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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 朱家瑩(Chia-Ying Chu) | |
| dc.contributor.author | Yen-Chun Chen | en |
| dc.contributor.author | 陳妍君 | zh_TW |
| dc.date.accessioned | 2021-06-08T00:12:12Z | - |
| dc.date.copyright | 2013-08-09 | |
| dc.date.issued | 2013 | |
| dc.date.submitted | 2013-08-05 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/17425 | - |
| dc.description.abstract | RCK/p54是DEAD-box蛋白質的一種，在轉錄後調控(post-transcriptional regulation)基因表現扮演重要角色。在酵母菌、非洲爪蟾、以及果蠅中的研究顯示出RCK/p54的同源基因會參與基因的轉譯抑制(translational repression)進而調控基因表現。在人類miRNA路徑中，RCK/p54會與Ago2及其他蛋白質形成靜默蛋白複合體(miRNA-induced silencing complex)，抑制目標RNA轉譯或是將其儲存。在人類大腸癌研究發現RCK/p54大量表現於癌細胞中。除此之外，RCK/p54也高度表現在C型肝炎引起的肝癌細胞中。然而，RCK/p54在癌症中所扮演的角色及其機制皆不甚清楚。為了探討RCK/p54於癌症中的所扮演的角色及其機制，我們利用次世代定序以及定量蛋白質體分析與RCK/p54蛋白質結合的RNA，及降低RCK/p54表現量後所造成的下游RNA及蛋白質表現量變化。研究結果顯示癌細胞的轉移能力會隨著RCK/p54過量表現而隨之提高。此外，藉由比對RCK/p54蛋白質所結合的RNA與蛋白質體的資料，我們發現RCK/p54可能直接經由轉譯抑制調控NUDT21以及GPRC5A兩個基因的蛋白表現。經由次世代定序分析去除RCK/p54時HeLa細胞的基因表現變化圖譜，發現當細胞去除RCK/p54時，MAGEE1的表現量上升，顯示其表現可能間接由RCK/p54調控。本研究顯示細胞轉移程度與RCK/p54表現量成正相關，並且可能是經由調控NUDT21、GPRC5A以及MAGEE1進而影響癌症細胞的惡性化。 | zh_TW |
| dc.description.abstract | RCK/p54 is a member of DEAD-box protein family, and it plays a role in post-transcriptional regulation. Previous studies of RCK/p54 homologues in yeast, Xenopus and Drosophila showed that RCK/p54 participated in regulation of translational repression and moderates RNA stability. RCK/p54 is also involved in miRNA pathway. It interacts with Ago2 in miRNA-induced silencing complex (miRISC), which represses mRNA translation or stores target mRNA in processing body (P-body). Several studies in cancer biology showed that RCK/p54 is highly expressed in tumors. However, the detail mechanism of RCK/p54 in regulation of cancer progression is unrevealed. In this study, we aim to identify the role of RCK/p54 in cancer metastasis. We found out that migration of cancer cell was increased in cells overexpressing RCK/p54. Comparing the deep sequencing result of RCK/p54-associated mRNA and the change of proteomic profile in RCK/p54- depleted cell, we found that RCK/p54 may directly regulate NUDT21 and GPRC5A. MAGEE1 mRNA was increased in RCK/p54-depleted cell, but did not associate with RCK/p54, suggesting that MAGEE1 is a potential downstream target of RCK/p54. These findings suggest that RCK/p54 protein regulates cancer metastasis through various pathways and the potential targets or downstream of RCK/p54 were identified. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-08T00:12:12Z (GMT). No. of bitstreams: 1 ntu-102-R99b41017-1.pdf: 9890870 bytes, checksum: d6519ac2460ec2175065e61283d04d64 (MD5) Previous issue date: 2013 | en |
| dc.description.tableofcontents | 口試委員審定書 i
誌謝 ii 摘要 iii Abstract iv 1. Introduction 1 1.1 Discovery of RCK/p54 1 1.2 DEAD-box proteins and post-transcriptional control of gene expression 1 1.3 Functions of RCK/p54 domains and motifs. 2 1.4 RCK/p54 participates in miRNA pathway. 3 1.5 The role of RCK/p54 in diseases. 4 2. Materials and methods 6 2.1 DNA constructs and siRNAs 6 2.2 Cell culture and transfection 6 2.3 Immunoprecipitation and Western blot 6 2.4 In vitro invasion and migration assays 7 2.5 RNA extraction, reverse transcription and real-time polymerase chain reaction (real-time PCR). 8 2.6 Next-generation sequencing (NGS) 8 2.7 Microarray 9 2.8 Quantitative proteomics 10 3. Results 11 3.1 RCK/p54 is highly expressed in highly malignant lung adenocarcinoma cell line. 11 3.2. The N-terminal RCK/p54 regulates the migration or invasion abilities of lung cancer cell lines 12 3.3 RCK/p54-Ago2 interaction in microRNA pathway was not involved in the regulation of cell migration. 14 3.4 Migration activity of HeLa cell was increased with overexpression of RCK/p54 15 3.5 Identification of RCK/p54-associated genes through NGS was available. 15 3.6 NUDT21 and GPRC5A were associated with RCK/p54 and the their expression of protein level were up-regulated in RCK/p54-deficient cell 17 3.7 Expression of MAGEE1 was increased in RCK/p54-depleted cell. 19 4. Discussion 21 4.1 Expression level of RCK/p54 in cells affects migration and invasion ability of cells. 21 4.2 The RCK/p54-regulated cell migration was not related with RCK/p54-Ago2 interaction. 21 4.3 ATP-binding ability of RCK/p54 was required for regulation of metastasis. 22 4.4 Protein expression of NUDT21 and GPRC5A were regulated by RCK/p54 directly. 23 4.5 The network of RCK/p54 regulated genes in cancer. 24 5. References 28 6. Figures and tables 33 7. Supplementary data 50 | |
| dc.language.iso | en | |
| dc.title | 人類RNA靜默複合體(RISC)組成蛋白質參與癌化進程之功能性探討 | zh_TW |
| dc.title | Functional Analysis of RCK/p54, a Human RISC Component, in Cancer Progression. | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 101-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 李心予(Hsinyu Lee),黃偉邦(Wei-Pang Huang),朱家瑜(Chia-Yu Chu) | |
| dc.subject.keyword | 轉錄後調控,RCK/p54,轉錄抑制,癌症,惡性化, | zh_TW |
| dc.subject.keyword | post-translational repression,RCK/p54,translational repression,cancer,cell migration, | en |
| dc.relation.page | 53 | |
| dc.rights.note | 未授權 | |
| dc.date.accepted | 2013-08-06 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 動物學研究所 | zh_TW |
| 顯示於系所單位： | 動物學研究所 | |
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