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研究生:王映酈
研究生(外文):Ying-Li Wang
論文名稱:斑馬魚OPTN基因之鑑定與功能分析
論文名稱(外文):Identification and functional analysis of zebrafish OPTN
指導教授:楊建洲
學位類別:碩士
校院名稱:中山醫學大學
系所名稱:生物醫學科學學系碩士班
學門:生命科學學門
學類:生物化學學類
論文種類:學術論文
論文出版年:2018
畢業學年度:106
語文別:中文
論文頁數:83
中文關鍵詞:斑馬魚青光眼
外文關鍵詞:zebrafishglaucoma optineurin optn
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  青光眼是全球引發雙眼失明的疾病主因之一,其特點是導致視網膜神經節細胞和神經纖維層逐漸病變且薄化,初期伴隨周邊的视野缺損,最終導致全盲。依病因可分為原發性和續發性,原發性意謂自發性。先前有研究指出原發性開放青光眼 (primary open-angle glaucoma) 與 optineurin 的突變有關,另外肌萎縮性脊髓侧索硬化症還有佩吉特氏病也與 OPTN 相關。Optineurin 是一種多功能蛋白,有一些捲曲螺旋和 C 端與泛素結的結構,其參與在許多訊息傳遞的路徑上,像是 NF-κB 的活化、自噬作用或者是自體免疫反應。青光眼發病的主要症狀為視野缺損、視力喪失,主要是由於神經節細胞中的視神經死亡。Optineurin 作為其中的關鍵蛋白,包括參與囊泡分泌和回收的運輸路徑、NF-κB 訊號、有絲分裂、高基氏體機轉和抗病毒信號,影響甚鉅,但此相關研究仍缺乏,機轉不明,因此釐清此基因相關的功能及分析,為本篇研究的主要目的。
  本研究針對斑馬魚 OPTN 基因進行研究。如以半定量 RT-PCR觀察斑馬魚早期胚胎各個發育時期的基因表現量,發現表現量隨著發育時期,呈正相關,接著利用全胚胎原位雜交的方式觀察基因在發育後期表現情形。為瞭解 OPTN 在斑馬魚體內的生理功能,我們利用反股寡核酸 (Antisense Morpholino oligonucleotides,MO) 顯微注射抑制 OPTN 基因的功能,為了確認 MO 對 OPTN 基因的抑制是專性的,我們建構了帶有紅色螢光蛋白的 OPTN 表現質體 (OPTN- TaqRFP),藉由統計螢光表現率來探討 MO 的專一性,結果發現OPTN 的 MO 可以有效的抑制紅色螢光表現,因此我們確認其具專一性 knockdown OPTN 的基因表現。取 3dpf 注射 MO 的斑馬魚,進行組織切片,量測其眼視網膜厚度變化情形,此數據可供往後研究原發性開放青光眼之早期病程。
Glaucoma is a leading cause of irreversible visual impairment and blindness resulting from progressive degeneration of retinal ganglion cells (RGCs) and optic neuropathy. Optineurin was originally identified as a gene responsible for primary open-angle glaucoma. The gene, OPTN, codes for the protein optineurin, which is involved in a variety of functions including regulation of endocytic trafficking, autophagy, immune response, mitosis and NF-κB signal transduction. Many missense mutations in optineurin have been reported and the association of the mutation with the disease varies in different populations. Optineurin is localized to pathological structures also seen in several neurodegenerative diseases such as amyotrophic lateral sclerosis, Alzheimer''s disease, Parkinson''s disease, etc. However, functional alterations caused by mutations in optineurin are poorly understood.
In this study, we investigated the physiological functions of OPTN in the zebrafish embryonic development. RT-PCR analysis of zebrafish OPTN transcripts were revealed significantly in the 120hpf embryos. Meanwhile, whole-mount in situ hybridization (WISH) signals were also observed in the 120hpf embryos. In addition, we established the fusion protein construct OPTN-TagRFP that was injected into embryos in the presence or absence of OPTN-targeting morpholinos. We have found that OPTN- targeting MO was able to specifically knockdown the expression of TagRFP-tagged OPTN fusion proteins in a dose-dependent manner. These
data may be useful for studying early POAG pathophysiology.
目錄
中文摘要       2
英文摘要       4
目錄         5
縮寫表        6
第一章 序論     8
第二章 材料與方法 18
第三章 結果    37
第四章 討論    43
第五章 未來展望  49
參考文獻      50
圖表        54
附錄一、附圖    69
附錄二、藥品製備  80
Resnikoff S, Pascolini D, Etya''ale D, et al. Global data on visual impairment in the year 2002. Bull World Health Organ. 2004 Nov;82:844-51.
Vrabec JP, Levin LA. The neurobiology of cell death in glaucoma. Eye (Lond). 2007 Dec;21 Suppl 1:S11-4.
Weinreb RN, Khaw PT. Primary open-angle glaucoma. Lancet. 2004 May 22;363:1711-20.
Vasiliou V, Gonzalez FJ. Role of CYPIBl in glaucoma. Annu Rev Pharmacol Toxicol.2008;48:333-58.
