DOCSLIB.ORG

Download File

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Download File STRUCTURAL AND FUNCTIONAL STUDIES OF TRPML1 AND TRPP2 Nicole Marie Benvin Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Graduate School of Arts and Sciences COLUMBIA UNIVERSITY 2017 © 2017 Nicole Marie Benvin All Rights Reserved ABSTRACT Structural and Functional Studies of TRPML1 and TRPP2 Nicole Marie Benvin In recent years, the determination of several high-resolution structures of transient receptor potential (TRP) channels has led to significant progress within this field. The primary focus of this dissertation is to elucidate the structural characterization of TRPML1 and TRPP2. Mutations in TRPML1 cause mucolipidosis type IV (MLIV), a rare neurodegenerative lysosomal storage disorder. We determined the first high-resolution crystal structures of the human TRPML1 I-II linker domain using X-ray crystallography at pH 4.5, pH 6.0, and pH 7.5. These structures revealed a tetramer with a highly electronegative central pore which plays a role in the dual Ca2+/pH regulation of TRPML1. Notably, these physiologically relevant structures of the I-II linker domain harbor three MLIV-causing mutations. Our findings suggest that these pathogenic mutations destabilize not only the tetrameric structure of the I-II linker, but also the overall architecture of full-length TRPML1. In addition, TRPML1 proteins containing MLIV- causing mutations mislocalized in the cell when imaged by confocal fluorescence microscopy. Mutations in TRPP2 cause autosomal dominant polycystic kidney disease (ADPKD). Since novel technological advances in single-particle cryo-electron microscopy have now enabled the determination of high-resolution membrane protein structures, we set out to solve the structure of TRPP2 using this technique. Our investigations offer valuable insight into the optimization of TRPP2 protein purification and sample preparation procedures necessary for structural analysis. Table of Contents List of Figures and Tables ........................................................................................................... vi Acknowledgements ...................................................................................................................... ix Dedication .................................................................................................................................... xii Chapter I Introduction to TRPML1 and TRPP2 1.1 Transient receptor potential (TRP) channel family .................................................................. 1 1.2 Introduction to the TRPML1 protein ........................................................................................ 3 1.2.1 TRPML1 and its role in mucolipidosis type IV (MLIV) ................................................... 4 1.2.2 Subcellular localization and channel regulation of TRPML1 ........................................... 4 1.2.3 Trafficking of TRPML1 ..................................................................................................... 6 1.2.4 The TRPML subfamily ..................................................................................................... 7 1.2.5 Electrophysiological properties of TRPML channels ........................................................ 7 1.3 Introduction to the TRPP2 protein and ADPKD ...................................................................... 9 1.3.1 Clinical manifestations of ADPKD .................................................................................. 10 1.3.2 Pathogenesis of ADPKD .................................................................................................. 11 1.3.3 Cellular and subcellular localization of TRPP2 ............................................................... 12 1.3.4 Mechanotransduction model of the PKD1/TRPP2 complex ............................................ 13 i 1.3.5 TRPP2 and its role in ADPKD ......................................................................................... 14 1.3.6 Stoichiometric assembly of the PKD1/TRPP2 complex .................................................. 14 1.3.7 TRPP2 agonist triptolide .................................................................................................. 15 1.3.8 Interaction between TRPP2 and additional channels ....................................................... 16 1.3.9 Ion selectivity in TRPP2 .................................................................................................. 16 1.3.10 Additional members of the TRPP subfamily ................................................................. 17 1.4 Structural determination of TRP channels .............................................................................. 18 1.5 Conclusion and thesis overview ............................................................................................. 20 1.6 References ............................................................................................................................... 22 Chapter II Structural and Functional Studies of the TRPML1 I-II Linker 2.1 Introduction ............................................................................................................................. 30 2.2 Experimental procedures for structrual studies ...................................................................... 31 2.2.1 Protein expression and purification of the TRPML1 I-II linker ...................................... 31 2.2.2 Protein crystallization of the TRPML1 I-II linker ........................................................... 33 2.2.3 Data collection and structure determination .................................................................... 34 2.3 Structural features of the TRPML1 I-II linker structures ....................................................... 