請用此 Handle URI 來引用此文件:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/67447
完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 湯志永 | |
dc.contributor.author | Yi-Ching Lee | en |
dc.contributor.author | 李宜靜 | zh_TW |
dc.date.accessioned | 2021-06-17T01:32:36Z | - |
dc.date.available | 2020-09-08 | |
dc.date.copyright | 2017-09-08 | |
dc.date.issued | 2017 | |
dc.date.submitted | 2017-08-03 | |
dc.identifier.citation | 參考資料
Accardi A, Pusch M (2000) Fast and slow gating relaxations in the muscle chloride channel CLC-1. The Journal of general physiology 116:433-444. Apaja PM, Foo B, Okiyoneda T, Valinsky WC, Barriere H, Atanasiu R, Ficker E, Lukacs GL, Shrier A (2013) Ubiquitination-dependent quality control of hERG K+ channel with acquired and inherited conformational defect at the plasma membrane. Molecular biology of the cell 24:3787-3804. Araki T, Milbrandt J (2003) ZNRF Proteins Constitute a Family of Presynaptic E3 Ubiquitin Ligases. The Journal of Neuroscience 23:9385-9394. Araki T, Nagarajan R, Milbrandt J (2001) Identification of genes induced in peripheral nerve after injury. Expression profiling and novel gene discovery. The Journal of biological chemistry 276:34131-34141. Aromataris EC, Rychkov GY (2006) ClC-1 chloride channel: Matching its properties to a role in skeletal muscle. Clinical and experimental pharmacology physiology 33:1118-1123. Arvan P, Zhao X, Ramos-Castaneda J, Chang A (2002) Secretory pathway quality control operating in Golgi, plasmalemmal, and endosomal systems. Traffic (Copenhagen, Denmark) 3:771-780. Asada S, Ikeda A, Nagao R, Hama H, Sudo T, Fukamizu A, Kasuya Y, Kishi T (2008) Oxidative stress-induced ubiquitination of RCAN1 mediated by SCFbeta-TrCP ubiquitin ligase. International journal of molecular medicine 22:95-104. Bagola K, Mehnert M, Jarosch E, Sommer T (2011) Protein dislocation from the ER. Biochimica et biophysica acta 1808:925-936. Behrends C, Harper JW (2011) Constructing and decoding unconventional ubiquitin chains. Nature structural molecular biology 18:520-528. Bernier V, Lagace M, Bichet DG, Bouvier M (2004) Pharmacological chaperones: potential treatment for conformational diseases. Trends in endocrinology and metabolism: TEM 15:222-228. Bienert GP, Schjoerring JK, Jahn TP (2006) Membrane transport of hydrogen peroxide. Biochimica et biophysica acta 1758:994-1003. Bienert GP, Moller AL, Kristiansen KA, Schulz A, Moller IM, Schjoerring JK, Jahn TP (2007) Specific aquaporins facilitate the diffusion of hydrogen peroxide across membranes. The Journal of biological chemistry 282:1183-1192. Blundell KL, Pal M, Roe SM, Pearl LH, Prodromou C (2017) The structure of FKBP38 in complex with the MEEVD tetratricopeptide binding-motif of Hsp90. PloS one 12:e0173543. Bonifacino JS, Glick BS (2004) The mechanisms of vesicle budding and fusion. Cell 116:153-166. Brandizzi F, Barlowe C (2013) Organization of the ER-Golgi interface for membrane traffic control. Nature reviews Molecular cell biology 14:382-392. Brandvold KR, Morimoto RI (2015) The Chemical Biology of Molecular Chaperones--Implications for Modulation of Proteostasis. Journal of molecular biology 427:2931-2947. Bretag AH (1987) Muscle chloride channels. Physiological reviews 67:618-724. Cao X, Fang Y (2015) Transducing oxidative stress to death signals in neurons. The Journal of cell biology 211:741-743. Cha-Molstad H et al. (2015) Amino-terminal arginylation targets endoplasmic reticulum chaperone BiP for autophagy through p62 binding. Nature cell biology 17:917-929. Chaudhuri TK, Paul S (2006) Protein-misfolding diseases and chaperone-based therapeutic approaches. The FEBS journal 273:1331-1349. Chen YA, Peng YJ, Hu MC, Huang JJ, Chien YC, Wu JT, Chen TY, Tang CY (2015) The Cullin 4A/B-DDB1-Cereblon E3 Ubiquitin Ligase Complex Mediates the Degradation of CLC-1 Chloride Channels. Scientific reports 5:10667. Chiang HL, Terlecky SR, Plant CP, Dice JF (1989) A role for a 70-kilodalton heat shock protein in lysosomal degradation of intracellular proteins. Science (New York, NY) 246:382-385. Chien YC and Tang CY (2015) Membrane distribution of molecular chaperones and ubiquitin ligases with human CLC-1 channels. 台灣大學碩士論文 Choi BH, Feng L, Yoon HS (2010) FKBP38 protects Bcl-2 from caspase-dependent degradation. The Journal of biological chemistry 285:9770-9779. Colding-Jorgensen E (2005) Phenotypic variability in myotonia congenita. Muscle nerve 32:19-34. Cole NB, Ellenberg J, Song J, DiEuliis D, Lippincott-Schwartz J (1998) Retrograde transport of Golgi-localized proteins to the ER. The Journal of cell biology 140:1-15. Cross DA, Alessi DR, Cohen P, Andjelkovich M, Hemmings BA (1995) Inhibition of glycogen synthase kinase-3 by insulin mediated by protein kinase B. Nature 378:785-789. Desaphy JF, Gramegna G, Altamura C, Dinardo MM, Imbrici P, George AL, Jr., Modoni A, Lomonaco M, Conte Camerino D (2013) Functional characterization of ClC-1 mutations from patients affected by recessive myotonia congenita presenting with different clinical phenotypes. Experimental neurology 248:530-540. Deshaies RJ, Joazeiro CA (2009) RING domain E3 ubiquitin ligases. Annual review of biochemistry 78:399-434. Dobson CM (2003) Protein folding and misfolding. Nature 426:884-890. DuBridge RB, Tang P, Hsia HC, Leong PM, Miller JH, Calos MP (1987) Analysis of mutation in human cells by using an Epstein-Barr virus shuttle system. Molecular and cellular biology 7:379-387. Edlich F, Fischer G (2006) Pharmacological targeting of catalyzed protein folding: the example of peptide bond cis/trans isomerases. Handbook of experimental pharmacology:359-404. Edlich F, Erdmann F, Jarczowski F, Moutty MC, Weiwad M, Fischer G (2007) The Bcl-2 regulator FKBP38-calmodulin-Ca2+ is inhibited by Hsp90. The Journal of biological chemistry 282:15341-15348. Edlich F, Weiwad M, Erdmann F, Fanghanel J, Jarczowski F, Rahfeld JU, Fischer G (2005) Bcl-2 regulator FKBP38 is activated by Ca2+/calmodulin. The EMBO journal 24:2688-2699. Ellgaard L, Helenius A (2003) Quality control in the endoplasmic reticulum. Nature reviews Molecular cell biology 4:181-191. Farhan H, Rabouille C (2011) Signalling to and from the secretory pathway. Journal of cell science 124:171-180. Flynn GC, Pohl J, Flocco MT, Rothman JE (1991) Peptide-binding specificity of the molecular chaperone BiP. Nature 353:726-730. Frykman S, Hur JY, Franberg J, Aoki M, Winblad B, Nahalkova J, Behbahani H, Tjernberg LO (2010) Synaptic and endosomal localization of active gamma-secretase in rat brain. PloS one 