DOCSLIB.ORG

Deficiency of Inositol Monophosphatase Activity Decreases Phosphoinositide Lipids and Enhances TRPV1 Function in Vivo

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Deficiency of Inositol Monophosphatase Activity Decreases Phosphoinositide Lipids and Enhances TRPV1 Function in Vivo Research Articles: Cellular/Molecular Deficiency of Inositol Monophosphatase Activity Decreases Phosphoinositide Lipids and Enhances TRPV1 Function in vivo https://doi.org/10.1523/JNEUROSCI.0803-20.2020 Cite as: J. Neurosci 2020; 10.1523/JNEUROSCI.0803-20.2020 Received: 6 April 2020 Revised: 12 November 2020 Accepted: 19 November 2020 This Early Release article has been peer-reviewed and accepted, but has not been through the composition and copyediting processes. The final version may differ slightly in style or formatting and will contain links to any extended data. Alerts: Sign up at www.jneurosci.org/alerts to receive customized email alerts when the fully formatted version of this article is published. Copyright © 2020 Caires et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license, which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed. 1 Deficiency of Inositol Monophosphatase Activity Decreases Phosphoinositide Lipids 2 and Enhances TRPV1 Function in vivo 3 Abbreviated title: Phosphoinositide Lipids Decrease TRPV1 function in vivo 4 5 Rebeca Caires1, Briar Bell1, 2,A, Jungsoo Lee1,A, Luis O. Romero1, 2, Valeria Vásquez1,*, Julio F. 6 Cordero-Morales1,*. A These authors contributed equally. 7 8 1Department of Physiology, College of Medicine, University of Tennessee Health Science Center, 9 Memphis, TN 38163, USA 10 11 2 Integrated Biomedical Sciences Graduate Program, College of Graduate Health Sciences, University of 12 Tennessee Health Science Center, Memphis, TN 38163, USA 13 14 *Corresponding Authors 15 Julio F. Cordero-Morales (JFC-M) Valeria Vásquez (VV) 16 Email: [email protected] Email: [email protected] 17 18 Ten figures 19 Abstract: 155 words; Introduction: 1388 words: Discussion: 907 words 20 21 Competing interests: The authors declare no competing financial interests. 22 Acknowledgements: The authors thank Dr. Avi Priel for kindly providing purified DkTx, and Dr. E. 23 Cao for critically reading the manuscript, and Dr. C. Bargmann for providing worm strains. Channel 24 constructs and worm strains DNA were sequenced at the Molecular Resource Center of Excellence at 25 The University of Tennessee Health Science Center with the assistance of Dr. Tom Cunningham. Lipid 26 analyses were provided by the Mass Spectrometry Lipidomics Core Facility at Wayne State University 27 (supported, in part, by the National Center for Research Resources, National Institutes of Health grant 28 S10RR027926) and the University of Colorado Anschutz Medical Campus in Aurora, Colorado. 29 Funding: This work was supported by the National Institutes of Health (R01GM133845 to V.V. and 30 R01GM125629 to J.F.C.-M.), the United States-Israel Binational Science Foundation (2015221 to 31 V.V.), the American Heart Association (16SDG26700010 to V.V. and 15SDG25700146 to J.F.C.-M.), 32 and the Neuroscience Institute at UTHSC (Postdoctoral Matching Salary Support to R.C. and J.L.). 33 34 Author contributions 35 Conceptualization, JFC-M; Methodology, JFC-M and VV; Investigation, VV, RC, BB, JL, and LOR; 36 Writing, JFC-M and VV; Supervision, JFC-M and VV. 37 38 Abstract 39 Membrane remodeling by inflammatory mediators influences the function of sensory ion channels. The 40 capsaicin- and heat-activated TRPV1 channel contributes to neurogenic inflammation and pain 41 hypersensitivity, in part due to its potentiation downstream of phospholipase C-coupled receptors that 42 regulate phosphoinositide lipid content. Here, we determined the effect of phosphoinositide lipids on 43 TRPV1 function by combining genetic dissection, diet supplementation, behavioral, biochemical, and 44 functional analyses in Caenorhabditis elegans. As capsaicin