Proteolytic and cellular mechanisms in prohormone and proprotein processing 1570595534

M OLECULAR BIOLOGY I N T E L L I G E N C E U N I T Vivian Y.H Hook Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing R.G LANDES C O M P A N Y MOLECULAR BIOLOGY INTELLIGENCE UNIT Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing Vivian Y.H Hook Department of Medicine University of California, San Diego La Jolla, California, U.S.A R.G LANDES COMPANY AUSTIN, TEXAS U.S.A MOLECULAR BIOLOGY INTELLIGENCE UNIT Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing R.G LANDES COMPANY Austin, Texas, U.S.A Copyright © 1998 R.G Landes Company All rights reserved No part of this book may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopy, recording, or any information storage and retrieval system, without permission in writing from the publisher Printed in the U.S.A Please address all inquiries to the Publishers: R.G Landes Company, 810 South Church Street, Georgetown, Texas, U.S.A 78626 Phone: 512/ 863 7762; FAX: 512/ 863 0081 ISBN: 1-57059-553-4 While the authors, editors and publisher believe that drug selection and dosage and the specifications and usage of equipment and devices, as set forth in this book, are in accord with current recommendations and practice at the time of publication, they make no warranty, expressed or implied, with respect to material described in this book In view of the ongoing research, equipment development, changes in governmental regulations and the rapid accumulation of information relating to the biomedical sciences, the reader is urged to carefully review and evaluate the information provided herein Library of Congress Cataloging-in-Publication Data Proteolytic and cellular mechanisms in prohormone processing / [edited by] Vivian Y.H Hook p cm (Molecular biology intelligence unit) ISBN 1-57059-553-4 (alk paper) Peptide hormones Metabolism Proteolytic enzymes Peptide hormones-Physiological transport Protein precursors Post-translational modifications I Hook, Vivian Yuan-Hen Ho, 1953- II Series QP572.P4P767 1998 572'.76 dc21 98-28730 CIP PUBLISHER’S NOTE Landes Bioscience produces books in six Intelligence Unit series: Medical, Molecular Biology, Neuroscience, Tissue Engineering, Biotechnology and Environmental The authors of our books are acknowledged leaders in their fields Topics are unique; almost without exception, no similar books exist on these topics Our goal is to publish books in important and rapidly changing areas of bioscience for sophisticated researchers and clinicians To achieve this goal, we have accelerated our publishing program to conform to the fast pace at which information grows in bioscience Most of our books are published within 90 to 120 days of receipt of the manuscript We would like to thank our readers for their continuing interest and welcome any comments or suggestions they may have for future books Judith Kemper Production Manager R.G Landes Company CONTENTS Targeting and Activation of Peptide Hormones in the Secretory Pathway Ken Teter and Hsiao-Ping H Moore Introduction Trafficking and Modification of Peptide Hormone Precursors Prohormone Sorting Mechanisms Site of Prohormone Sorting 10 Prohormone Activation 12 Summary and Future Perspectives 15 The Mechanism of Sorting Proopiomelanocortin to Secretory Granules and Its Processing by Aspartic and PC Enzymes 29 Niamh X Cawley, David R Cool, Emmanuel Normant, Fu-Sheng Shen, Vicki Olsen and Y Peng Loh General Introduction 29 Mechanism of Sorting POMC to the Regulated Secretory Pathway 31 Endoproteolytic Processing of Proopiomelanocortin 34 Future Directions 42 The Mammalian Precursor Convertases: Paralogs of the Subtilisin/ Kexin Family of Calcium-Dependent Serine Proteinases 49 Nabil G Seidah, Majambu Mbikay, Mieczyslaw Marcinkiewicz, Michel Chrétien Introduction 49 Subtilisin/Kexin-like Precursor Convertases (PCs): Structural and Cellular Considerations 51 Ontogeny, Tissue Expression and Subcellular Localization 59 Structure, Loci, and Evolution of PC Genes 62 Antisense Transgene Inhibition 63 Heritable Deficiency of PC in Human and Mouse 65 Inhibitors of PCs 66 Enzymatic Cascades: ADAM Family and PCs 67 Conclusions 68 The Neuroendocrine Prohormone Convertases PC1, PC2 and PC5 77 Margery C Beinfeld Introduction 77 The Discovery of the Subtilisin Family of Prohormone Convertases 77 Distribution of PC1, PC2 and