38 2.3.1 TRPML1 I-II linker structure at pH 6.0 ........................................................................... 38 2.3.2 Features of the central luminal pore ................................................................................ 39 2.3.3 TRPML1 I-II linker structures at pH 4.5 and pH 7.5 ....................................................... 41 ii 2.3.4 Verification of the tetrameric I-II linker .......................................................................... 43 2.3.5 Role of the I-II linker in TRPML1 assembly .................................................................. 45 2.4 Structural basis of pH and Ca2+ regulation of TRPML1......................................................... 47 2.4.1 Dual regulation of TRPML1 activity by pH and Ca2+ ..................................................... 47 2.4.2 The central pore is essential for pH and Ca2+ regulation .................................................. 48 2.4.3 Model of pH and Ca2+ dual regulation of TRPML1 ......................................................... 50 2.5 Characterization of MLIV-causing mutations ........................................................................ 53 2.5.1 MLIV-causing mutations in the TRPML1 I-II linker ...................................................... 53 2.5.2 Cell culture, transfection, and confocal live-cell imaging ............................................... 54 2.5.3 Confocal imaging of MLIV-causing mutations .............................................................. 55 2.5.4 Biochemical analysis and circular dichroism spectroscopy ............................................ 58 2.6 Conclusion .............................................................................................................................. 60 2.7 References .............................................................................................................................. 62 Chapter III Structural Studies of TRPP2 3.1 Introduction ............................................................................................................................. 64 3.2 Screening of TRPP2 constructs using FSEC ......................................................................... 65 3.2.1 Rationale for cloning truncated constructs ....................................................................... 65 3.2.2 Transient HEK cell transfection and FSEC...................................................................... 66 3.2.3 FSEC analysis of human TRPP2 constructs transfected in HEK293 cells ..................... 67 3.3 Insect cell baculovirus-based expression system .................................................................... 69 iii 3.3.1 Generation of recombinant baculoviruses ........................................................................ 69 3.3.2 FSEC analysis of human TRPP2 constructs transfected into Sf9 cells ............................ 70 3.4 FSEC of mouse TRPP2 constructs transfected in HEK293 cells ........................................... 71 3.5 Purification of hTRPP2 in Hi5 cells using DDM and amphipol ............................................ 73 3.5.1 Small-scale purification in DDM ..................................................................................... 74 3.5.2 Reconstitution into amphipols .......................................................................................... 75 3.5.3 Inhibition of N-glycosylation ........................................................................................... 79 3.5.4 Large-scale purification of TRPP2 in DDM ................................................................... 82 3.6 Purification of hTRPP2 in Hi5 cells using LMNG ................................................................
Load more

Recommended publications
  • Natural Product Modulators of Transient Receptor Potential (TRP) Channels As Potential Cite This: Chem
    Chem Soc Rev View Article Online TUTORIAL REVIEW View Journal | View Issue Natural product modulators of transient receptor potential (TRP) channels as potential Cite this: Chem. Soc. Rev., 2016, 45,6130 anti-cancer agents Tiago Rodrigues,a Florian Sieglitza and Gonçalo J. L. Bernardes*ab Treatment of cancer is a significant challenge in clinical medicine, and its research is a top priority in chemical biology and drug discovery. Consequently, there is an urgent need for identifying innovative chemotypes capable of modulating unexploited drug targets. The transient receptor potential (TRPs) channels persist scarcely explored as targets, despite intervening in a plethora of pathophysiological events in numerous diseases, including cancer. Both agonists and antagonists have proven capable of evoking phenotype changes leading to either cell death or reduced cell migration. Among these, natural products entail biologically pre-validated and privileged architectures for TRP recognition. Furthermore, several natural products have significantly contributed to our current knowledge on TRP biology. In this Creative Commons Attribution 3.0 Unported Licence. Tutorial Review we focus on selected natural products, e.g. capsaicinoids, cannabinoids and terpenes, by highlighting challenges and opportunities in their use as starting points for designing natural product- Received 17th December 2015 inspired TRP channel modulators. Importantly, the de-orphanization of natural products as TRP channel DOI: 10.1039/c5cs00916b ligands may leverage their exploration as viable strategy for developing anticancer therapies. Finally, we foresee that TRP channels may be explored for the selective pharmacodelivery of cytotoxic payloads to www.rsc.org/chemsocrev diseased tissues, providing an innovative platform in chemical biology and molecular medicine.