elicits hot and pain sensation in mammals, 45 transgenic TRPV1 worms exhibit an aversive response to capsaicin. TRPV1 worms with low levels of 46 phosphoinositide lipids display an enhanced response to capsaicin, whereas phosphoinositide lipid 47 supplementation reduces TRPV1-mediated responses. A worm carrying a TRPV1 construct lacking the 48 distal C-terminal domain features an enhanced response to capsaicin, independent of the 49 phosphoinositide lipid content. Our results demonstrate that TRPV1 activity is enhanced when the 50 phosphoinositide lipid content is reduced, and the C-terminal domain is key to determining agonist 51 response in vivo. 52 53 54 55 56 57 58 59 60 61 62 1 63 64 65 Significance Statement 66 TRPV1 is an essential protein for the mechanism whereby noxious stimuli, such as high temperatures 67 and chemicals, cause pain. TRPV1 undergoes sensitization, a process in which inflammatory molecules 68 enhance its response to other stimuli, thereby promoting pain hypersensitivity. Proalgesic agents 69 produced in response to tissue injury activate PLC-coupled receptors and alter the membrane 70 phosphoinositide lipid content. The mechanism by which phosphoinositide lipids modulate TRPV1 71 function has remained controversial. Determining whether membrane phosphoinositides are positive or 72 negative regulators of TRPV1 function is critical for developing therapeutic strategies to ameliorate 73 TRPV1-mediated inflammatory pain. We address the role of phosphoinositide lipids on TRPV1 function 74 using an in vivo approach and report that phosphoinositide lipids reduce TRPV1 activity in vivo. 75 76 77 78 79 80 81 82 83 84 85 86 87 2 88 89 90 Introduction 91 The transient receptor potential vanilloid 1 (TRPV1) is a polymodal ion channel activated by 92 noxious heat (Caterina et al., 1997), capsaicin (Caterina et al., 1997), animal toxins (Siemens et al., 93 2006; Bohlen et al., 2010; Yang et al., 2015), extracellular protons (Jordt et al., 2000), and 94 lysophosphatidic acid (Nieto-Posadas et al., 2012). TRPV1 is modulated by proalgesic inflammatory 95 agents, such as bradykinin (Chuang et al., 2001), nerve growth factor (Lewin et al., 1993; Chuang et al., 96 2001; Stratiievska et al., 2018), and bioactive lipids (Zygmunt et al., 1999; Prescott and Julius, 2003; 97 Cao et al., 2013b) produced in response to tissue injury. TRPV1 is expressed in nociceptors, which are 98 specialized peripheral sensory neurons that detect and respond to a variety of potentially harmful 99 stimuli, including extreme temperatures, pressure, and chemicals (Basbaum et al., 2009). TRPV1 100 undergoes sensitization, a process in which inflammatory agents enhance its response to other stimuli, 101 thereby promoting pain hypersensitivity (Basbaum et al., 2009). Sensitization is an intrinsic feature of 102 TRPV1 since protons (e.g., pH 6), fatty acids (e.g., arachidonic acid), and the lack of phosphoinositide 103 lipids produce a leftward shift in the thermal and chemical response profile of TRPV1-containing 104 liposomes (Cao et al., 2013b). However, TRPV1 is also indirectly sensitized through the activation of 105 phospholipase C (PLC)-coupled receptors by a variety of proalgesic agents (e.g., bradykinin) and the 106 subsequent hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) (Chuang et al., 2001) and/or 107 phosphorylation via protein kinase C (Vellani et al., 2001; Numazaki et al., 2002; Bhave et al., 2003). It 108 has been reported that reduction of plasma membrane PIP2, via PLC-mediated hydrolysis, sensitizes 109 TRPV1, mimicking the effect observed with proalgesic agents such as bradykinin and nerve growth 110 factor (Chuang et al., 2001; Prescott and Julius, 2003). However, later work demonstrated that TRPV1 is 111 also modulated by other phosphoinositide lipids, including PI, PI3P, PI4P, and PI5P (Lukacs et al., 112 2007; Klein et al., 2008; Cao et al., 2013b). Hence, particular attention has been given to the mechanism 3 113 by which this lipid class modulates channel function (Rohacs, 2015). Defining the mechanisms through 114 which bioactive lipids modulate TRPV1 will help elucidate its role in controlling sensory neuron 115 excitability. 