PC5 79 Biosynthesis and Activation of PC1, PC2 and PC5 79 Regulation of PC Expression 80 Experimental Systems Used to Study Processing 80 Enzymatic Activity of PC1, PC2, and PC5 81 Antisense PC1 and PC2 Strategies to Study Proneuropeptide Processing Endoproteases in CCK Processing, a Case in Point Processing Enzyme Knockouts and Mutations Future Challenges 81 82 82 83 ‘Prohormone Thiol Protease’ (PTP), a Novel Cysteine Protease for Proenkephalin and Prohormone Processing 89 Vivian Y.H Hook, Yuan-Hsu Kang, Martin Schiller, Nikolaos Tezapsidis, Jane M Johnston and Ada Azaryan Introduction 89 The Novel ‘Prohormone Thiol Protease’ (PTP): A Major Proenkephalin Processing Enzyme in Chromaffin Granules 92 Participation of PC1/3 and PC2 Subtilisin-Like Proteases, and 70 kDa Aspartyl Protease (PCE) in Proenkephalin Processing in Chromaffin Granules 100 Conclusions 100 Regulation of Prohormone Conversion by Coordinated Control of Processing Endopeptidase Biosynthesis with That of the Prohormone Substrate 105 Terence P Herbert, Cristina Alarcon, Robert H Skelly, L Cornelius Bollheimer, George T Schuppin and Christopher J Rhodes Introduction 105 Coordinated Regulation of Prohormone and Processing Enzyme mRNA Levels 106 Coordinated Translational Regulation of Specific Prohormone and Processing Enzyme Biosynthesis 110 Carboxypeptidase and Aminopeptidase Proteases in Proneuropeptide Processing 121 Vivian Y.H Hook and Sukkid Yasothornsrikul Introduction 121 Neuroendocrine-specific Carboxypeptidase E/H 122 Molecular Genetic Analysis of Mutant Carboxypeptidase E/H in fat/fat Obese Mice: Effects of Inactive CPE/H on Prohormone Processing 129 Mutant CPE/H in fat/fat Mice Leads to Discovery of Novel Carboxypeptidase D and Carboxypeptidase Z 130 Evidence for CPE/H as a Sorting Receptor for the Intracellular Routing of POMC and Possibly Other Prohormones to the Secretory Vesicle 132 Aminopeptidase(s) for Prohormone Processing 133 Conclusions and Future Perspectives 134 The Neuroendocrine Polypeptide 7B2 as a Molecular Chaperone and Naturally Occurring Inhibitor of Prohormone Convertase PC2 141 A Martin Van Horssen and Gerard J.M Martens Introduction 141 History of 7B2 141 The 7B2 Gene and Its Regulation 142 Evolutionary Aspects 144 7B2 is a Neuroendocrine-Specific Polypeptide 144 Biochemical Characteristics of 7B2 145 Posttranslational Modifications of 7B2 145 Regulated Secretion of 7B2 146 The Quest for the Role of 7B2 146 Model of the Interaction Between 7B2 and PC2 149 Implications and Future Prospects 151 Neuroendocrine α1-Antichymotrypsin as a Possible Regulator of Prohormone and Neuropeptide Precursor Processing 159 Shin-Rong Hwang and Vivian Y.H Hook Introduction 159 Biochemical Evidence for α1-Antichymotrypsin (ACT) as an Endogenous Regulator of the ‘Prohormone Thiol Protease’ (PTP) and Other Prohormone Processing Proteases 160 Molecular Cloning Reveals Multiple Isoforms of Bovine ACT Expressed in Neuroendocrine Tissues 164 10 Proteolytic Inactivation of Secreted Neuropeptides 173 Eva Csuhai, Afshin Safavi, Michael W Thompson and Louis B Hersh Introduction 173 Neprilysin 174 Aminopeptidases 176 Angiotensin Converting Enzyme 178 Pyroglutamyl Peptidase II 178 Proline Specific Peptidases 179 Soluble Neuropeptidases 180 Endopeptidase 24.15 and Endopeptidase 24.16 181 Summary 182 11 Stimulation of Peptidergic Receptors by Peptide Hormones and Neurotransmitters: Studies of Opioid Receptors 191 George Bot, Allan D Blake and Terry Reisine Introduction 191 Opioid Receptor Types 191 Endogenous Opioids 192 Endogenous Peptide Receptor Selectivity 192 Opioid Ligands 194 Opioid Cellular Activity 195 Opioid Receptor Cloning 196 ORL1 and Nociceptin/Orphanin FQ 196 Structure-Function Analysis of Cloned Opioid Receptors 198 µ Receptor Knockout Mice Model 201 Agonist Regulation of Cloned Opioid Receptors 201 G Protein Role in Differential Agonist Activity 204 Conclusion 204 Index 213 EDITORS Vivian Y.H Hook Department of Medicine University of California, San Diego La Jolla, California, U.S.A Chapters 5, 7, CONTRIBUTORS Cristina Alarcon Gifford Laboratories for Diabetes Research Department of of Internal Medicine and Pharmacology University of Texas Southwestern Medical Center Dallas, Texas, U.S.A Chapter Cornelius Bollheimer Gifford Laboratories for Diabetes Research Department of of Internal Medicine and Pharmacology University of Texas Southwestern