- 學位論文
探討乳癌代謝壓力下粒線體融合蛋白MFN2表現量之調控機制
Investigating the regulatory mechanisms of mitofusin 2 (MFN2) expression under metabolic stress in breast cancer
摘要
在實質固態瘤 (solid tumor) 中心區域之癌細胞會因缺乏氧氣與養分而面臨代謝壓力，粒線體的動態平衡對於癌細胞在此等不利環境下之生長具有重要影響。 KRAB-associated protein-1 (KAP1)是個多方參與細胞生理功能調控的共轉錄調節因子。本實驗室先前研究發現在乳癌細胞缺乏養分時，活性氧物質(reactive oxygen species, ROS) 與p38 MAPK會促進KAP1 S473位點磷酸化，進而抑制粒線體融合蛋白 mitofusin 2 (MFN2) 之表現量，避免在缺乏養分時粒線體過度融合、引發不利細胞生存的氧化壓力 (oxidative stress)。然而，KAP1 S473位點磷酸化調控MFN2之相關機制尚未被探討。本論文分別就轉錄調控、轉錄後調控與蛋白質降解等不同機制進行探討。 根據實驗室先前研究，首先挑選出了會被KAP1 S473位點磷酸化調控的微小核醣核酸(microRNA, miRNA) miR-20b-5p作為探討目標。miR-20b-5p可結合至MFN2 3’UTR且可抑制MFN2，但其抑制效果有限，因此初步認為miR-20b-5p可能非主要調控MFN2表現量之因子。另外，轉錄因子EGR1之表現量雖然在缺乏養分時明顯增加，且其被誘發之機制與KAP1 S473位點磷酸化修飾機制相同，但是抑制EGR1後並未影響MFN2表現量，因此EGR1參與調控MFN2之可能性也較低。最終，在抑制蛋白酶體活性後發現MFN2蛋白量明顯增加，顯示在缺乏養分時MFN2下降是與蛋白質降解相關，未來須更進一步探討KAP1 S473位點磷酸化與MFN2蛋白質降解之間的關係。
並列摘要
Cancer cells in the core region of a solid tumor encounter metabolic stress induced by hypoxia and insufficient nutrition. Mitochondrial dynamics has a profound effect on the survival of cancer cells under this unfavorable condition. KRAB-associated protein-1 (KAP1) is a transcriptional co-regulator participating in multiple cellular physiological processes. Previously, our study has revealed that KAP1 is phosphorylated at serine 473 (S473) under nutrient deprivation by reactive oxygen species (ROS)-p38 MAPK signaling in breast cancer cells. pS473KAP1 reduces mitochondrial fusion protein mitofusin 2 (MFN2) expression and thus prevents mitochondrial hyperfusion. Mitochondrial hyperfusion leads to ROS overproduction causing detrimental effects to cancer cells under metabolic stress. Therefore, it has been demonstrated that phosphorylation at KAP1 S473 site benefits breast cancer cell survival under nutrient insufficiency-induced metabolic stress. However, pS473KAP1 mediated regulation of MFN2 remains unclear. In this thesis, the regulatory mechanisms of MFN2 including transcriptional regulation, post-transcriptional regulation and protein degradation will be discussed. Based on previous study, microRNA (miRNA) miR-20b-5p, which can be upregulated by pS473KAP1, was chosen. Although miR-20b-5p bound to MFN2 3’UTR to repress MFN2 expression, miR-20b-5p had only limited effect on MFN2 reduction. Current results suggested that miR-20b-5p might not be the main regulator of MFN2. Besides, even though the stress response transcription factor EGR1 seemed to share the same regulatory route of KAP1 S473 phosphorylation under nutrient deprivation, knockdown of EGR1 did not affect MFN2 expression. Hence, EGR1 was not likely to involve in the MFN2 regulation. At last, inhibition of proteasomal degradation accumulated MFN2 protein under nutrient deprivation, which suggested the downregulation of MFN2 under nutrient deprivation was associated with protein degradation. In the future, the detailed regulatory mechanism of pS473-KAP1 and MFN2 degradation will be further investigated.
並列關鍵字
breast cancer ； metabolic stress ； nutrient deprivation ； mitochondrial dynamics ； KAP1 ； MFN2 ； miR-20b-5p ； EGR1 ； proteasomal degradation參考文獻
延伸閱讀
- 楊爵豪（2014）。乳癌細胞株MDA-MB-361中不可逆的第一型MHC分子和抗原呈現機制缺陷的機轉研究〔碩士論文，國立清華大學〕。華藝線上圖書館。https://www.airitilibrary.com/Article/Detail?DocID=U0016-2912201413500379
- Lin, M. S. (2007). 探討乳癌激酶對其受質p190之功能調控 [master's thesis, National Taiwan University]. Airiti Library. https://doi.org/10.6342/NTU.2007.02750
- Chen, H. W. (2007). 乳癌之基因微陣列分析研究—探討基因表現與單核苷酸多型性及微型核醣核酸之關係 [master's thesis, National Taiwan University]. Airiti Library. https://doi.org/10.6342/NTU.2007.02255
- 黃姿蓉（2013）。Exploring the Molecular Mechanisms of Anti-Proliferative Effect for TF-7 on Human Breast Cancer Cells〔碩士論文，高雄醫學大學〕。華藝線上圖書館。https://www.airitilibrary.com/Article/Detail?DocID=U0011-1907201312012900
- Pai, H. C. (2015). Investigation of the anti-cancer mechanisms of Moscatilin and MT-4 in human breast and ovarian cancer [doctoral dissertation, National Taiwan University]. Airiti Library. https://doi.org/10.6342/NTU.2015.00654