116 Several lines of evidence are consistent with the idea that phosphoinositides negatively regulate 117 channel function. At low capsaicin concentration and moderate heat, it has been shown that a soluble 118 PIP2 partially inhibits TRPV1 in a whole-cell environment (Lukacs et al., 2007). Positively-charged 119 residues (lysine/arginine) within the distal TRPV1 C-terminal domain (L777–L792 in rat TRPV1) have 120 been identified as the site required for phosphoinositide lipid-mediated inhibition, and mutations that 121 eliminate these residues enhance channel heat response (Prescott and Julius, 2003). Interestingly, a 122 vampire bat TRPV1 splice variant lacking the distal C-terminal domain (putative phosphoinositide- 123 interaction site) displayed enhanced sensitivity to thermal stimuli (Gracheva et al., 2011). These results 124 are further supported by experiments in which TRPV1 displays enhanced sensitivity to thermal and 125 chemical stimuli when reconstituted into liposomes without phosphoinositide lipids (Cao et al., 2013b). 126 Furthermore, incorporation of phosphoinositide lipids (e.g., PI, PI3P, PI4P, PI5P, and PIP2) in the 127 proteoliposomes produces a rightward shift (i.e., decreased response) in the channel’s agonist response 128 profile. Likewise, stabilization of the TRPV1 C terminus-membrane interaction, using nitrilotriacetic 129 acid (NTA)-modified lipid (DGS-NTA) in a nickel-dependent manner, displays a TRPV1 agonist 130 response comparable to those observed in the presence of phosphoinositide lipids (Cao et
Load more

Recommended publications
  • Phosphorylation of Synaptojanin Differentially Regulates Endocytosis of Functionally Distinct Synaptic Vesicle Pools
    8882 • The Journal of Neuroscience, August 24, 2016 • 36(34):8882–8894 Cellular/Molecular Phosphorylation of Synaptojanin Differentially Regulates Endocytosis of Functionally Distinct Synaptic Vesicle Pools X Junhua Geng,1* Liping Wang,1,2* Joo Yeun Lee,1,4 XChun-Kan Chen,1 and Karen T. Chang1,3,4 1Zilkha Neurogenetic Institute, 2Department of Biochemistry and Molecular Biology, and 3Department of Cell and Neurobiology, Keck School of Medicine, University of Southern California, Los Angeles, California 90089, and 4Neuroscience Graduate Program, University of Southern California, Los Angeles, California 90089 The rapid replenishment of synaptic vesicles through endocytosis is crucial for sustaining synaptic transmission during intense neuronal activity. Synaptojanin (Synj), a phosphoinositide phosphatase, is known to play an important role in vesicle recycling by promoting the uncoating of clathrin following synaptic vesicle uptake. Synj has been shown to be a substrate of the minibrain (Mnb) kinase, a fly homolog of the dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A); however, the functional impacts of Synj phosphorylation by Mnb are not well understood. Here we identify that Mnb phosphorylates Synj at S1029 in Drosophila. We find that phosphorylation of Synj at S1029 enhances Synj phosphatase activity, alters interaction between Synj and endophilin, and promotes efficient endocytosis of the active cycling vesicle pool (also referred to as exo-endo cycling pool) at the expense of reserve pool vesicle endocytosis. Dephosphorylated Synj, on the other hand, is deficient in the endocytosis of the active recycling pool vesicles but maintains reserve pool vesicle endocytosis to restore total vesicle pool size and sustain synaptic transmission. Together, our findings reveal a novel role for Synj in modulating reserve pool vesicle endocytosis and further indicate that dynamic phosphorylation and dephosphorylation of Synj differentially maintain endocytosis of distinct functional synaptic vesicle pools.