Quigley HA. Glaucoma. Lancet. 2011 Apr 16;377:1367-77.
Hilal L, Boutayeb S, Serrou A, et al. Screening of CYPIBl and MYOC in Moroccan families with primary congenital glaucoma: three novel mutations in CYPIBI. Mol Vis. 2010;16:1215-26.
Melki R, Colomb E, Lefort N, et al. CYP1BI mutations in French patients with earlyonset primary open-angle glaucoma. J Med Genet. 2004 Sep;41:647-51.
Rezaie T, Child A, Hitchings R, et al. Adult-onset primary open- angle glaucoma caused by mutations in optineurin. Science. 2002 Feb 8;295:1077-9.
Ayala-Lugo RM, Pawar H, Reed DM, et al. Variation in optineurin (OPTN) allele frequencies between and within populations. Mol Vis. 2007;13:151-63.
Leung YF, Fan BJ, Lam DS, et al. Different optineurin mutation pattern in primary open-angle glaucoma. Invest Ophthalmol Vis Sci. 2003 Sep;44:3880-4.
Zhu G, Wu CJ, Zhao Y, et al. Optineurin negatively regulates TNFalpha- induced NF-kappaB activation by competing with NEMO for ubiquitinated RIP. Curr Biol.2007 Aug 21;17:1438-43.
Sahlender DA, Roberts RC, Arden SD, et al. Optineurin links myosin VI to the Golgi complex and is involved in Golgi organization and exocytosis. J Cell Biol. 2005 Apr 25;169:285-95.
Chalasani ML, Swarup G, Balasubramanian D. Optineurin and its mutants: molecules associated with some forms of glaucoma. Ophthalmic Res. 2009;42:176-84.
Morton S, Hesson L, Peggie M, et al. Enhanced binding of TBKI by an optineurin mutant that causes a familial form of primary open angle glaucoma. FEBS Lett. 2008 Mar 19;582:997-1002.
Vaibhava V, Nagabhushana A, Chalasani ML, et al. Optineurin mediates negative regulation of Rab8 function by TBCID17, a GTPase activating protein. J Cell Sci. 2012 Aug 1.
Ying H, Yue BY. Cellular and molecular biology of optineurin. Int Rev Cell Mol Biol.2012;294:223-58.
Hutagalung AH, Novick PJ. Role of Rab GTPases in membrane traffic and cell physiology. Physiol Rev. 2011 Jan;91:119-49.
Henry L, Sheff DR. Rab8 regulates basolateral secretory, but not recycling, traffic at the recycling endosome. Mol Biol Cell. 2008 May;19:2059-68.
Hattula K, Furuhjelm J, Tikkanen J, et al. Characterization of the Rab8-specific membrane traffic route linked to protrusion formation. J Cell Sci. 2006 Dec 1;119:4866-77.
Peranen J. Rab8 GTPase as a regulator of cell shape. Cytoskeleton (Hoboken). 2011 Oct;68:527-39.
Chibalina MV, Poliakov A, Kendrick-Jones J, et al. Myosin VI and optineurin are required for polarized EGFR delivery and directed migration. Traffic. 2010 Oct; 11:1290-303.
Bond LM, Peden AA, Kendrick-Jones J, et al. Myosin VI and its binding partner optineurin are involved in secretory vesicle fusion at the plasma membrane. Mol Biol Cell. 2010 Jan 1;22:54-65.
Gilmore TD. Introduction to NF-kappaB: players, pathways, perspectives. Oncogene.2006 Oct 30;25:6680-4.
Skaug B, Jiang X, Chen ZJ. The role of ubiquitin in NF-kappaB regulatory pathways.Annu Rev Biochem. 2009;78:769-96.
Trompouki E, Hatzivassiliou E, Tsichritzis T, et al. CYLD is a deubiquitinating enzyme that negatively regulates NF-kappalB activation by TNFR family members. Nature. 2003 Aug 14;424:793- 6.
Kovalenko A, Chable-Bessia C, Cantarella G, et al. The tumour suppressor CYLD negatively regulates NF-kappaB signalling by deubiquitination. Nature. 2003 Aug 14:424:801-5.
Sun SC. CYLD: a tumor suppressor deubiquitinase regulating NF- kappaB activation and diverse biological processes. Cell Death Differ. 2010 Jan; 17:25-34.
Sudhakar C, Nagabhushana A, Jain N, et al. NF-kappaB mediates tumor necrosis factor alpha-induced expression of optineurin, a negative regulator of NF-kappaB. PLoS One. 2009;4:e5114.
De Marco N, Buono M, Troise F, et al. Optineurin increases cell survival and translocates to the nucleus in a Rab8-dependent manner upon an apoptotic stimulus. J Biol Chem. 2006 Jun 9;281:16147-56.
Shen X, Ying H, Qiu Y, et al. Processing of optineurin in neuronal cells. J Biol Chem. 2010 Feb 4;286:3618-29.
Chalasani ML, Radha V, Gupta V, et al. A glaucoma-associated mutant of optineurin selectively induces death of retinal ganglion cells which is inhibited by antioxidants. Invest Ophthalmol Vis Sci. 2007 Apr;48;1607-14.
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