5:e8948. Gallagher E, Gao M, Liu YC, Karin M (2006) Activation of the E3 ubiquitin ligase Itch through a phosphorylation-induced conformational change. Proceedings of the National Academy of Sciences of the United States of America 103:1717-1722. Ge L, Melville D, Zhang M, Schekman R (2013) The ER-Golgi intermediate compartment is a key membrane source for the LC3 lipidation step of autophagosome biogenesis. eLife 2:e00947. Graham JM (2001) Isolation of Golgi membranes from tissues and cells by differential and density gradient centrifugation. Current protocols in cell biology Chapter 3:Unit 3.9. Harrar Y, Bellini C, Faure JD (2001) FKBPs: at the crossroads of folding and transduction. Trends in plant science 6:426-431. Hartl FU (1996) Molecular chaperones in cellular protein folding. Nature 381:571-579. Hartl FU, Hayer-Hartl M (2002) Molecular chaperones in the cytosol: from nascent chain to folded protein. Science (New York, NY) 295:1852-1858. Hartl FU, Hayer-Hartl M (2009) Converging concepts of protein folding in vitro and in vivo. Nature structural molecular biology 16:574-581. Hartl FU, Bracher A, Hayer-Hartl M (2011) Molecular chaperones in protein folding and proteostasis. Nature 475:324-332. Hoeller D, Dikic I (2010) Regulation of ubiquitin receptors by coupled monoubiquitination. Sub-cellular biochemistry 54:31-40. Hoxhaj G, Najafov A, Toth R, Campbell DG, Prescott AR, MacKintosh C (2012) ZNRF2 is released from membranes by growth factors and, together with ZNRF1, regulates the Na+/K+ATPase. Journal of cell science 125:4662-4675. Hutt DM, Roth DM, Chalfant MA, Youker RT, Matteson J, Brodsky JL, Balch WE (2012) FK506 binding protein 8 peptidylprolyl isomerase activity manages a late stage of cystic fibrosis transmembrane conductance regulator (CFTR) folding and stability. The Journal of biological chemistry 287:21914-21925. Jentsch TJ, Stein V, Weinreich F, Zdebik AA (2002) Molecular structure and physiological function of chloride channels. Physiological reviews 82:503-568. Kampinga HH, Craig EA (2010) The HSP70 chaperone machinery: J proteins as drivers of functional specificity. Nature reviews Molecular cell biology 11:579-592. Kee Y, Huibregtse JM (2007) Regulation of catalytic activities of HECT ubiquitin ligases. Biochemical and biophysical research communications 354:329-333. Kim YE, Hipp MS, Bracher A, Hayer-Hartl M, Hartl FU (2013) Molecular chaperone functions in protein folding and proteostasis. Annual review of biochemistry 82:323-355. Klausner RD, Donaldson JG, Lippincott-Schwartz J (1992) Brefeldin A: insights into the control of membrane traffic and organelle structure. The Journal of cell biology 116:1071-1080. Kloetzel PM, Ossendorp F (2004) Proteasome and peptidase function in MHC-class-I-mediated antigen presentation. Current opinion in immunology 16:76-81. Koch MC, Steinmeyer K, Lorenz C, Ricker K, Wolf F, Otto M, Zoll B, Lehmann-Horn F, Grzeschik KH, Jentsch TJ (1992) The skeletal muscle chloride channel in dominant and recessive human myotonia. Science (New York, NY) 257:797-800. Komander D (2009) The emerging complexity of protein ubiquitination. Biochemical Society transactions 37:937-953. Kullmann DM (2010) Neurological channelopathies. Annual review of neuroscience 33:151-172. Lam E, Martin