    [Show full text]
  • Gene Targeting Therapies (Roy Alcalay)
    Recent Developments in Gene - Targeted Therapies for Parkinson’s Disease Roy Alcalay, MD, MS Alfred and Minnie Bressler Associate Professor of Neurology Division of Movement Disorders Columbia University Medical Center Disclosures Funding: Dr. Alcalay is funded by the National Institutes of Health, the DOD, the Michael J. Fox Foundation and the Parkinson’s Foundation. Dr. Alcalay receives consultation fees from Genzyme/Sanofi, Restorbio, Janssen, and Roche. Gene Localizations Identified in PD Gene Symbol Protein Transmission Chromosome PARK1 SNCA α-synuclein AD 4q22.1 PARK2 PRKN parkin (ubiquitin ligase) AR 6q26 PARK3 ? ? AD 2p13 PARK4 SNCA triplication α-synuclein AD 4q22.1 PARK5 UCH-L1 ubiquitin C-terminal AD 4p13 hydrolase-L1 PARK6 PINK1 PTEN-induced kinase 1 AR 1p36.12 PARK7 DJ-1 DJ-1 AR 1p36.23 PARK8 LRRK2 leucine rich repeat kinase 2 AD 12q12 PARK9 ATP13A2 lysosomal ATPase AR 1p36.13 PARK10 ? ? (Iceland) AR 1p32 PARK11 GIGYF2 GRB10-interacting GYF protein 2 AD 2q37.1 PARK12 ? ? X-R Xq21-q25 PARK13 HTRA2 serine protease AD 2p13.1 PARK14 PLA2G6 phospholipase A2 (INAD) AR 22q13.1 PARK15 FBXO7 F-box only protein 7 AR 22q12.3 PARK16 ? Discovered by GWAS ? 1q32 PARK17 VPS35 vacuolar protein sorting 35 AD 16q11.2 PARK18 EIF4G1 initiation of protein synth AD 3q27.1 PARK19 DNAJC6 auxilin AR 1p31.3 PARK20 SYNJ1 synaptojanin 1 AR 21q22.11 PARK21 DNAJC13 8/RME-8 AD 3q22.1 PARK22 CHCHD2 AD 7p11.2 PARK23 VPS13C AR 15q22 Gene Localizations Identified in PD Disorder Symbol Protein Transmission Chromosome PD GBA β-glucocerebrosidase AD 1q21 SCA2
    [Show full text]
  • 1 Metabolic Dysfunction Is Restricted to the Sciatic Nerve in Experimental
    Page 1 of 255 Diabetes Metabolic dysfunction is restricted to the sciatic nerve in experimental diabetic neuropathy Oliver J. Freeman1,2, Richard D. Unwin2,3, Andrew W. Dowsey2,3, Paul Begley2,3, Sumia Ali1, Katherine A. Hollywood2,3, Nitin Rustogi2,3, Rasmus S. Petersen1, Warwick B. Dunn2,3†, Garth J.S. Cooper2,3,4,5* & Natalie J. Gardiner1* 1 Faculty of Life Sciences, University of Manchester, UK 2 Centre for Advanced Discovery and Experimental Therapeutics (CADET), Central Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Sciences Centre, Manchester, UK 3 Centre for Endocrinology and Diabetes, Institute of Human Development, Faculty of Medical and Human Sciences, University of Manchester, UK 4 School of Biological Sciences, University of Auckland, New Zealand 5 Department of Pharmacology, Medical Sciences Division, University of Oxford, UK † Present address: School of Biosciences, University of Birmingham, UK *Joint corresponding authors: Natalie J. Gardiner and Garth J.S. Cooper Email: [email protected]; [email protected] Address: University of Manchester, AV Hill Building, Oxford Road, Manchester, M13 9PT, United Kingdom Telephone: +44 161 275 5768; +44 161 701 0240 Word count: 4,490 Number of tables: 1, Number of figures: 6 Running title: Metabolic dysfunction in diabetic neuropathy 1 Diabetes Publish Ahead of Print, published online October 15, 2015 Diabetes Page 2 of 255 Abstract High glucose levels in the peripheral nervous system (PNS) have been implicated in the pathogenesis of diabetic neuropathy (DN). However our understanding of the molecular mechanisms which cause the marked distal pathology is incomplete. Here we performed a comprehensive, system-wide analysis of the PNS of a rodent model of DN.