    [Show full text]
  • A Computational Model of Large Conductance Voltage and Calcium
    Journal of Computational Neuroscience (2019) 46:233–256 https://doi.org/10.1007/s10827-019-00713-9 A computational model of large conductance voltage and calcium activated potassium channels: implications for calcium dynamics and electrophysiology in detrusor smooth muscle cells Suranjana Gupta1 · Rohit Manchanda1 Received: 11 September 2018 / Revised: 14 February 2019 / Accepted: 19 February 2019 / Published online: 25 April 2019 © Springer Science+Business Media, LLC, part of Springer Nature 2019 Abstract The large conductance voltage and calcium activated potassium (BK) channels play a crucial role in regulating the excitability of detrusor smooth muscle, which lines the wall of the urinary bladder. These channels have been widely characterized in terms of their molecular structure, pharmacology and electrophysiology. They control the repolarising and hyperpolarising phases of the action potential, thereby regulating the firing frequency and contraction profiles of the smooth muscle. Several groups have reported varied profiles of BK currents and I-V curves under similar experimental conditions. However, no single computational model has been able to reconcile these apparent discrepancies. In view of the channels’ physiological importance, it is imperative to understand their mechanistic underpinnings so that a realistic model can be created. This paper presents a computational model of the BK channel, based on the Hodgkin-Huxley formalism, constructed by utilising three activation processes — membrane potential, calcium inflow from voltage-gated calcium channels on the membrane and calcium released from the ryanodine receptors present on the sarcoplasmic reticulum. In our model, we attribute the discrepant profiles to the underlying cytosolic calcium received by the channel during its activation. The model enables us to make heuristic predictions regarding the nature of the sub-membrane calcium dynamics underlying the BK channel’s activation.
    [Show full text]
  • The Role of Transient Receptor Potential Cation Channels in Ca2þ Signaling
    Downloaded from http://cshperspectives.cshlp.org/ on October 7, 2021 - Published by Cold Spring Harbor Laboratory Press The Role of Transient Receptor Potential Cation Channels in Ca2þ Signaling Maarten Gees, Barbara Colsoul, and Bernd Nilius KU Leuven, Department of Molecular Cell Biology, Laboratory Ion Channel Research, Campus Gasthuisberg, Herestraat 49, bus 802, Leuven, Belgium Correspondence: [email protected] The 28 mammalian members of the super-family of transient receptor potential (TRP) channels are cation channels, mostly permeable to both monovalent and divalent cations, and can be subdivided into six main subfamilies: the TRPC (canonical), TRPV (vanilloid), TRPM (melastatin), TRPP (polycystin), TRPML (mucolipin), and the TRPA (ankyrin) groups. TRP channels are widely expressed in a large number of different tissues and cell types, and their biological roles appear to be equally diverse. In general, considered as poly- modal cell sensors, they play a much more diverse role than anticipated. Functionally, TRP channels, when activated, cause cell depolarization, which may trigger a plethora of voltage-dependent ion channels. Upon stimulation, Ca2þ permeable TRP channels 2þ 2þ 2þ generate changes in the intracellular Ca concentration, [Ca ]i,byCa entry via the plasma membrane. However, more and more evidence is arising that TRP channels are also located in intracellular organelles and serve as intracellular Ca2þ release channels. This review focuses on three major tasks of TRP channels: (1) the function of TRP channels as Ca2þ entry channels; (2) the electrogenic actions of TRPs; and (3) TRPs as Ca2þ release channels in intracellular organelles. ransient receptor potential (TRP) channels choanoflagellates, yeast, and fungi are primary Tconstitute a large and functionally versatile chemo-, thermo-, or mechanosensors (Cai 2008; family of cation-conducting channel proteins, Wheeler and Brownlee 2008; Chang et al.
