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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 梁博煌 | |
dc.contributor.author | Che-Kuan Chao | en |
dc.contributor.author | 趙哲寬 | zh_TW |
dc.date.accessioned | 2021-06-07T17:59:03Z | - |
dc.date.copyright | 2012-08-18 | |
dc.date.issued | 2012 | |
dc.date.submitted | 2012-08-09 | |
dc.identifier.citation | Abe Y, Yoon SO, Kubota K, Mendoza MC, Gygi SP, Blenis J (2009) p90 Ribosomal S6 Kinase and p70 Ribosomal S6 Kinase Link Phosphorylation of the Eukaryotic Chaperonin Containing TCP-1 to Growth Factor, Insulin, and Nutrient Signaling. J Biol Chem. 284(22), 14939–14948.
Abolhoda A, Wilson AE, Ross H, Danenberg PV, Burt M, Scotto KW (1999) Rapid Activation of MDR1 Gene Expression in Human Metastatic Sarcoma after in Vivo Exposure to Doxorubicin. Clin Cancer Res. 5, 3352–3356. Amit M, Weisberg SJ, Nadler-Holly M, McCormack EA, Ester Feldmesser E, Kaganovich D, Willison KR, Horovitz A (2010) J. Mol. Biol. 401, 532–543. Baguley BC (2010) Multiple Drug Resistance Mechanisms in Cancer. Mol Biotechnol 46, 308–316. Besnard A, Galan-Rodriguez B, Vanhoutte P, Caboche J (2011) Elk-1 a transcription factor with multiple facets in the brain. Front Neurosci. 5, 1-11. Blade K, Menick DR, Cabral F (1999) Overexpression of class I, II or IVb β-tubulin isotypes in CHO cells is insufficient to confer resistance to paclitaxel. J Cell Sci. 112, 2213-2221. Bradley G, Ling V (1994) P-glycoprotein, multidrug resistance and tumor progression. Cancer Metast Rev. 13, 223-233. Camasses A, Bogdanova A, Shevchenko A, Zachariae W (2003) The CCT Chaperonin Promotes Activation of the Anaphase-Promoting Complex through the Generation of Functional Cdc20 (2003) Mol cell. 12, 87–100 Casalou C, Cyrne L, Rosa MR, Soares H (2001) Microtubule cytoskeleton perturbation induced by taxol and colchicines affects chaperonin containing TCP-1 (CCT) subunit gene expression in Tetrahymena cells. Biochim Biophys Acta. 1522, 9-21. Coghlin C, Carpenter B, Dundas SR, Lawrie LC, Telfer C, Murray GI (2006) Characterization and over-expression of chaperonin t-complex proteins in colorectal cancer. J Pathol. 210: 351–357. Dekker C (2010) On the role of the chaperonin CCT in the just-in-time assembly process of APC/CCdc20. FEBS Lett. 584, 477–481. Dekker C, Roe SM, McCormack EA, Beuron F, Pear LH, Willison KR (2011) The crystal structure of yeast CCT reveals intrinsic asymmetry of eukaryotic cytosolic chaperonins. EMBO J. 30, 3078–3090. Ferrandina G, Zannoni GF, Martinelli E, Paglia A, Gallotta V, Mozzetti S, Scambia G, Ferlini C (2006) Class III | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/16043 | - |
dc.description.abstract | 近年來,對於癌症的運作機制已經越來越被研究透徹,治療方式也持續地有突破性的發展。然而,對於各式抗癌藥物,在長時間的治療下,癌症細胞所出現的抗藥性,仍然是癌症治療上難以突破的癥結點。Chaperonin containing t-complex polypeptide 1 (CCT) subunit β (CCT-β) 被發現在許多不同種類的癌症以及具抗藥性的癌症細胞中,都有過量表現的狀況(Lin et al., 2009)。因此在本論文中,我主要的目的即為觀察過量表現的CCT-β在癌症細胞中的特性,為此,我首先建立了CCT-β過量表現以及CCT-β抑制表現的兩種穩定細胞株,以觀察CCT-β在癌症細胞中所造成的影響。
首先檢視的,是CCT的蛋白質折疊能力。CCT的兩種受質actin與tubulin,是細胞骨架actin filament與microtubule不可或缺的組成物。在CCT-β過量表現的穩定細胞株中,actin與tubulin的兩種亞型均發現有顯著增加的情形,顯示過量表現的CCT-β能夠增加actin與tubulin的折疊及成熟。另一方面,CCT- | zh_TW |
dc.description.abstract | Chaperonin containing t-complex polypeptide 1 (CCT) is a molecular chaperonin that facilitates the protein folding process in eukaryotic cytosol. The expression level of the CCT-β subunit was found elevated in a variety of drug-resistant tumor types (Lin et al., 2009). In this study, the main purpose is to examine the characteristics of up-regulating CCT-β in cancer cells.
