揭示CEP55基因在大腸直腸癌轉移中所扮演的角色

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林揚芷(Yang-Chih Lin) 查詢紙本館藏

畢業系所

系統生物與生物資訊研究所

論文名稱

揭示CEP55基因在大腸直腸癌轉移中所扮演的角色
(Clarifying role of CEP55 in colorectal cancer metastasis)

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摘要(中)

大腸直腸癌好發率高居世界第三，且轉移性大腸直腸癌會使治療更加困難。本研究討論CEP55在大腸直腸癌扮演的角色，以釐清轉移性大腸直腸癌的機制。CEP55是一個轉錄中心體蛋白，在細胞增生上扮演重要角色，且已被發現在肝癌、肺癌、頭頸癌和鼻咽癌中過度表現。此研究包含三個部分，第一個部分是利用四個獨立公開的數據組來比較大腸直腸癌患者與健康族群的CEP55的表現量。第二部分是利用免疫組織染色程度來評估110例大腸直腸癌的CEP55表現量和淋巴結擴散程度的相關性。第三部分，藉由質體轉染及小分子干擾核糖核酸來改變HCT116細胞中CEP55的表現程度，再利用高通量信息RNA定序來得到CEP55的完整調控資訊。結果顯示，CEP55在大腸直腸癌的表現量升高。且CEP55的表現量與淋巴結擴散程度呈正相關性。最後結果顯示CEP55能促進細胞的爬行，並利用高通量定序技術，揭示跟CEP55共同調控的基因、功能及路徑。以上的整體結果能幫助深入了解轉移性大腸直腸癌的分子機制，也示意CEP55可能做為治療轉移性大腸直腸癌的標靶。

摘要(英)

Colorectal cancer (CRC) has the third highest incidence rate among all cancers worldwide and becomes radically more difficult to treat when it is metastatic. Here, to provide insight into the mechanisms underlying CRC metastasis, we describe the role in CRC metastasis of CEP55, which is the encoding gene of a centrosomal protein essential to proliferation and has been reported to be overexpressed in hepatocellular carcinoma, lung cancer, head and neck cancer and nasopharyngeal carcinomas. The study comprises three steps. The first step was to measure the relative expression levels of CEP55 in CRC patients compared with healthy controls, using four independent public datasets. The second step was to assess correlation between metastasis and CEP55 expression level, using 110 staged CRCs and their Immunohistochemistry-based grades. The third step was to artificially alter CEP55 expression levels in HCT116 cells, using plasmid-implanted CEP55 clones and siRNA knockdowns, and to apply high-throughput mRNA sequencing to holistically profile the resultant co-regulation information. The results show elevated CEP55 levels in CRCs over controls. And the results reveal positive correlation between CEP55 level and spreading status to lymph nodes. CEP55 promotes cell migration and delineates the involved genes, functions and pathways in CEP55 co-regulation. The results overall pave the way for deeper understandings of the underlying molecular mechanisms and suggest CEP55 as a therapeutic target for CRC metastasis.

關鍵字(中)

★ 大腸直腸癌
★ 轉移
★ CEP55
★ 高通量定序技術

關鍵字(英)

★ Colorectal cancer
★ metastasis
★ CEP55
★ high-throughput mRNA sequencing

論文目次

中文摘要 i
Abstract ii
誌謝 iii
Table of contents iii
List of tables vi
List of figures vii
List of supplements tables viii
List of supplements figures ix
1. Introduction 1
1-1. Colorectal cancer 1
1-2. CRC metastasis 2
1-3. Role of centrosomal protein CEP55 3
2. Materials and methods 5
2-1. Public data collection 5
2-2. CRC tissue preparation 5
2-3. Cell culture conditions 5
2-4. Cell transient transfections and stabilization 6
2-5. Immunohistochemical staining (IHC) 6
2-6. RNA extraction 7
2-7. Protein extraction 8
2-8. RT-PCR 9
2-9. Western blot assay 10
2-10. Wound-healing assay 10
2-11. Cell migration assay 11
2-12. RNA-seq library construction and sequencing 11
2-3-2. RNA-seq data analysis 12
3. Results 14
3-1. Transcriptome analysis 14
3-2. Clinicopathological analysis 20
3-3. Cytological analysis 23
3-3-1. Promotion of CRC migration by CEP55 23
3-3-2. RNA-seq data analysis 28
3-3-3. Differential gene expression and bioinformatics analysis 30
3-3-4. CEP55 co-expression analysis 35
4. Discussion 43
5. Reference 46
6. Supplements 48

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指導教授

蘇立仁(Li-Jen Su)

審核日期

2015-7-14

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