Medical Center Dallas, Texas, U.S.A Chapter Ada Azaryan Department of Pharmacology Uniformed Services University of the Health Sciences Bethesda, Maryland, U.S.A Chapter George Bot Department of Pharmacology University of Pennsylvania School of Medicine Philadelphia, Pennsylvania, U.S.A Chapter 11 Margery C Beinfeld Department of Pharmacology and Experimental Therapeutics Tufts University School of Medicine Boston, Massachusetts, U.S.A Chapter Niamh X Cawley Section on Cellular Neurobiology Laboratory of Developmental Neurobiology National Institutes of Child Health and Human Development National Institutes of Health Bethesda, Maryland, U.S.A Chapter Allan D Blake Department of Pharmacology University of Pennsylvania School of Medicine Philadelphia, Pennsylvania, U.S.A Chapter 11 Michel Chrétien J.A De Sève Laboratories of Molecular Neuroendocrinology Clinical Research Institute of Montreal Montreal, Quebec, Canada Chapter 202 Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing showed that the expressed µ receptor was downregulated by DAMGO treatment and that this downregulation event was correlated with phosphorylation of the receptor However, it is unclear from this study101 whether the downregulation and phosphorylation events that were observed in HEK 293 cells also correlated with functional desensitization of the µ receptor Zhang et al102 have reported the phosphorylation of the µ receptor in CHO cells, when the cells were challenged with either morphine or an activator of protein kinase C, although the impact of the phosphorylation of receptor function was unclear Chakrabarti et al103 observed a time and concentration-dependent effect of morphine and DAMGO on receptor downregulation and receptor desensitization In contrast, Blake et al104 found that morphine and DAMGO pretreatment sensitized their subsequent actions on µ receptor function in HEK 293 cells and that neither agonist caused receptor desensitization, with only DAMGO causing receptor downregulation However, methadone and buprenorphine, two opioids used clinically in treating opioid dependence, caused a pronounced receptor desensitization, suggesting that the therapeutic effect of these agents may be linked to the inhibition of receptor function.104 Studies have also shown that those compounds, fentanyl, sufentanil, lofentanil and nalbuphine, that desensitized the µ receptor, were not dependent on the Asp-114 residue for activation of the µ receptor to inhibit cAMP accumulation.70 The necessity of Asp-114 for morphine and levorphanol to stimulate the µ receptor, and the lack of its requirement for the fentanyl analogs and nalbuphine activation, indicate that these compounds have different determinants in the µ receptor for activation By interacting with the µ receptor differently, the fentanyl analogs and nalbuphine may activate adaptive cellular responses that result in µ receptor/adenylyl cyclase uncoupling In contrast, morphine may not stimulate these cellular pathways, even though fentanyl and morphine both bind to the same receptor and are equally effective in inhibiting adenylyl cyclase Studies have also shown that receptor desensitization occurred independently of receptor internalization70,104 and that the internalization induced did not correlate with the magnitude of µ receptor desensitization.70 Binding and immunofluorescent techniques demonstrated that morphine had little effect on receptor internalization, while etorphine induced a rapid receptor sequestration and desensitization.70,105 However, lofentanil, which abolished coupling of the µ receptor to adenylyl cyclase, caused no greater magnitude of internalization than fentanyl, which caused only a small reduction in maximal accumulation of cAMP.70 While differences in receptor regulation may be an inherent property of the surrogate cell lines used in the different studies, these results suggest that opioids differ markedly in their abilities to regulate the µ receptor at the cellular level A recent study has noted that the agonist regulation effects observed in µ receptor-transfected surrogate cell lines reflect biologically relevant actions that occur at the µ receptor in native tissue preparations.106 κ receptor Agonist regulation studies on the opioid κ receptor have yielded contrasting results Raynor et al107 were able to show a role of G protein-coupled protein kinases