M, Wiederrecht G (1995) Isolation of a cDNA encoding a novel human FK506-binding protein homolog containing leucine zipper and tetratricopeptide repeat motifs. Gene 160:297-302. Lamark T, Kirkin V, Dikic I, Johansen T (2009) NBR1 and p62 as cargo receptors for selective autophagy of ubiquitinated targets. Cell cycle (Georgetown, Tex) 8:1986-1990. Lecker SH, Goldberg AL, Mitch WE (2006) Protein degradation by the ubiquitin-proteasome pathway in normal and disease states. Journal of the American Society of Nephrology : JASN 17:1807-1819. Lee C, Goldberg J (2010) Structure of coatomer cage proteins and the relationship among COPI, COPII, and clathrin vesicle coats. Cell 142:123-132. Lee CY, Lai TY, Tsai MK, Chang YC, Ho YH, Yu IS, Yeh TW, Chou CC, Lin YS, Lawrence T, Hsu LC (2017) The ubiquitin ligase ZNRF1 promotes caveolin-1 ubiquitination and degradation to modulate inflammation. Nature communications 8:15502. Lee TT, Zhang XD, Chuang CC, Chen JJ, Chen YA, Chen SC, Chen TY, Tang CY (2013) Myotonia congenita mutation enhances the degradation of human CLC-1 chloride channels. PloS one 8:e55930. Li L, Ren CH, Tahir SA, Ren C, Thompson TC (2003) Caveolin-1 maintains activated Akt in prostate cancer cells through scaffolding domain binding site interactions with and inhibition of serine/threonine protein phosphatases PP1 and PP2A. Molecular and cellular biology 23:9389-9404. Li WW, Li J, Bao JK (2012) Microautophagy: lesser-known self-eating. Cellular and molecular life sciences : CMLS 69:1125-1136. Lossin C, George AL, Jr. (2008) Myotonia congenita. Advances in genetics 63:25-55. Loureiro CA, Matos AM, Dias-Alves A, Pereira JF, Uliyakina I, Barros P, Amaral MD, Matos P (2015) A molecular switch in the scaffold NHERF1 enables misfolded CFTR to evade the peripheral quality control checkpoint. Science signaling 8:ra48. MacGurn JA, Hsu PC, Emr SD (2012) Ubiquitin and membrane protein turnover: from cradle to grave. Annual review of biochemistry 81:231-259. Macias MJ, Teijido O, Zifarelli G, Martin P, Ramirez-Espain X, Zorzano A, Palacin M, Pusch M, Estevez R (2007) Myotonia-related mutations in the distal C-terminus of ClC-1 and ClC-0 chloride channels affect the structure of a poly-proline helix. The Biochemical journal 403:79-87. McMahon HT, Mills IG (2004) COP and clathrin-coated vesicle budding: different pathways, common approaches. Current opinion in cell biology 16:379-391. Mellman I, Warren G (2000) The road taken: past and future foundations of membrane traffic. Cell 100:99-112. Meyer S, Dutzler R (2006) Crystal structure of the cytoplasmic domain of the chloride channel ClC-0. Structure (London, England : 1993) 14:299-307. Nilius B, Droogmans G (2003) Amazing chloride channels: an overview. Acta physiologica Scandinavica 177:119-147. O'Brien EP, Ciryam P, Vendruscolo M, Dobson CM (2014) Understanding the influence of codon translation rates on cotranslational protein folding. Accounts of chemical research 47:1536-1544. Okiyoneda T, Barriere H, Bagdany M, Rabeh WM, Du K, Hohfeld J, Young JC, Lukacs GL (2010) Peripheral protein quality control removes unfolded CFTR from the plasma membrane. Science (New York, NY) 329:805-810. Pal R, Mamidi MK, Das AK, Bhonde R (2012) Diverse effects of dimethyl sulfoxide (DMSO) on