    [Show full text]
  • The Intracellular Ca2+ Release Channel TRPML1 Regulates Lower Urinary Tract Smooth Muscle Contractility
    The intracellular Ca2+ release channel TRPML1 regulates lower urinary tract smooth muscle contractility Caoimhin S. Griffina, Michael G. Alvaradoa, Evan Yamasakia, Bernard T. Drummb,c, Vivek Krishnana, Sher Alia, Eleanor M. Naglea, Kenton M. Sandersb, and Scott Earleya,1 aDepartment of Pharmacology, Center for Molecular and Cellular Signaling in the Cardiovascular System, Reno School of Medicine, University of Nevada, Reno, NV 89557-0318; bDepartment of Physiology and Cell Biology, Reno School of Medicine, University of Nevada, Reno, NV 89557-0318; and cDepartment of Life & Health Sciences, Dundalk Institute of Technology, Louth, Ireland A91 K584 Edited by Mark T. Nelson, University of Vermont, Burlington, VT, and approved October 13, 2020 (received for review August 12, 2020) TRPML1 (transient receptor potential mucolipin 1) is a Ca2+-perme- including dense granulomembranous storage bodies in neurons, able, nonselective cation channel that is predominantly localized to elevated plasma gastrin, vacuolization in the gastric mucosa, and the membranes of late endosomes and lysosomes (LELs). Intracellular retinal degeneration (14). Interestingly, however, an anatomical release of Ca2+ through TRPML1 is thought to be pivotal for mainte- examination of these mice reveals dramatically distended bladders nance of intravesicular acidic pH as well as the maturation, fusion, and (14), leading us to question how TRPML1, an intracellular Ca2+- trafficking of LELs. Interestingly, genetic ablation of TRPML1 in mice release channel important in LEL function, affects bladder −/− (Mcoln1 ) induces a hyperdistended/hypertrophic bladder phenotype. physiology. Here, we investigated this phenomenon further by exploring an un- The lower urinary tract (LUT) is composed of the urinary conventional role for TRPML1 channels in the regulation of Ca2+-signal- bladder and urethra—structures that serve the simple, reciprocal ing activity and contractility in bladder and urethral smooth muscle cells functions of storing and voiding urine (15).
    [Show full text]
  • Chapter Four – TRPA1 Channels: Chemical and Temperature Sensitivity
    CHAPTER FOUR TRPA1 Channels: Chemical and Temperature Sensitivity Willem J. Laursen1,2, Sviatoslav N. Bagriantsev1,* and Elena O. Gracheva1,2,* 1Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, CT, USA 2Program in Cellular Neuroscience, Neurodegeneration and Repair, Yale University School of Medicine, New Haven, CT, USA *Corresponding author: E-mail: [email protected], [email protected] Contents 1. Introduction 90 2. Activation and Regulation of TRPA1 by Chemical Compounds 91 2.1 Chemical activation of TRPA1 by covalent modification 91 2.2 Noncovalent activation of TRPA1 97 2.3 Receptor-operated activation of TRPA1 99 3. Temperature Sensitivity of TRPA1 101 3.1 TRPA1 in mammals 101 3.2 TRPA1 in insects and worms 103 3.3 TRPA1 in fish, birds, reptiles, and amphibians 103 3.4 TRPA1: Molecular mechanism of temperature sensitivity 104 Acknowledgments 107 References 107 Abstract Transient receptor potential ankyrin 1 (TRPA1) is a polymodal excitatory ion channel found in sensory neurons of different organisms, ranging from worms to humans. Since its discovery as an uncharacterized transmembrane protein in human fibroblasts, TRPA1 has become one of the most intensively studied ion channels. Its function has been linked to regulation of heat and cold perception, mechanosensitivity, hearing, inflam- mation, pain, circadian rhythms, chemoreception, and other processes. Some of these proposed functions remain controversial, while others have gathered considerable experimental support. A truly polymodal ion channel, TRPA1 is activated by various stimuli, including electrophilic chemicals, oxygen, temperature, and mechanical force, yet the molecular mechanism of TRPA1 gating remains obscure. In this review, we discuss recent advances in the understanding of TRPA1 physiology, pharmacology, and molecular function.