in the desensitization of the mouse κ receptor and the desensitization appeared to be independent of receptor downregulation Tallent et al108 observed that agonist pretreatment of the mouse κ receptor resulted in a pronounced homologous desensitization of adenylyl cyclase activity, in addition to desensitizing a receptor-coupled K+ current On the other hand, Blake et al109 found that agonist regulation of the human κ receptor was dependent on the agonist used, with κ-selective agonists desensitizing and downregulating the receptor, while nonselective agonists were without effect In contrast, Avidor Reiss et al110 observed no agonist-mediated desensitization or downregulation with the rat κ receptor At present, the cellular basis for the differences observed in the agonist regulation of the opioid v receptor remains unre- Stimulation of Peptidergic Receptors by Peptide Hormones and Neurotransmitters 203 solved, although the differences may reflect properties unique to the surrogate cell lines used in the studies, or the species isoforms of the receptors studied δ receptor The opioid δ receptor appears to undergo desensitization and downregulation in response to δ-selective peptide and nonpeptide agonists more readily than either the µ or κ receptors.95,111-115 But like the µ and κ receptors, the δ receptor may also be capable of undergoing differential agonist regulation, since pretreatment with δ-selective peptides desensitized the receptor, whereas pretreatment with the δ-selective nonpeptide SIOM did not.95 Agonist-induced desensitization of the δ receptor appears to fit the general paradigm that has been described for G protein-coupled receptors,116 with agonist treatment uncoupling the receptor from G proteins111 and G protein-coupled protein kinases playing a role in attenuating the functional response.112 Agonist mediated sequestration and downregulation of the δ receptor appear to involve the receptor carboxyl terminal domain, with differences in the amino acids required for short-term receptor sequestration and downregulation.113,116 In this regard, agonist regulation of the opioid δ receptor appears to differ from that of the µ receptor, since studies indicate that the carboxyl terminal domain of the µ receptor is not necessary for receptor internalization89 but may be still necessary for desensitization.117 Electrophysiological Studies In Xenopus laevis oocytes expressing the cloned µ, δ or κ receptor and the cloned inward rectifier GIRK1, selective µ, δ and κ agonists increased K+ conductance.118-122 Similar results have been observed in the AtT20 cell line expressing the cloned µ receptor.108 In either systems expressing the µ receptor, the continuous presence of opiates uncoupled the µ receptor from the K+ channel and abolished the subsequent opiate potentiation of K+ conductance in a heterologous manner In the cell line AtT20 expressing the κ receptor, pretreatment with agonist U50,488 resulted in desensitization which was homologous since AtT20 cells treated with U50,488 still responded to somatostatin agonists with an increase in K+ currents.108 Chen and Yu119 observed a differential regulation, by intracellular protein kinases, of the human µ receptor activation of an inwardly rectifying K+ current, as protein kinase C activation potentiated DAMGO-mediated desensitization of the response, whilst protein kinase A activation abolished the current desensitization Mestek et al120 were able to demonstrate that intracellular protein kinases known to be dependent on phospholipase C activation were involved in potentiating the K+ current desensitization, as both protein kinase C and calcium\calmodulin-dependent protein kinase accentuated the desensitization elicited by DAMGO Zhang et al123 confirmed the earlier observations on protein kinase C effects with the µ receptor and found that these effects also occurred with the κ receptor The latter group proposed that two distinct pathways are involved in K+ current desensitization, based on observed differences in the time course of the agonist-mediated effect and the effects of the protein kinase C inhibitor staurosporine.123 In contrast, Kovoor et al118 suggested a post receptor mechanism for the agonist regulation of the K+ current in oocytes, with the desensitization occurring independently of protein kinase C activation Hence opiates appear to have a wide