    [Show full text]
  • Yeast Genome Gazetteer P35-65
    gazetteer Metabolism 35 tRNA modification mitochondrial transport amino-acid metabolism other tRNA-transcription activities vesicular transport (Golgi network, etc.) nitrogen and sulphur metabolism mRNA synthesis peroxisomal transport nucleotide metabolism mRNA processing (splicing) vacuolar transport phosphate metabolism mRNA processing (5’-end, 3’-end processing extracellular transport carbohydrate metabolism and mRNA degradation) cellular import lipid, fatty-acid and sterol metabolism other mRNA-transcription activities other intracellular-transport activities biosynthesis of vitamins, cofactors and RNA transport prosthetic groups other transcription activities Cellular organization and biogenesis 54 ionic homeostasis organization and biogenesis of cell wall and Protein synthesis 48 plasma membrane Energy 40 ribosomal proteins organization and biogenesis of glycolysis translation (initiation,elongation and cytoskeleton gluconeogenesis termination) organization and biogenesis of endoplasmic pentose-phosphate pathway translational control reticulum and Golgi tricarboxylic-acid pathway tRNA synthetases organization and biogenesis of chromosome respiration other protein-synthesis activities structure fermentation mitochondrial organization and biogenesis metabolism of energy reserves (glycogen Protein destination 49 peroxisomal organization and biogenesis and trehalose) protein folding and stabilization endosomal organization and biogenesis other energy-generation activities protein targeting, sorting and translocation vacuolar and lysosomal
    [Show full text]
  • Supplemental Information
    Supplemental information Dissection of the genomic structure of the miR-183/96/182 gene. Previously, we showed that the miR-183/96/182 cluster is an intergenic miRNA cluster, located in a ~60-kb interval between the genes encoding nuclear respiratory factor-1 (Nrf1) and ubiquitin-conjugating enzyme E2H (Ube2h) on mouse chr6qA3.3 (1). To start to uncover the genomic structure of the miR- 183/96/182 gene, we first studied genomic features around miR-183/96/182 in the UCSC genome browser (http://genome.UCSC.edu/), and identified two CpG islands 3.4-6.5 kb 5’ of pre-miR-183, the most 5’ miRNA of the cluster (Fig. 1A; Fig. S1 and Seq. S1). A cDNA clone, AK044220, located at 3.2-4.6 kb 5’ to pre-miR-183, encompasses the second CpG island (Fig. 1A; Fig. S1). We hypothesized that this cDNA clone was derived from 5’ exon(s) of the primary transcript of the miR-183/96/182 gene, as CpG islands are often associated with promoters (2). Supporting this hypothesis, multiple expressed sequences detected by gene-trap clones, including clone D016D06 (3, 4), were co-localized with the cDNA clone AK044220 (Fig. 1A; Fig. S1). Clone D016D06, deposited by the German GeneTrap Consortium (GGTC) (http://tikus.gsf.de) (3, 4), was derived from insertion of a retroviral construct, rFlpROSAβgeo in 129S2 ES cells (Fig. 1A and C). The rFlpROSAβgeo construct carries a promoterless reporter gene, the β−geo cassette - an in-frame fusion of the β-galactosidase and neomycin resistance (Neor) gene (5), with a splicing acceptor (SA) immediately upstream, and a polyA signal downstream of the β−geo cassette (Fig.
