division of Neurology Barrow Neurological Institute St Joseph's Hospital and Medical Center Phoenix USA

Alzheimer’s disease (AD) is the most common form of brain dementia characterizedby the accumulation of β-amyloid peptides (Aβ) and loss of forebrain cholinergicneurons.Aβ accumulation and aggregation are thought to contribute to cholin-ergic neuronal degeneration,in turn causing learning and memory deficits,but thespecific targets that mediate Aβ neurotoxicity remain ***,accumu-lating lines of evidence have demonstrated that Aβ directly modulates the func-tion of neuronal nicotinic acetylcholine receptors (nAChRs),which leads to thenew hypothesis that neuronal nAChRs may serve as important targets that medi-ate Aβ *** this review,we summarize current studies performed inour laboratory and in others to address the question of how Aβ modulates neu-ronal nAChRs,especially nAChR subunit function.

关键词： nicotinic acetylcholine receptor β-amyloid peptides neurodegeneration Alzheimer’s disease

Two Potent α3/5 Conotoxins from Piscivorous Conus achatinus

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Acta Biochimica et Biophysica Sinica 2007年第6期39卷 438-444页

作者： Geoffrey CHEW Edward HAWROT Department of Molecular Pharmacology Physiology and Biotechnology Brown University Providence Rhode Island 02903 USA

Every cone snail produces a mixture of different conotoxins and secretes them to immobilize their prey and predators. α3/5 Conotoxins, isolated from fish-hunting cone snails, target muscle nicotinic acetylcholine receptors. The structure and function of α3/5 conotoxin from the piscivorous Conus achatinus have not been studied. We synthesized two pentadecamer peptides, Ac 1.1 a and Ac 1.1 b, with appropriate disulfide bonding, based on cDNA sequences of α3/5 conotoxins from C. achatinus. Ac 1.1 a and Ac 1.1 b differ by only one amino acid residue. They have similar potency on blocking recombinant mouse muscle acetylcholine receptor expressed in Xenopus laevis oocytes, with IC50values of 36 nM and 26 nM, respectively. For Ac 1.1b, deletion of the first three N-terminal amino acids did not change its activity, indicating that the Nterminus is not involved in the interaction with its receptor. Furthermore, our experiments indicate that both toxins strongly prefer the α1-δ subunit interface instead of the α1-γ binding site on the mouse muscle nicotinic acetylcholine receptor. These peptides provide additional tools for the study of the structure and function of nicotinic receptor.

关键词： cone snail α3/5 conotoxin disulfide bond nicotinic acetylcholine receptor