and varied effect upon intracellular effector systems However, it should be noted that adaptive responses occurring during administration of a particular opiate may not be identical in all opiate-sensitive neuronal populations and that a particular opiate may selectively desensitize some, but not all, intracellular functions 204 Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing G Protein Role in Differential Agonist Activity The receptor binding differences exhibited by opioids suggests that different opioids may interact differently with the receptor and induce a myriad of intracellular effector systems In support of this, there is also reasonable evidence to suggest that various opioid analgesics have different intrinsic efficacies and that these differences are likely due to the different abilities of these agonists to interact with the receptors and differentially activate G proteins.124,125 Hence opioids exhibit different efficacy and/or potency in the activation of different classes of G proteins.43,126 For example, antisense oligodeoxyribonucleotide against the subtype Gi2α protein antagonized morphine but not sufentanil-induced antinociception.124 Evidence also indicates that a single opioid receptor type can interact with several G proteins127 which, in turn, can couple to more than one effector128 and these may integrate coincident signals from different G protein subtypes.129 This coupling may differ in cell lines and neuronal membranes For example, in neuroblastoma glioma hybrid NG108-15 cells, δ receptors are coupled to inhibition of adenylyl cyclase via Gi2,42,130 whereas in human neuroblastoma SH-SY5Y cells they are coupled mainly via Gi1 and Go.131 The coupling efficacy to G proteins may also differ, as it has been reported that the δ receptor is more efficiently coupled to Gi2 protein than the µ receptor.130,132 Hence multiple G protein subunits are able to influence the actions of a single opioid agonist This suggests that although these compounds may be interacting with the same opioid receptor type, different intracellular effector mechanisms may be induced by them in producing their effects Hence the molecular determinants of receptor recognition may be different than for cellular activation The degree and efficiency of coupling to different cellular effectors in different systems may explain why agonist pretreatment has been shown to desensitize the coupling of the cloned µ, δ and κ receptors to K+ channels,109,133 and the δ and κ receptors from inhibition of adenylyl cyclase activity,95,107,112 whereas morphine has been reported not to uncouple the cloned µ receptor from adenylyl cyclase even though it effectively inhibited the activity of this enzyme.104 It may also explain the reported heterologous/homologous desensitization demonstrated for the opioid receptors The type of desensitization reported, i.e., homologous or heterologous, seems to be dependent on the opioid receptor type and the intracellular effector under investigation Interaction of the opioid receptor with the K+ channel may be of different magnitude and involve different G proteins than interaction with adenylyl cyclase Hence, adaptive responses occurring during opiate administration may not be identical in all opiate-sensitive neuronal populations and opiates may selectively desensitize some, but not all, intracellular functions of the opioid receptor Conclusion The use of heterologous expression studies in examining opioid receptor structure and function, in combination with genetic manipulations on the cloned opioid receptors, has revealed receptor domains involved in opioid ligand recognition and in the cellular regulation of receptor function However, binding of opiates and subsequent activation of intracellular function is agonist dependent Agonists and antagonists appear to have different determinants for binding to and activating the opioid receptors Furthermore, opioid compounds which produce tolerance readily, such as morphine, bind and activate the opioid receptor differently than opioids such as methadone and buprenorphine These differences may be linked to long-term functional consequences associated with their use Since the effectiveness of opioids in controlling chronic pain is limited by the undesirable side effects associated with long-term treatment, a better understanding of the molecular pharmacology