    [Show full text]
  • Toxic Effects As a Result of Herbal Medicine Intake Toxic Effects As a Result of Herbal Medicine Intake
    ProvisionalChapter chapter 9 Toxic Effects as a Result of Herbal Medicine Intake Toxic Effects as a Result of Herbal Medicine Intake Nudrat Fatima and Naira Nayeem Nudrat Fatima and Naira Nayeem Additional information is available at the end of the chapter Additional information is available at the end of the chapter http://dx.doi.org/10.5772/64468 Abstract Concurrent use of herbs with therapeutic drugs increases the potential of herb-drug interactions. The clinical importance of herb-drug interactions is associated with the particular herb, drug, and patient profile. Herbs are potentially potent as they affect body functions. The use herbal medicine and supplements can be risky as they are not subject to review by the FDA. In this chapter, we make an attempt to discuss the possible reasons for toxic effects, types of toxicities, some reported cases of toxicities involving the use of herbal medicine alone, and some herb-drug interactions. In addition to this, possible ways to reduce toxic effects of herbal medicines have also been discussed. Keywords: herbal medicine, toxicity, reported cases, herb-drug interaction 1. Introduction Herbal medicines are advertised to be free from side effects, which is a myth. A large number of people still rely on herbal medicines, and some people take herbal medicines along with routine allopathic medicines especially in cases of diabetes, hypertension, thyroid disease, etc., where the patient is on long-term or lifelong treatment. Many commercial websites are available on Internet, which insist that herbal medicines have no side effects. In underdeveloped and developing countries, there are no specific laws for herbal practitioners and companies marketing herbal products.
    [Show full text]
  • Recessive Mutations of the Gene TRPM1 Abrogate on Bipolar Cell Function and Cause Complete Congenital Stationary Night Blindness in Humans
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector REPORT Recessive Mutations of the Gene TRPM1 Abrogate ON Bipolar Cell Function and Cause Complete Congenital Stationary Night Blindness in Humans Zheng Li,1 Panagiotis I. Sergouniotis,1 Michel Michaelides,1,2 Donna S. Mackay,1 Genevieve A. Wright,2 Sophie Devery,2 Anthony T. Moore,1,2 Graham E. Holder,1,2 Anthony G. Robson,1,2 and Andrew R. Webster1,2,* Complete congenital stationary night blindness (cCSNB) is associated with loss of function of rod and cone ON bipolar cells in the mammalian retina. In humans, mutations in NYX and GRM6 have been shown to cause the condition. Through the analysis of a consan- guineous family and screening of nine additional pedigrees, we have identified three families with recessive mutations in the gene TRPM1 encoding transient receptor potential cation channel, subfamily M, member 1, also known as melastatin. A number of other variants of unknown significance were found. All patients had myopia, reduced central vision, nystagmus, and electroretinographic evidence of ON bipolar cell dysfunction. None had abnormalities of skin pigmentation, although other skin conditions were reported. RNA derived from human retina and skin was analyzed and alternate 50 exons were determined. The most 50 exon is likely to harbor an initiation codon, and the protein sequence is highly conserved across vertebrate species. These findings suggest an important role of this specific cation channel for the normal function of ON bipolar cells in the human retina. Congenital stationary night blindness (CSNB) is a group of of the gene encoding transient receptor potential cation genetically determined, nondegenerative disorders of the channel, subfamily M, member 1 (TRPM1 [MIM *603576]) retina associated with lifelong deficient vision in the dark has been discovered in the skin and retina of horses homo- and often nystagmus and myopia.
    [Show full text]
  • Entropy-Based Regulation of Cluster Ion Channel Density
    www.nature.com/scientificreports OPEN Molecular and cellular correlates in Kv channel clustering: entropy‑based regulation of cluster ion channel density Limor Lewin1, Esraa Nsasra1, Ella Golbary1, Uzi Hadad2, Irit Orr1 & Ofer Yifrach1* Scafold protein-mediated ion channel clustering at unique membrane sites is important for electrical signaling. Yet, the mechanism(s) by which scafold protein-ion channel interactions lead to channel clustering or how cluster ion channel density is regulated is mostly not known. The voltage‑activated potassium channel (Kv) represents an excellent model to address these questions as the mechanism underlying its interaction with the post-synaptic density 95 (PSD-95) scafold protein is known to be controlled by the length of the extended ‘ball and chain’ sequence comprising the C-terminal channel region. Here, using sub-difraction high-resolution imaging microscopy, we show that Kv channel ‘chain’ length regulates Kv channel density with a ‘bell’-shaped dependence, refecting a balance between thermodynamic considerations controlling ‘chain’ recruitment by PSD-95 and steric hindrance due to the spatial proximity of multiple channel molecules. Our results thus reveal an entropy‑based mode of channel cluster density regulation that mirrors the entropy‑based regulation of the Kv channel-PSD-95 interaction. The implications of these fndings for electrical signaling are discussed. Action potential generation, propagation and the evoked synaptic potential all rely on precisely timed events associated with activation and inactivation gating transitions of voltage-dependent Na + and K + channels, clus- tered in multiple copies at unique membrane sites, such as the initial segment of an axon, nodes of Ranvier, pre-synaptic terminals or at the post-synaptic density (PSD)1–3.