the differentiation potential of human embryonic stem cells. Archives of toxicology 86:651-661. Papponen H, Nissinen M, Kaisto T, Myllyla VV, Myllyla R, Metsikko K (2008) F413C and A531V but not R894X myotonia congenita mutations cause defective endoplasmic reticulum export of the muscle-specific chloride channel CLC-1. Muscle nerve 37:317-325. Pelham HR (1986) Speculations on the functions of the major heat shock and glucose-regulated proteins. Cell 46:959-961. Pelham HR (1988) Evidence that luminal ER proteins are sorted from secreted proteins in a post-ER compartment. The EMBO journal 7:913-918. Peng YJ, Huang JJ, Wu HH, Hsieh HY, Wu CY, Chen SC, Chen TY, Tang CY (2016) Regulation of CLC-1 chloride channel biosynthesis by FKBP8 and Hsp90beta. Scientific reports 6:32444. Pickart CM (2001) Mechanisms underlying ubiquitination. Annual review of biochemistry 70:503-533. Picollo A, Pusch M (2005) Chloride/proton antiporter activity of mammalian CLC proteins ClC-4 and ClC-5. Nature 436:420-423. Pletjushkina OY, Fetisova EK, Lyamzaev KG, Ivanova OY, Domnina LV, Vyssokikh MY, Pustovidko AV, Alexeevski AV, Alexeevski DA, Vasiliev JM, Murphy MP, Chernyak BV, Skulachev VP (2006) Hydrogen peroxide produced inside mitochondria takes part in cell-to-cell transmission of apoptotic signal. Biochemistry Biokhimiia 71:60-67. Powers ET, Morimoto RI, Dillin A, Kelly JW, Balch WE (2009) Biological and chemical approaches to diseases of proteostasis deficiency. Annual review of biochemistry 78:959-991. Pusch M (2002) Myotonia caused by mutations in the muscle chloride channel gene CLCN1. Human mutation 19:423-434. Riordan JR (2008) CFTR function and prospects for therapy. Annual review of biochemistry 77:701-726. Saitoh F, Araki T (2010) Proteasomal degradation of glutamine synthetase regulates schwann cell differentiation. The Journal of neuroscience : the official journal of the Society for Neuroscience 30:1204-1212. Santos NC, Figueira-Coelho J, Martins-Silva J, Saldanha C (2003) Multidisciplinary utilization of dimethyl sulfoxide: pharmacological, cellular, and molecular aspects. Biochemical pharmacology 65:1035-1041. Saviane C, Conti F, Pusch M (1999) The muscle chloride channel ClC-1 has a double-barreled appearance that is differentially affected in dominant and recessive myotonia. The Journal of general physiology 113:457-468. Scheel O, Zdebik AA, Lourdel S, Jentsch TJ (2005) Voltage-dependent electrogenic chloride/proton exchange by endosomal CLC proteins. Nature 436:424-427. Shiber A, Ravid T (2014) Chaperoning proteins for destruction: diverse roles of Hsp70 chaperones and their co-chaperones in targeting misfolded proteins to the proteasome. Biomolecules 4:704-724. Shirane M, Nakayama KI (2003) Inherent calcineurin inhibitor FKBP38 targets Bcl-2 to mitochondria and inhibits apoptosis. Nature cell biology 5:28-37. Spang A (2009) On vesicle formation and tethering in the ER-Golgi shuttle. Current opinion in cell biology 21:531-536. Spiess C, Meyer AS, Reissmann S, Frydman J (2004) Mechanism of the eukaryotic chaperonin: protein folding in the chamber of secrets. Trends in cell biology 14:598-604. Steinmeyer K, Ortland C, Jentsch TJ (1991) Primary structure and functional expression of a developmentally regulated skeletal muscle