    [Show full text]
  • 140503 IPF Signatures Supplement Withfigs Thorax
    Supplementary material for Heterogeneous gene expression signatures correspond to distinct lung pathologies and biomarkers of disease severity in idiopathic pulmonary fibrosis Daryle J. DePianto1*, Sanjay Chandriani1⌘*, Alexander R. Abbas1, Guiquan Jia1, Elsa N. N’Diaye1, Patrick Caplazi1, Steven E. Kauder1, Sabyasachi Biswas1, Satyajit K. Karnik1#, Connie Ha1, Zora Modrusan1, Michael A. Matthay2, Jasleen Kukreja3, Harold R. Collard2, Jackson G. Egen1, Paul J. Wolters2§, and Joseph R. Arron1§ 1Genentech Research and Early Development, South San Francisco, CA 2Department of Medicine, University of California, San Francisco, CA 3Department of Surgery, University of California, San Francisco, CA ⌘Current address: Novartis Institutes for Biomedical Research, Emeryville, CA. #Current address: Gilead Sciences, Foster City, CA. *DJD and SC contributed equally to this manuscript §PJW and JRA co-directed this project Address correspondence to Paul J. Wolters, MD University of California, San Francisco Department of Medicine Box 0111 San Francisco, CA 94143-0111 [email protected] or Joseph R. Arron, MD, PhD Genentech, Inc. MS 231C 1 DNA Way South San Francisco, CA 94080 [email protected] 1 METHODS Human lung tissue samples Tissues were obtained at UCSF from clinical samples from IPF patients at the time of biopsy or lung transplantation. All patients were seen at UCSF and the diagnosis of IPF was established through multidisciplinary review of clinical, radiological, and pathological data according to criteria established by the consensus classification of the American Thoracic Society (ATS) and European Respiratory Society (ERS), Japanese Respiratory Society (JRS), and the Latin American Thoracic Association (ALAT) (ref. 5 in main text). Non-diseased normal lung tissues were procured from lungs not used by the Northern California Transplant Donor Network.
    [Show full text]
  • Regulation of Synaptojanin 1 by Cyclin-Dependent Kinase 5 at Synapses
    Regulation of synaptojanin 1 by cyclin-dependent kinase 5 at synapses Sang Yoon Lee*†, Markus R. Wenk*†, Yong Kim‡, Angus C. Nairn§, and Pietro De Camilli*†¶ Departments of *Cell Biology and §Psychiatry and †Howard Hughes Medical Institute, Yale University School of Medicine, 295 Congress Avenue, New Haven, CT 06510; and ‡Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University, New York, NY 10021 Contributed by Pietro De Camilli, November 24, 2003 Synaptojanin 1 is a polyphosphoinositide phosphatase concen- substrates in vitro include PI 4-phosphate [PI(4)P] (15, 25, 26). trated in presynaptic nerve terminals, where it dephosphorylates The PRD region functions primarily as a targeting domain via a pool of phosphatidylinositol 4,5-bisphosphate implicated in syn- the interaction with a large number of proteins, among which aptic vesicle recycling. Like other proteins with a role in endocy- endophilin 1, via its COOH-terminal Src homology 3 (SH3) tosis, synaptojanin 1 undergoes constitutive phosphorylation in domain, appears to be the most important physiological partner resting synapses and stimulation-dependent dephosphorylation (9, 27, 28). In neurons of synaptojanin 1 knockout mice, which by calcineurin. Here, we show that cyclin-dependent kinase 5 contain elevated levels of PI(4,5)P2, an abnormal accumulation (Cdk5) phosphorylates synaptojanin 1 and regulates its function of coated vesicles within a cytoskeletal cytomatrix was observed both in vitro and in intact synaptosomes. Cdk5 phosphorylation at endocytic zones of nerve terminals (16). Similar observations inhibited the inositol 5-phosphatase activity of synaptojanin 1, were made after disruption of synaptojanin function by antibody whereas dephosphorylation by calcineurin stimulated such activ- and peptide microinjection into giant lamprey synapses (17), or ity.
    [Show full text]
  • Identifying New Substrates and Functions for an Old Enzyme: Calcineurin
    Downloaded from http://cshperspectives.cshlp.org/ on October 4, 2021 - Published by Cold Spring Harbor Laboratory Press Identifying New Substrates and Functions for an Old Enzyme: Calcineurin Jagoree Roy and Martha S. Cyert Department of Biology, Stanford University, Stanford, California 94305-5020 Correspondence: [email protected] Biological processes are dynamically regulated by signaling networks composed of protein kinases and phosphatases. Calcineurin, or PP3, is a conserved phosphoserine/phospho- threonine-specific protein phosphatase and member of the PPP family of phosphatases. Calcineurin is unique, however, in its activation by Ca2+ and calmodulin. This ubiquitously expressed phosphatase controls Ca2+-dependent processes in all human tissues, but is best known for driving the adaptive immune response by dephosphorylating the nuclear factor of the activated T-cells (NFAT) family of transcription factors. Therefore, calcineurin inhibi- tors, FK506 (tacrolimus), and cyclosporin A serve as immunosuppressants. We describe some of the adverse effects associated with calcineurin inhibitors that result from inhibition of calcineurin in nonimmune tissues, illustrating the many functions of this enzyme that have yet to be elucidated. In fact, calcineurin has essential roles beyond the immune system, from yeast to humans, but since its discovery more than 30 years ago, only a small number of direct calcineurin substrates have been shown (∼75 proteins). This is because of limitations in current methods for identification of phosphatase substrates. Here we discuss recent insights into mechanisms of calcineurin activation and substrate recognition that have been critical in the development of novel approaches for identifying its targets systematically. Rather than comprehensively reviewing known functions of calcineurin, we highlight new approaches to substrate identification for this critical regulator that may reveal molecular mechanisms un- derlying toxicities caused by calcineurin inhibitor-based immunosuppression.