    [Show full text]
  • Transient Receptor Potential (TRP) Channels in Haematological Malignancies: an Update
    biomolecules Review Transient Receptor Potential (TRP) Channels in Haematological Malignancies: An Update Federica Maggi 1,2 , Maria Beatrice Morelli 2 , Massimo Nabissi 2 , Oliviero Marinelli 2 , Laura Zeppa 2, Cristina Aguzzi 2, Giorgio Santoni 2 and Consuelo Amantini 3,* 1 Department of Molecular Medicine, Sapienza University, 00185 Rome, Italy; [email protected] 2 Immunopathology Laboratory, School of Pharmacy, University of Camerino, 62032 Camerino, Italy; [email protected] (M.B.M.); [email protected] (M.N.); [email protected] (O.M.); [email protected] (L.Z.); [email protected] (C.A.); [email protected] (G.S.) 3 Immunopathology Laboratory, School of Biosciences and Veterinary Medicine, University of Camerino, 62032 Camerino, Italy * Correspondence: [email protected]; Tel.: +30-0737403312 Abstract: Transient receptor potential (TRP) channels are improving their importance in differ- ent cancers, becoming suitable as promising candidates for precision medicine. Their important contribution in calcium trafficking inside and outside cells is coming to light from many papers published so far. Encouraging results on the correlation between TRP and overall survival (OS) and progression-free survival (PFS) in cancer patients are available, and there are as many promising data from in vitro studies. For what concerns haematological malignancy, the role of TRPs is still not elucidated, and data regarding TRP channel expression have demonstrated great variability throughout blood cancer so far. Thus, the aim of this review is to highlight the most recent findings Citation: Maggi, F.; Morelli, M.B.; on TRP channels in leukaemia and lymphoma, demonstrating their important contribution in the Nabissi, M.; Marinelli, O.; Zeppa, L.; perspective of personalised therapies.
    [Show full text]
  • Functional P53 Is Required for Triptolide-Induced Apoptosis and AP-1 and Nuclear Factor-Κb Activation in Gastric Cancer Cells
    Oncogene (2001) 20, 8009 ± 8018 ã 2001 Nature Publishing Group All rights reserved 0950 ± 9232/01 $15.00 www.nature.com/onc Functional p53 is required for triptolide-induced apoptosis and AP-1 and nuclear factor-kB activation in gastric cancer cells Xiao-Hua Jiang1,2,5, Benjamin Chun-Yu Wong*,2,5, Marie Chia-Mi Lin3, Geng-Hui Zhu2, Hsiang-Fu Kung3, Shi-Hu Jiang1, Dan Yang4 and Shiu-Kum Lam2 1Department of Gastroenterology, Rui-jin Hospital, Shanghai, Peoples Republic of China; 2Department of Medicine, University of Hong Kong, Hong Kong; 3Institute of Molecular Biology, University of Hong Kong, Hong Kong; 4Department of Chemistry, University of Hong Kong, Hong Kong Triptolide, a major component in the extract of Chinese Introduction herbal plant Tripterygium wilfordii Hook f (TWHf), has potential anti-neoplastic eect. In the present study we Tripterygium wilfordii Hook f (TWHf) has been used investigated the potential therapeutic eects and mechan- in traditional Chinese medicine for centuries. Its crude isms of triptolide against human gastric cancer cells. extracts continue to be used to treat a variety of Four gastric cancer cell lines with dierent p53 status, autoimmune diseases, such as rheumatoid arthritis, AGS and MKN-45 (wild type p53); MKN-28 and SGC- nephritis, and systemic lupus erythematosus (Qin et al., 7901 (mutant p53) were observed as to cell growth 1981; Tao et al., 1989; Jiang et al., 1994). It has been inhibition and induction of apoptosis in response to suggested that the major therapeutic eects of TWHf triptolide treatment. We showed that triptolide inhibited are from ingredients such as triptolide, tripdiolide, cell growth, induced apoptosis and suppressed NK-kB triptonide, and triptophenolide (Zhang et al., 1990).