of opioids is necessary Modeling of the receptor-binding sites, together with the known structure of the synthetic opiates, should facilitate the rational development of new compounds with improved therapeutic profile and limited side effects Stimulation of Peptidergic Receptors by Peptide Hormones and Neurotransmitters 205 Abbreviations TIPP[ψ] DPDPE DADLE DSLET EKC SIOM BW373U86 NTB BNTX TAN-67 U50,488 nor-BNI U69,593 NMDA DAMGO CTOP CTAP Deltorphin II H-Tyr-Ticψ(CH2-NH]-Phe-Phe-OH cyclic [D-Pen2, D-Pen5]enkephalin [D-Ala2, D-Leu5]enkephalin [D-Ser2, D-Leu5]enkephalin-Thr6 ethylketocyclazocine 7-spiroindanyloxymorphone (±)-4-{(α-R*)-α-[(2S*,5R*)-4-allyl-2,5-dimethyl-1-piperazinyl]-3hydroxybenzyl}-N,N-diethylbenzamide naltriben methanesulfonate 7-benylidenenaltrexone 2-methyl-4aα-(3-hydroxyphenyl)-1,2,3,4,4a,5,12,12aαoctahydroquinolino [2,3,3,-g]isoquinoline 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Interaction of δ-opioid receptors with multiple G proteins: A non-relationship between agonist potency to inhibit adenylyl cyclase and to activate G proteins Mol Pharmacol 1994; 45:997-1003 128 Gintzler AR, Xu H Different G proteins mediate the opioid inhibition or enhancement of evoked [5-methionine]enkephalin release Proc Natl Acad Sci (USA) 1991; 88:4741-4745 129 Lustig KD, Conklin BR, Herzmark P et al Type II adenylyl cyclase integrates coincident signals from GS, Gi and Gq J Biol Chem 1993; 268:13900-13905 130 McKenzie FR, Milligan G delta-opioid-receptor-mediated inhibition of adenylyl cyclase is transduced specifically by the guanine-nucleotide-binding protein Gi2 Biochem J 1990; 267:391-398 131 Laugwitz K-L, Offermanns S, Spicher K et al µ and δ opioid receptors differentially couple to G protein subtypes in membranes of human neuroblastoma SH-SY5Y cells Neuron 1993; 10:233-242 132 Offermanns S, Schultz G, Rosenthal W Evidence for opioid receptor-mediated activation of the G proteins, Go and Gi2, in membranes of neuroblastoma x glioma (NG108-15) hybrid cells J Biol Chem 1991; 266:3365-3368 133 Ma GH, Miller R, Kuznestov A et al 1995 Kappa-opioid receptor activates an inwardly rectifying K+ channel by a G protein-linked mechanism: Coexpression in Xenopus oocytes Mol Pharmacol 1995; 47:1035-1040 Index A α-melanocyte stimulating hormone (α-MSH) 29, 30, 42, 45, 55, 60, 68, 73, 87 α1-antichymotrypsin (ACT) 160-168 α1-antitrypsin portland (a1-PDX) 66 ACTH (adrenocorticotropin hormone) 10, 29, 30, 32, 34, 36-42, 61, 63, 80, 97, 107, 124, 127, 128, 130, 133, 135, 142 ADAM family of metalloproteases 60, 67, 68 Adrenocorticotropin 29, 30, 107, 124 Alzheimer’s disease 51, 161 Amastatin 133, 176, 177 Aminopeptidase 36, 90, 95, 96, 121, 123, 133-135, 174-178, 181 Aminopeptidase N 175, 176, 177 Angiotensin converting enzyme 175, 178, 181 Antisense 34, 41, 61, 63, 65, 66, 81, 82, 132, 160, 163, 204 Arterial restenosis 65 Aspartyl protease 67, 94, 97, 98, 100, 101, 121, 135, 159, 161 Atherosclerosis 65 Carboxypeptidase Z (CPZ) 130-132 Catalytic domain 51, 53, 55, 65, 77, 78, 148, 149, 176 Chaperone 3, 13, 51, 79, 80, 109, 144, 148, 149, 151, 152 Cholecystokinin 38, 63 Cholesterol metabolism 51 Chromaffin granule 36, 79, 92, 93, 95, 97, 99-101, 123, 127, 128, 133, 134, 161, 162 Chromogranin 9, 16, 32, 52, 81, 141, 159 Constitutive secretory pathway 1, 31, 49, 59, 66, 77 Cysteine protease 92, 93, 99-101, 121, 159, 161, 162, 166 D DAMGO 194, 195, 198-200, 202, 203, 205 Dipeptidyl peptidase IV (DPP IV) 179 Diprotin A and B 179 DPDPE 194, 195, 199, 200, 205 Dynorphin 81, 82, 130, 176, 178-180, 192, 194, 197, 200 B E β-endorphin (β-END) 29, 30, 36, 38-42, 50, Ectoenzyme 173, 177-181 Ectopeptidase 173 Endo-oligopeptidase A 181 Endomorphin 193, 194 Endoprotease 13, 29, 34, 41, 82, 89, 105-107, 110, 112, 134, 146, 147, 149, 174, 179, 180-182 Endoproteolytic processing 34, 89, 90, 121, 123, 133-135 Endothelin converting enzyme 175 Enkephalin 81, 82, 92-94, 97, 99, 100, 121-123, 126-128, 130, 133, 161-163, 174-178, 192-195, 198, 205 Enkephalinase 173-177 Exocytosis 1, 4, 8, 12, 31, 112 Exopeptidase 34, 106, 112, 121, 124, 133, 134, 176 97, 107, 180, 192-194 β-lipotropic hormone (β-LPH) 29, 30, 36, 38-42, 55, 107 Bestatin 133, 176, 177 Biosynthesis 29, 79, 99, 105, 106, 110-116, 127, 129, 142, 147, 152, 179, 198 Buprenorphine 195, 198, 202, 204 C CALLA 174 Captopril 178 Carboxypeptidase D (CPD) 131, 132 Carboxypeptidase E (CPE) 