chloride channel. Nature 354:301-304. Stolting G, Fischer M, Fahlke C (2014) CLC channel function and dysfunction in health and disease. Frontiers in physiology 5:378. Tang CY, Chen TY (2011) Physiology and pathophysiology of CLC-1: mechanisms of a chloride channel disease, myotonia. Journal of biomedicine biotechnology 2011:685328. Tartaglia GG, Pechmann S, Dobson CM, Vendruscolo M (2007) Life on the edge: a link between gene expression levels and aggregation rates of human proteins. Trends in biochemical sciences 32:204-206. Tsigelny I, Hotchko M, Yuan JX, Keller SH (2005) Identification of molecular determinants that modulate trafficking of DeltaF508 CFTR, the mutant ABC transporter associated with cystic fibrosis. Cell biochemistry and biophysics 42:41-53. Tsujino A, Kaibara M, Hayashi H, Eguchi H, Nakayama S, Sato K, Fukuda T, Tateishi Y, Shirabe S, Taniyama K, Kawakami A (2011) A CLCN1 mutation in dominant myotonia congenita impairs the increment of chloride conductance during repetitive depolarization. Neuroscience letters 494:155-160. Waghray M, Cui Z, Horowitz JC, Subramanian IM, Martinez FJ, Toews GB, Thannickal VJ (2005) Hydrogen peroxide is a diffusible paracrine signal for the induction of epithelial cell death by activated myofibroblasts. FASEB journal : official publication of the Federation of American Societies for Experimental Biology 19:854-856. Wakatsuki S, Saitoh F, Araki T (2011) ZNRF1 promotes Wallerian degeneration by degrading AKT to induce GSK3B-dependent CRMP2 phosphorylation. Nature cell biology 13:1415-1423. Wakatsuki S, Furuno A, Ohshima M, Araki T (2015) Oxidative stress-dependent phosphorylation activates ZNRF1 to induce neuronal/axonal degeneration. The Journal of cell biology 211:881-896. Walker VE, Atanasiu R, Lam H, Shrier A (2007) Co-chaperone FKBP38 Promotes HERG Trafficking. Journal of Biological Chemistry 282:23509-23516. Wang HQ, Nakaya Y, Du Z, Yamane T, Shirane M, Kudo T, Takeda M, Takebayashi K, Noda Y, Nakayama KI, Nishimura M (2005) Interaction of presenilins with FKBP38 promotes apoptosis by reducing mitochondrial Bcl-2. Human molecular genetics 14:1889-1902. Wang J, Maldonado MA (2006) The ubiquitin-proteasome system and its role in inflammatory and autoimmune diseases. Cellular molecular immunology 3:255-261. Wang X, Robbins J (2014) Proteasomal and lysosomal protein degradation and heart disease. Journal of molecular and cellular cardiology 71:16-24. Wieland FT, Gleason ML, Serafini TA, Rothman JE (1987) The rate of bulk flow from the endoplasmic reticulum to the cell surface. Cell 50:289-300. Wolins N, Bosshart H, Kuster H, Bonifacino JS (1997) Aggregation as a determinant of protein fate in post-Golgi compartments: role of the luminal domain of furin in lysosomal targeting. The Journal of cell biology 139:1735-1745. Wright MA, Aprile FA, Arosio P, Vendruscolo M, Dobson CM, Knowles TP (2015) Biophysical approaches for the study of interactions between molecular chaperones and protein aggregates. Chemical communications (Cambridge, England) 51:14425-14434. Xu D, Hay JC (2004) Reconstitution of COPII vesicle fusion to generate a pre-Golgi intermediate compartment. The Journal of cell biology 167:997-1003. Young JC, Agashe VR, Siegers K, Hartl FU (2004) Pathways of chaperone-mediated protein folding in the cytosol. Nature reviews Molecular cell biology 5:781-791. | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/67447 | - |