    [Show full text]
  • Molecular Characterization of TRPA Subfamily Genes and Function in Temperature Preference in Tuta Absoluta (Meyrick) (Lepidoptera: Gelechiidae)
    International Journal of Molecular Sciences Article Molecular Characterization of TRPA Subfamily Genes and Function in Temperature Preference in Tuta absoluta (Meyrick) (Lepidoptera: Gelechiidae) Xiao-Di Wang 1, Ze-Kai Lin 1 , Shun-Xia Ji 1, Si-Yan Bi 1, Wan-Xue Liu 1, Gui-Fen Zhang 1, Fang-Hao Wan 1,2 and Zhi-Chuang Lü 1,* 1 State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China; [email protected] (X.-D.W.); [email protected] (Z.-K.L.); [email protected] (S.-X.J.); [email protected] (S.-Y.B.); [email protected] (W.-X.L.); [email protected] (G.-F.Z.); [email protected] (F.-H.W.) 2 Agricultural Genome Institute at Shenzhen, Chinese Academy of Agricultural Sciences, Shenzhen 518120, China * Correspondence: [email protected]; Tel.: +86-10-8210-9572 Abstract: To reveal the mechanism of temperature preference in Tuta absoluta, one of the top 20 plant pests in the world, we cloned and identified TaTRPA1, TaPain, and TaPyx genes by RACE and bioinformatic analysis, and clarified their expression profiles during different development stages using real-time PCR, and revealed their function in preference temperature by RNAi. The full-length cDNA of TaPain was 3136 bp, with a 2865-bp open reading frame encoding a 259.89-kDa Citation: Wang, X.-D.; Lin, Z.-K.; Ji, protein; and the partial length cDNA of TaPyx was 2326-bp, with a 2025-bp open reading frame S.-X.; Bi, S.-Y.; Liu, W.-X.; Zhang, G.-F.; encoding a 193.16-kDa protein.
    [Show full text]
  • Structure of the Mouse TRPC4 Ion Channel 1 2 Jingjing
    bioRxiv preprint doi: https://doi.org/10.1101/282715; this version posted March 15, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 1 Structure of the mouse TRPC4 ion channel 2 3 Jingjing Duan1,2*, Jian Li1,3*, Bo Zeng4*, Gui-Lan Chen4, Xiaogang Peng5, Yixing Zhang1, 4 Jianbin Wang1, David E. Clapham2, Zongli Li6#, Jin Zhang1# 5 6 7 1School of Basic Medical Sciences, Nanchang University, Nanchang, Jiangxi, 330031, China. 8 2Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, VA 20147, USA 9 3Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, KY 10 40536, USA. 11 4Key Laboratory of Medical Electrophysiology, Ministry of Education, and Institute of 12 Cardiovascular Research, Southwest Medical University, Luzhou, Sichuan, 646000, China 13 5The Key Laboratory of Molecular Medicine, the Second Affiliated Hospital of Nanchang 14 University, Nanchang 330006, China. 15 6Howard Hughes Medical Institute, Department of Biological Chemistry and Molecular 16 Pharmacology, Harvard Medical School, Boston, MA 02115, USA. 17 18 *These authors contributed equally to this work. 19 # Corresponding authors bioRxiv preprint doi: https://doi.org/10.1101/282715; this version posted March 15, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 20 Abstract 21 Members of the transient receptor potential (TRP) ion channels conduct cations into cells. They 22 mediate functions ranging from neuronally-mediated hot and cold sensation to intracellular 23 organellar and primary ciliary signaling.
    [Show full text]