10, 11, 13-15, 32-35, 42, 132 Carboxypeptidase E (CpE) 5, 32, 35, 82, 121, 123 Carboxypeptidase E/H (CPE/H) 29, 121, 123-132, 134, 135 Carboxypeptidase H (CPH) 32, 80, 106, 109, 115, 121, 123 F Furin 5-7, 12, 13, 39, 49, 51, 53, 55, 59-64, 66-68, 77-79, 81, 82, 146-149, 151 214 Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing G O γ-endorphin generating enzyme (γ-EGE) 180 Gene expression 29, 105-109, 127 Glucose 2, 79, 83, 105-107, 110-116, 127, 148 Gluzincin 174, 176, 177 GroEL 144 Growth factor 59, 63, 65, 66, 142, 147 Guanine nucleotide binding protein 108, 195-197, 199, 201-204 Guanine nucleotide binding protein (G protein) Obesity 64, 65, 83, 121, 129, 130 Opiate 29, 191, 194-201, 203, 204 Opioid receptor 191-199, 201, 204 Orphan receptor 176 I Immature secretory granule (ISG) 3-5, 8-16, 29, 31, 35, 49, 51, 67, 146-150 Inactivation 64-66, 175 Insulin degrading enzyme (IDE) 180, 181 Integrin 50, 55, 67, 68, 79 Inverzincin 174, 180 K KELL blood group protein 175 Kexin 49, 51, 53-55, 59, 68 M Metalloprotease 67, 124, 131, 133, 134, 180 Methadone 191, 194, 195, 198, 202, 204 Morphine 191, 192, 194, 195, 198-202, 204 MRNA 38-41, 59-63, 65, 68, 79, 80, 82, 89, 105-113, 115, 116, 126, 127, 131, 142, 144, 166, 175, 178 Mutagenesis 32, 148, 174, 198, 199 N Neprilysin 174-176, 180 Neuroendocrine 9, 10, 34, 39, 77, 89, 90, 101, 105, 121, 122, 124-128, 130, 134, 141, 142, 144-149, 151, 152, 159-161, 163-166, 168 Neurolysin 181 Neuropeptide 29, 38, 89, 90, 92, 94, 101, 121-123, 125-130, 133-135, 141, 147, 149, 152, 159-161, 168, 173, 174, 179-182 Nociceptin 176, 193, 196-198 P Peptide hormone 1, 3, 5, 12, 14, 30, 31, 34, 38, 89, 101, 105, 106, 110, 121-123, 126, 127, 130, 135, 141, 142, 145, 147, 149, 150, 152, 159, 168 Peptide neurotransmitter 1, 159 Pituitary 8, 29, 30, 32-34, 36-42, 59-61, 79, 80, 90, 91, 94, 100, 101, 107-109, 122, 124-134, 141, 142, 144-149, 159, 161, 162, 164-166, 168 POMC sorting signal 32, 33 Posttranslational modification 34, 110, 129, 145 Precursor convertase 49, 51, 68 Precursor convertase (PC) 49-51, 53-55, 57, 59, 60-68, 77-83, 94, 95, 97, 98, 100, 101, 106-112, 114-116, 121, 130, 131, 135, 142, 146-152, 159, 161, 164 Proenkephalin 32, 34, 37, 79, 81, 92-94, 97-101, 121, 122, 142, 159, 161-163, 192 Prohormone 1, 3-5, 7-16, 29, 31, 32, 34, 36-39, 42, 65, 77-83, 89-95, 97, 98, 100, 101, 105-110, 112, 121-125, 129, 130, 132-135, 141, 142, 146-152, 159-161, 163, 164, 166, 168 Prohormone convertase 1, 5, 12, 16, 29, 34, 37-39, 77, 78, 83, 94, 101, 105, 121, 130, 135, 142, 146, 147, 152, 159, 160, 164 Prohormone processing 4, 10, 14, 15, 36, 38, 42, 77, 78, 81, 82, 89-92, 98, 100, 101, 105, 107, 110, 121-124, 129, 130, 133, 134, 135, 146, 149, 159-161, 163, 166, 168 Prohormone thiol protease (PTP) 37, 92-101, 121, 134, 135, 159-164, 166, 168 Proinsulin 3, 5, 7, 9, 10, 32, 34, 36, 41, 42, 50, 65, 83, 90, 91, 97, 101, 106-116, 121, 122, 130, 134, 142, 147 Promoter 63, 80, 82, 92, 107-109, 142, 175 Proneuropeptide 31, 32, 37, 39, 63, 65, 77, 81, 92, 97, 159, 160 Proopiomelanocortin (POMC) 9, 10, 13, 29-42, 50, 55, 63, 65, 80, 81, 94, 97, 98, 100, 101, 106-110, 121, 122, 124, 130, 132-135, 142, 146, 147, 159, 161, 192 Index ProPC2 maturation 13, 147, 149 Proprotein convertase 67, 105, 110, 143, 144, 146, 151 Protease inhibitor 36, 99, 100, 124, 131, 134, 159-163, 166, 168 Proteolysis 12, 13, 49, 65, 89, 90, 105, 129, 160 R Reactive site loop (RSL) 160, 161, 164-166, 168 Receptor desensitization 202, 203 Receptor internalization 173, 202, 203 Receptor mutagenesis 198, 199 Receptor subtype 194-196, 198, 199, 201 Regulated secretory pathway 3, 8, 10, 11, 31, 32, 34, 39, 42, 49, 59, 77, 78, 89, 107, 129, 132, 141, 145, 146, 148, 159, 160 Regulation 1, 4, 12, 34, 39, 40, 42, 51, 59, 60, 68, 80, 83, 99, 105-108, 110-115, 127, 128, 135, 142, 152, 164, 176, 178, 179, 181, 201-204 Renin 50, 52, 67, 81 215 Secretogranin 81, 141 Secretory granule 1, 3, 8, 11, 16, 31-35, 41, 42, 49, 51, 55, 59, 66, 67, 77, 79, 81, 110, 129, 141, 142, 144, 146-150, 173 Secretory vesicle 1, 15, 36, 42, 77, 81, 89-93, 97, 99-101, 122-125, 128, 129, 131-134, 160-163 Serine proteinase 49, 59, 68 Serpin 66, 160-162, 164-167 Seven-transmembrane protein 196, 197 Sorting receptor 3, 5, 10, 32-34, 42, 132 Subtilisin 38, 49, 51, 54, 55, 59, 68, 77, 78, 83, 94, 100, 101, 121, 135, 144, 149, 159, 160, 161, 164, 174, 179 T Trans-Golgi (TGN) 1, 3-16, 29, 31, 32, 34, 35, 49, 51, 55, 59, 66, 67, 145, 147-150, 160 Transcription factor 107-109, 142 Translational regulation 105, 110, 112, 113 V Viral surface glycoprotein 66 S Secretase 52 Secretion 1-5, 11, 12, 16, 32, 34, 36, 55, 59, 78, 80, 89, 105-107, 112, 128, 132-134, 141, 142, 145-147, 149 Z Zymogen 51, 55, 59, 67, 97, 149, 168 [...]