    [Show full text]
  • Human Induced Pluripotent Stem Cell–Derived Podocytes Mature Into Vascularized Glomeruli Upon Experimental Transplantation
    BASIC RESEARCH www.jasn.org Human Induced Pluripotent Stem Cell–Derived Podocytes Mature into Vascularized Glomeruli upon Experimental Transplantation † Sazia Sharmin,* Atsuhiro Taguchi,* Yusuke Kaku,* Yasuhiro Yoshimura,* Tomoko Ohmori,* ‡ † ‡ Tetsushi Sakuma, Masashi Mukoyama, Takashi Yamamoto, Hidetake Kurihara,§ and | Ryuichi Nishinakamura* *Department of Kidney Development, Institute of Molecular Embryology and Genetics, and †Department of Nephrology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan; ‡Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Hiroshima, Japan; §Division of Anatomy, Juntendo University School of Medicine, Tokyo, Japan; and |Japan Science and Technology Agency, CREST, Kumamoto, Japan ABSTRACT Glomerular podocytes express proteins, such as nephrin, that constitute the slit diaphragm, thereby contributing to the filtration process in the kidney. Glomerular development has been analyzed mainly in mice, whereas analysis of human kidney development has been minimal because of limited access to embryonic kidneys. We previously reported the induction of three-dimensional primordial glomeruli from human induced pluripotent stem (iPS) cells. Here, using transcription activator–like effector nuclease-mediated homologous recombination, we generated human iPS cell lines that express green fluorescent protein (GFP) in the NPHS1 locus, which encodes nephrin, and we show that GFP expression facilitated accurate visualization of nephrin-positive podocyte formation in
    [Show full text]
  • Chemistry-To-Gene Screens in Caenorhabditis Elegans
    REVIEWS CHEMISTRY-TO-GENE SCREENS IN CAENORHABDITIS ELEGANS Andrew K. Jones, Steven D. Buckingham and David B. Sattelle Abstract | The nematode worm Caenorhabditis elegans is a genetic model organism linked to an impressive portfolio of fundamental discoveries in biology. This free-living nematode, which can be easily and inexpensively grown in the laboratory, is also a natural vehicle for screening for drugs that are active against nematode parasites. Here, we show that chemistry-to-gene screens using this animal model can define targets of antiparasitic drugs, identify novel candidate drug targets and contribute to the discovery of new drugs for treating human diseases. MODEL ORGANISMS Nematode parasites cause substantial mortality and Discovering a novel target can offer considerably higher ANTHELMINTIC A chemical that controls morbidity in the human population throughout the commercial returns than developing a known target, parasitic worms. An example tropical and sub-tropical world. The main species but such discovery in the field of endoparasites is hin- is levamisole, which controls involved — Ascaris lumbricoides (intestinal round- dered by difficulties in culturing parasites outside a parasitic nematodes. worms), Trichuris trichiura (whip worms) and several host. The free-living nematode worm Caenorhabditis ENDECTOCIDE species of hookworms — each currently infects around elegans,whose genome has been fully sequenced, is well 1 An antiparasitic drug which a billion people worldwide .Parasitic nematodes also suited for mutagenesis studies, lends itself to high- controls both internal parasites cause considerable losses in livestock and domestic throughput assays, and shares some physiological and (such as worms) and external animals, and are responsible for more than US$100 pharmacological characteristics with parasitic nema- parasites (such as ticks).