dc.description.abstract | 先天性肌肉強直症(myotonia congenita)是一種遺傳性的骨骼肌病變,是由於第七對染色體上的CLCN1基因發生突變,導致骨骼肌電壓敏感性氯離子通道CLC-1的功能改變。本實驗室先前利用yeast two-hybrid篩選技術,發現分子伴護蛋白FKBP8與泛素連接酶ZNRF1皆可以和CLC-1的C端區域產生交互作用。先前的實驗結果顯示FKBP8可能參與CLC-1在內質網的品質控管,進而提升CLC-1的總表現量和細胞膜上的表現量,至於FKBP8是否能夠直接穩定細胞膜上的CLC-1蛋白質表現則尚不清楚。另外,雖然ZNRF1被指出因為其N端具有豆蔻酸化(myristoylation)修飾,並且可以連結到細胞膜上,但ZNRF1是否會影響細胞膜上的CLC-1蛋白質表現也仍未確定。因此,本篇論文的目的,是想要探討FKBP8與ZNRF1是否可能參與CLC-1在細胞膜上的的品質控管。 首先,我們利用次細胞分群法(subcellular fractionation)去探討FKBP8在有無CLC-1的情況下的次細胞分布狀況。我們發現在與CLC-1共同表現的情形下,部分的FKBP8會從原本存在於內質網的分層轉為分布到細胞膜的分層。接著我們也利用差速離心法(differential centrifugation)觀察內質網、高基氏體及細胞膜蛋白質的分布趨勢。我們發現在與CLC-1共同表現的情形下,FKBP8在高基氏體及細胞膜的分布比例會明顯增加,也觀察到大量表現FKBP8時,CLC-1在內質網的分佈比例下降,而在細胞膜的分佈比例上升。由此推測,FKBP8可伴隨著CLC-1運出內質網,並一起經由高基氏體運送至細胞膜。 當我們利用shRNA壓制內生性ZNRF1的表現,會使CLC-1的蛋白質總表現量上升。當我們利用過氧化氫提升HEK293T細胞內生性ZNRF1活性時, CLC-1的總表現量也會明顯下降。此外,在有過氧化氫的情況下,大量表現ZNRF1則會進一步降低CLC-1的總表現量。我們還利用brefeldin A (BFA)來觀察細胞膜上CLC-1的代謝時程(turn over rate),初步的結果顯示,利用Ub-K0干擾聚泛素鏈(polyubiquitin chain)的形成,會增加CLC-1於細胞膜上的穩定性。這些結果顯示,ZNRF1對細胞膜上CLC-1的降解效果會因細胞承受過氧化壓力而進一步提升,而且ZNRF1有可能經由對CLC-1催化聚泛素鏈的方式,促進細胞膜上的CLC-1降解。 | zh_TW |
dc.description.abstract | Myotonia congenita is a hereditary skeletal muscle disease that is linked to mutations in the human CLCN1 gene, which encode the voltage-gated chloride channel CLC-1. Previously, through yeast two-hybrid screening, our lab demonstrated that cochaperone FKBP8 and E3 ligase ZNRF1 may interact with the C-terminal region of CLC-1. Previous data from our lab further showed that FKBP8 may be involved in the protein quality control system in endoplasmic reticulum (ER) and therefore increase both the total protein and the plasma membrane protein level of CLC-1. Whether FKBP8 could directly stabilize the protein level of CLC-1 at the plasma membrane remain unknown. On the other hand, although the E3 ubiquitin ligase ZNRF1may be anchored at the plasma membrane via N-myristoylation, it is still an open question whether ZNRF1 could regulate CLC-1 protein level at the plasma membrane. In this thesis, we aim to understand whether FKBP8 and ZNRF1 may involved in the protein quality control of CLC-1 at the plasma membrane. First, we investigated the subcellular distribution of FKBP8 by using subcellular fractionation. We found that, when coexpressed with CLC-1, a significant fraction of FKBP8 was translocated from the endoplasmic reticulum fraction to the plasma membrane fraction. Next, by performing differential centrifugation, we examined the subcellular distribution of FKBP8 between endoplasmic reticulum and Golgi apparatus. We found that, upon coexpression with CLC-1, there was a significant increase in the distribution of FKBP8 in the Golgi apparatus fraction. Moreover, the same fractionation and centrifugation experiment also showed that coexpression with FKBP8 led to a notable enhamcement of the membrane localization of CLC-1. Thus, our data suggest that, together with CLC-1, FKBP8 may be exported out of endoplasmic reticulum and transported to Golgi apparatus and finally to plasma membrane. shRNA suppression of endogenous ZNRF1expression resulted in significant