... regarding the site of prohormone sorting and activation Location in the Presence of BfA Definition Table 1.4 Definitions of the TGN and responses to Brefeldin A Targeting and Activation of Peptide Hormones in the Secretory Pathway 7 8 Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing Prohormone cleavage and activation continues in the ISG, the vesicular intermediate linking... protein to the AtT20 SG initially demonstrated that regulated proteins contain positive sorting information,122 and subsequent work has found that the propeptides of several hormone precursors contain SG targeting signals.123-128 However, deletional analysis of proopiomelanocortin (POMC), protrypsinogen 10 Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing and proinsulin has indicated... understanding of current knowledge concerning proteases involved in prohormone processing, and cellular aspects that must be considered for proper processing, storage, and secretion of bioactive peptides It is well known that prohormone processing occurs in well-defined subcellular compartments of the regulated secretory pathway Knowledge of the cell biology of prohormone processing is required in the... animal cells 2 Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing Targeting and Activation of Peptide Hormones in the Secretory Pathway 3 prohormones and prohormone converting enzymes along the biosynthetic regulated secretory pathway Trafficking and Modification of Peptide Hormone Precursors Proteins destined for either regulated or constitutive release are transported and modified... allows inhibitor association and hence suppression of PC activity until the acidic lumen of the target compartment inactivates the inhibitor and allows full expression of PC activity 14 Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing Compartmental pH and Ca2+ levels thus play a pivotal role in trafficking within the regulated pathway, influencing: 1 formation of macromolecular... pro-region.73,161-164 Synthesis and cleavage of the propeptide appear to be essential for proper folding, trafficking and activation of furin since unprocessed pro-furin is retained in the ER and has no substrate processing activity.73,163,164 The cleaved propeptide remains bound to the mature protein and serves as a trans-acting inhibitor.165 The enzyme therefore remains inactive in the proximal part of the... destined for secretion appear to be excluded from these retrograde transport carriers, as recent studies indicate that proinsulin and VSV G protein are segregated into distinct vesicles from those containing the ER retrieval KDEL receptor.46 Interestingly, the proinsulin-containing vesicles are also COPI-coated The possibility that COPI functions in both anterograde and retrograde transport remains... carriers, the ISG and constitutive vesicle The relative partitioning of cargo into each vesicle type appears to vary significantly for different proteins and cell 12 Proteolytic and Cellular Mechanisms in Prohormone and Proprotein Processing types.118,145,146,148 “Sorting for entry” enriches the ISG with regulated secretory proteins, but does not efficiently exclude constitutive secretory proteins or lysosomal... P Calnexin and BiP act as sequential molecular chaperones during thyroglobulin folding in the endoplasmic reticulum J Cell Biol 1995; 128(1-2):29-38 31 Schmitz A, Maintz M, Kehle T et al In vivo iodination of a misfolded proinsulin reveals co-localized signals for Bip binding and for degradation in the ER EMBO J 1995; 14(6):1091-8 32 Huang XF, Arvan P Intracellular transport of proinsulin in pancreatic... Cellular Mechanisms in Prohormone and Proprotein Processing Targeting and Activation of Peptide Hormones in the Secretory Pathway 5 identified, trafficking of these proteins may also involve ER export signals and cargo sorting receptors By contrast, bulk flow may apply to intra-Golgi transport since further concentration of migrant proteins does not occur as trafficking continues across the Golgi stacks.38,39,46