Initially, the classic substrates of CCT, actin and tubulin, were examined. Overexpression of CCT-β resulted in significantly elevated expression of both molecules, while knockdown of CCT-β gave no apparent difference. Therefore, up-regulation of CCT-β in cancer cells promotes more vigorous cytoskeleton formation. Next, other subunits of CCT were examined. In both CCT-β overexpress and knockdown stable clone, as well as paclitaxel-resistant cells, some but not all of the eight subunits were found to have synchronous changes with CCT-β in final protein amount. This phenomenon could be explained by the sub-assemblies during the complete CCT complex formation. Abnormally high level of CCT-β in cancer cells may hence break the balance of CCT subunits and affect the folding efficiency. On the other hand, expression level of CCT-β also correlated with the phosphorylation of Elk1, a transcription factor which is usually activated by MAPK family member p38. CCT-β up-regulation was found to induce the multi-drug resistance in cancer cells and stable clone, and the expression of multidrug-resistant protein (MDR1) was induced by CCT-β up-regulation. RNA interference experiments revealed that Elk1 knockdown effectively repressed MDR1 expression and drug resistance in CCT-β up-regulated cells. Interestingly, CCT-β seemed able to affect the phosphorylation level of Elk1, indicating that MDR1 expression and drug resistance in cancer cells may be controlled by MAPK signaling and stress-induced CCT-β. Moreover, a recently screened compound called R379603 showed significant apoptosis effects on CCT-β up-regulated cancer cells. R379603 led to CCT-β up-regulated cell death by introducing ER stress in cells, causing caspase activation and apoptosis. Since this is the route bypassing MDR1-related drug resistance, R379603 may therefore be a promising candidate for treating patients with elevated CCT-βand MDR1 expression. Overall, these studies uncovered two possible roles of CCT-β in CCT-β up-regulated cancer cells: enhancing folding efficiencies of cytoskeleton molecules, and inducing multi-drug resistance. Since CCT-β may be leading CCT assembly and substrate folding, the importance of CCT-β should be further investigated. | en |
dc.description.provenance | Made available in DSpace on 2021-06-07T17:59:03Z (GMT). No. of bitstreams: 1 ntu-101-R99b46009-1.pdf: 3759094 bytes, checksum: aecbddc5ee4f511f5fc97781926bf589 (MD5) Previous issue date: 2012 | en |
dc.description.tableofcontents | 中文摘要 … 1
ABSTRACT …3 ABBREVIATIONS … 5 (1) INTRODUCTION …7 1.1 Chaperonin containing t-complex polypptide 1 (CCT) … 7 1.1.1 Classification … 7 1.1.2 Functions… 8 1.2 Tubulin and microtubule assembly … 9 1.3 Paclitaxel … 11 1.4 Mechanisms of paclitaxel resistance … 12 1.5 Elk1 …13 1.6 MDR1 … 15 1.7 Previous study and present work … 17 (2) MATERIALS AND METHODS … 19 2.1 Cell lines and culture conditions … 19 2.2 Preparation of CCT-β transfection and stable clone selection … 19 2.3 Preparation of small interfering RNA transfection … 20 2.4 shRNA lentiviral partical transduction … 21 2.5 Drug sensitivity assay … 22 2.6 Preparation of cell lysate and isolation of nuclear and cytoplasmic fractionations … 23 2.7 SDS-PAGE and Western blot analysis … 24 2.8 Flow cytometry …25 (3) RESULTS … 26 3.1 Paclitaxel-domesticated A549 cell line possesses multi-drug resistance … 26 3.2 CCT-β is overexpressed in multi-drug resistant cancer cell lines … 26 3.3 CCT-β overexpression stable clone showed more actin and tubulin synthesis … 27 3.4 Repressed CCT-β expression did not affect the amount of actin and tubulin … 28 3.5 Some protein quantities of CCT subunits were elevated in CCT-β overexpression cells … 29 3.6 Some protein quantities of CCT subunits were decreased in CCT-β knockdown cells … 31 3.7 p38 and Elk1 were highly phosphorylated in CCT-βoverexpression stable clone … 31 3.8 CCT-β knock down stable clone showed decreased phosphorylation of Elk1 … 32 3.9 CCT-β overexpression induced MDR1 expression …32 3.10 MDR1 knock down by siRNA showed restored vulnerability to anticancer drugs … 33 3.11 Knockdown of Elk1 showed decreased MDR1 expression and vulnerability to anticancer drugs … 33 3.12 HSF1 was translocated into nucleus in TR cells … 34 3.13 Compound targeting CCT-β/β-tubulin complex showed cytotoxic effect on CCT-βoverexpressed cell lines … 35 (4) DISCUSSION … 37 REFERENCE … 49 FIGURE … 57 SUPPLEMENTARY … 79 | |
dc.language.iso | en | |
dc.title | 癌細胞中調控CCT-β表現程度對伴侶蛋白折疊與抗藥性之影響 | zh_TW |
dc.title | Regulating CCT-β expression level affects chaperonin folding machinery and drug resistance in cancer cell | en |
dc.type | Thesis | |
dc.date.schoolyear | 100-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 張茂山,張?仁 | |
dc.subject.keyword | 抗藥性癌症細胞,多重抗藥性蛋白,伴侶蛋白,細胞骨架,新藥開發, | zh_TW |
dc.subject.keyword | chaperonin,CCT-β,Multidrug resistance protein 1,tubulin, | en |
dc.relation.page | 85 | |
dc.rights.note | 未授權 | |
dc.date.accepted | 2012-08-09 | |
dc.contributor.author-college | 生命科學院 | zh_TW |
dc.contributor.author-dept | 生化科學研究所 | zh_TW |
顯示於系所單位: | 生化科學研究所 |
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