upregelation of CLC-1 protein level. Activation of endogenous ZNRF1 activity with hydrogen peroxide led to notable reduction in CLC-1 protein level. Additionally, in the presence of hydrogen peroxide, coexpression with ZNRF1 significantly down-regulated CLC-1 protein expression. We also investigated the turn over rate of CLC-1 at the plasma membrane by using brefeldin A (BFA) treatment. We observed that overexpression of Ub-K0, which prevent the formation of polyubiquitin chain, seemed to stabilize CLC-1 protein level at the plasma membrane. Together, these results suggest that the degradation effect of ZNRF1 on CLC-1 is enhanced under oxidative stress, and that ZNRF1 may enhance protein degradation of CLC-1 at the plasma membrane by promoting CLC-1 polyubiquitination. | en |
dc.description.provenance | Made available in DSpace on 2021-06-17T01:32:36Z (GMT). No. of bitstreams: 1 ntu-106-R04441004-1.pdf: 3367446 bytes, checksum: 748e1265db4f21f6d4e1f33c09016b50 (MD5) Previous issue date: 2017 | en |
dc.description.tableofcontents | 目錄
中文摘要 i Abstract iii 目錄 v 圖表目錄 viii 第一章 導論 1 1.1 氯離子通道 1 1.2 CLC-1的結構與電生理特性 2 1.3 CLC-1的生理功能 2 1.4 先天性肌肉強直症(Myotonia congenita) 3 1.5蛋白質平衡(proteostasis) 4 1.6蛋白質的摺疊與堆積 5 1.7分子伴護蛋白的基本概念 5 1.8分子伴護蛋白參與蛋白質的摺疊 6 1.9 FKBP8 (FK506 Binding Protein 8) 7 1.10蛋白質降解系統 8 1.11蛋白質泛素化(protein ubiquitination) 9 1.12 ZNRF1 (Zinc finger and RING finger protein 1) 10 1.13蛋白質品質控管(protein quality control) 11 1.14 Peripheral quality control 11 1.15蛋白質運送機轉 12 1.16研究目的 14 第二章 材料與方法 16 2.1 DNA construct 16 2.2 Cell culture 17 2.3 DNA transfection 17 2.4 Western Blot 18 2.5次細胞分群法(Subcellular fractionation) 19 2.6差速離心法(Differential centrifugation) 19 2.7 H2O2 treatment 20 2.8 Lentivirus production and infection 21 2.9 Brefeldin A treatment 21 2.10 Biotinylation 22 2.11統計分析 22 第三章 結果 23 3.1以次細胞分群法(subcellular fractionation)觀察FKBP8及CLC-1在內質網和細胞膜上的分布 23 3.1.1 大量表現CLC-1會增加FKBP8在細胞膜分布的比例 24 3.1.2 大量表現FKBP8會增加CLC-1在細胞膜分布的比例 25 3.2以差速離心法(diffenertial centrifugation)觀察FKBP8及CLC-1在內質網和高基氏體的分布 25 3.2.1大量表現CLC-1會增加FKBP8在高基氏體分布的比例 26 3.2.2大量表現FKBP8會增加CLC-1在高基氏體分布的比例 27 3.3泛素連接酶ZNRF1對CLC-1蛋白質表現量的影響 28 3.4 ZNRF1的dominant-negative mutant (C184A)對CLC-1蛋白質表現量的影響 29 3.5 H2O2活化ZNRF1的活性並影響CLC-1的蛋白質表現量 29 3.6大量表現Ub-K0會增加細胞膜上CLC-1的穩定性 30 第四章 討論 31 4.1次細胞分群法和差速離心實驗 31 4.2 FKBP8原本於細胞中的分布狀況以及大量表現CLC-1會改變FKBP8於細胞中的分布狀況 33 4.3 FKBP8對於CLC-1的可能調控角色 33 4.4 ZNRF1於細胞中的分布及對CLC-1表現量之影響 35 4.5 ZNRF1的活化及調控蛋白質降解的機制 37 4.6 Ub-K0可能增加細胞膜上CLC-1的穩定性 38 4.7未解決的問題 39 4.8未來實驗方向 39 結論 42 圖表 43 附圖 65 參考資料 69 | |
dc.language.iso | zh-TW | |
dc.title | 人類第一型氯離子通道蛋白於細胞膜上蛋白質穩定性之調控 | zh_TW |
dc.title | Protein stability of human CLC-1 channels at the plasma membrane | en |
dc.type | Thesis | |
dc.date.schoolyear | 105-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 鄭瓊娟,胡孟君,徐立中 | |
dc.subject.keyword | 人類第一型氯離子通道蛋白,蛋白質穩定性,細胞膜, | zh_TW |
dc.subject.keyword | human CLC-1 channels,protein stability,plasma membrane, | en |
dc.relation.page | 77 | |
dc.identifier.doi | 10.6342/NTU201702471 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2017-08-03 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 生理學研究所 | zh_TW |
顯示於系所單位: | 生理學科所 |
文件中的檔案:
檔案 | 大小 | 格式 | |
---|---|---|---|
ntu-106-1.pdf 目前未授權公開取用 | 3.29 MB | Adobe PDF |
系統中的文件,除了特別指名其著作權條款之外,均受到著作權保護,並且保留所有的權利。