    [Show full text]
  • 11056.Full.Pdf
    11056 • The Journal of Neuroscience, October 10, 2007 • 27(41):11056–11064 Neurobiology of Disease Huntingtin-Interacting Protein 1 Influences Worm and Mouse Presynaptic Function and Protects Caenorhabditis elegans Neurons against Mutant Polyglutamine Toxicity J. Alex Parker,1,2 Martina Metzler,3 John Georgiou,4 Marilyne Mage,1,2 John C. Roder,4,5 Ann M. Rose,6 Michael R. Hayden,3 and Christian Ne´ri1,2 1Inserm, Unit 857 “Neuronal Cell Biology and Pathology,” and 2University of Paris Rene´ Descartes, Equipe d’Accueil 4059, Centre Paul Broca, 75014 Paris, France, 3Centre for Molecular Medicine and Therapeutics, Child and Family Research Institute, Department of Medical Genetics, University of British Columbia, Vancouver, British Columbia, Canada V5Z 4H4, 4Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada M5G 1X5, 5Department of Physiology, University of Toronto, Toronto, Ontario, Canada M5S 1A8, and 6Department of Medical Genetics, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z3 Huntingtin-interacting protein 1 (HIP1) was identified through its interaction with htt (huntingtin), the Huntington’s disease (HD) protein. HIP1 is an endocytic protein that influences transport and function of AMPA and NMDA receptors in the brain. However, little is known about its contribution to neuronal dysfunction in HD. We report that the Caenorhabditis elegans HIP1 homolog hipr-1 modulates presynaptic activity and the abundance of synaptobrevin, a protein involved in synaptic vesicle fusion. Presynaptic function was also altered in hippocampal brain slices of HIP1 Ϫ/Ϫ mice dem- onstrating delayed recovery from synaptic depression and a reduction in paired-pulse facilitation, a form of presynaptic plasticity.
    [Show full text]
  • The Lipid Phosphatase Synaptojanin 1 Undergoes a Significant Alteration In
    Ando et al. Acta Neuropathologica Communications (2020) 8:79 https://doi.org/10.1186/s40478-020-00954-1 RESEARCH Open Access The lipid phosphatase Synaptojanin 1 undergoes a significant alteration in expression and solubility and is associated with brain lesions in Alzheimer’s disease Kunie Ando1,2,3*, Marième Ndjim3, Sabrina Turbant2,3, Gaëlle Fontaine3, Gustavo Pregoni3, Luce Dauphinot3, Zehra Yilmaz1, Valérie Suain1, Salwa Mansour1, Michèle Authelet1, Robert De Dekker1, Karelle Leroy1, Benoît Delatour3, Brain Bank NeuroCEB Neuropathology Network, Charles Duyckaerts2,3, Marie-Claude Potier3† and Jean-Pierre Brion1*† Abstract Synaptojanin 1 (SYNJ1) is a brain-enriched lipid phosphatase critically involved in autophagosomal/endosomal trafficking, synaptic vesicle recycling and metabolism of phosphoinositides. Previous studies suggest that SYNJ1 polymorphisms have significant impact on the age of onset of Alzheimer’s disease (AD) and that SYNJ1 is involved in amyloid-induced toxicity. Yet SYNJ1 protein level and cellular localization in post-mortem human AD brain tissues have remained elusive. This study aimed to examine whether SYNJ1 localization and expression are altered in post- mortem AD brains. We found that SYNJ1 is accumulated in Hirano bodies, plaque-associated dystrophic neurites and some neurofibrillary tangles (NFTs). SYNJ1 immunoreactivity was higher in neurons and in the senile plaques in AD patients carrying one or two ApolipoproteinE (APOE) ε4 allele(s). In two large cohorts of APOE-genotyped controls and AD patients, SYNJ1 transcripts were significantly increased in AD temporal isocortex compared to control. There was a significant increase in SYNJ1 transcript in APOEε4 carriers compared to non-carriers in AD cohort. SYNJ1 was systematically co-enriched with PHF-tau in the sarkosyl-insoluble fraction of AD brain.
    [Show full text]