DC 欄位 | 值 | 語言 |
dc.contributor | 陳瑞華 | zh-TW |
dc.contributor | Chen, Ruey-Hwa | en |
dc.contributor | 臺灣大學:生化科學研究所 | zh-TW |
dc.contributor.author | 賴俊凱 | zh-TW |
dc.contributor.author | Lai, Chun-Kai | en |
dc.creator | 賴俊凱 | zh-TW |
dc.creator | Lai, Chun-Kai | en |
dc.date | 2009 | en |
dc.date.accessioned | 2010-05-04T08:04:09Z | - |
dc.date.accessioned | 2018-07-06T06:11:26Z | - |
dc.date.available | 2010-05-04T08:04:09Z | - |
dc.date.available | 2018-07-06T06:11:26Z | - |
dc.date.issued | 2009 | - |
dc.identifier.other | U0001-2807200916010600 | en |
dc.identifier.uri | http://ntur.lib.ntu.edu.tw//handle/246246/178895 | - |
dc.description.abstract | DIP-2為BTB-kelch蛋白家族的一員。許多BTB-kelch蛋白已被發現高度表現於神經系統中,並參與包括神經型態生成以及突觸訊息傳遞在內的神經發育機轉。但就DIP-2而言,儘管我們已發現它可作為Cul3泛素接合酶與受質間的轉接者,進而導致死亡相關蛋白激酶DAPK以及抑癌蛋白PML的泛素化和降解,但對於DIP-2的表現分佈以及其詳細的功能機制所知甚少。本篇論文中,我們觀察DIP-2於成鼠腦部的表現情形,發現DIP-2主要分佈於大腦皮層、海馬迴、嗅球及小腦皮層等區域。值得注意的是,DIP-2於絕大多數區域呈現以細胞體和樹突為主的表現型態,但於海馬迴CA3處則特別表現於軸突中。另一方面,為求更進一步瞭解DIP-2的功能,本篇論文亦深入探討DIP-2對PML進行蛋白降解的調控機制,從而發現細胞週期素依賴性激酶(CDKs)參與DIP-2對PML的調控。我們證實PML確實為CDK1/2/4/6的受質,而Ser 518則為主要發生磷酸化的殘基。實驗結果顯示,這些CDK都參與由DIP-2所誘發的PML蛋白降解。總的來說,這項研究不僅呈現DIP-2蛋白於鼠腦中的表現型態特徵,亦將DIP-2所媒介的PML蛋白降解與細胞週期對PML的調控機制作緊密結合,為抑癌蛋白PML穩定性的研究提供新的一層認知。 | zh-TW |
dc.description.abstract | DIP-2 is a member of the BTB-kelch protein family, of which several members are highly expressed in the nervous system and have been demonstrated to elicit neuronal functions, including neural morphogenesis and synapse transmission. However, little is known about the expression patterns of DIP-2 and its biological functions, although previous studies in our laboratory revealed that DIP-2 functions as a substrate adaptor of Cul3-based ubiquitin ligase to promote the ubiquitination of DAPK and PML. Here, we characterized the expression pattern of DIP-2 protein in the adult mouse brain and found its high expression levels in the cerebral cortex, the hippocampus, the olfactory bulb and the cerebellar cortex. Notably, DIP-2 displays both somatic and dendritic patterns in most regions observed, whereas it exhibits an axonal pattern in CA3 of hippocampus. In an attempt to study the function of DIP-2, we investigated the mechanism through which DIP-2 mediates PML protein degradation. First, we identified a role of CDK in DIP-2-mediated PML degradation. Furthermore, we demonstrated PML as the substrate of CDK1/2/4/6 in vitro and the Ser 518 residue as the major phosphorylation site. Finally, we showed that the each of these CDKs contributes to PML degradation induced by DIP-2. Together, this study not only presents the expression features of DIP-2 in mouse brain, but also provides a linkage between DIP-2-mediated PML degradation and cell cycle-dependent regulation of PML, thus adding another aspect of proper control for the stability of tumor suppressor PML protein. | en |
dc.description.tableofcontents | Table of Content 2文摘要 4bstract 5ntroduction 6he kelch-repeat superfamily 6he BTB-kelch proteins 7he BTB-kelch proteins in the nervous system 10IP-2 (DAPK-interacting protein 2) 13ML (Promyelocytic Leukemia) 14ML nuclear bodies 15unctions of PML and PML-NBs in transcriptional regulation 16unctions of PML and PML-NBs in apoptosis 17ML as a tumor suppressor 18he control of PML protein stability 19aterials and Methods 21NA purification and RT-PCR 21lasmids 21stablishment of DIP-2 knockdown cell by lentivirus system 22ell culture and transient transfection 22ntibodies 23estern blot 23mmunohistochemistry 24mmunofluorescence 24inase inhibitors 25n vitro kinase assay 25esults 26he expression pattern of DIP-2 in adult mouse brain 26IP-2 is localized mainly in cell bodies and dendrites but also in axons of certain neurons 28DK inhibitor blocks DIP-2-mediated PML degradation 29DK1 phosphorylates PML at serine 518 and serine 527 in vitro 30DK1, 2, 4 and 6 are all involved in DIP-2-mediated PML degradation. 30yclins trigger DIP-2-mediated PML degradation 31iscussion 32eferences 36igures 49igure 1. Schematic illustration of DIP-2 domain structure and its function as an E3 ubiquitin ligase adaptor. 49igure 2. Schematic illustration of PML domain structure and its known residues for post-translational modifications. 50igure 3. Anti-DIP-2 antiserum specifically recognizes DIP-2. 51igure 4. Immunohistochemistry analysis for characterization of DIP-2 distribution in adult mouse brain. 52igure 5. DIP-2 is localized mainly in cell bodies and dendrites but also in axons of certain neurons. 54igure 6. CDK inhibitor blocks DIP-2-mediated degradation of ectopically expressed PML. 56igure 7. DIP-2-mediated degradation of endogenous PML is blocked by CDK inhibitor. 57igure 8. CDK1 phosphorylates PML principally at serine 518 in vitro. 58igure 9. CDK1, 2, 4 and 6 are all involved in DIP-2-mediated PML degradation. 59igure 10. Cyclins trigger DIP-2-mediated PML degradation. 60ppendix 61able S1. Primers used in this thesis. 61igure S1. Expression pattern of DIP-2 mRNA in embryonic mice and adult mouse brain. 62 | en |
dc.format | application/pdf | en |
dc.format.extent | 3359049 bytes | - |
dc.format.mimetype | application/pdf | - |
dc.language | en | en |
dc.language.iso | en_US | - |
dc.subject | 降解 | zh-TW |
dc.subject | 泛素化 | zh-TW |
dc.subject | 腦 | zh-TW |
dc.subject | 細胞週期素依賴性激酶DIP-2 | zh-TW |
dc.subject | Pin1 | zh-TW |
dc.subject | brain | en |
dc.subject | BTB-kelch | en |
dc.subject | CDK | en |
dc.subject | DIP-2 | en |
dc.subject | PML | en |
dc.title | BTB-kelch蛋白DIP-2的表現及功能特性分析 | zh-TW |
dc.title | Characterization of the Expression and Function of a Novel BTB-kelch Protein DIP-2 | en |
dc.identifier.uri.fulltext | http://ntur.lib.ntu.edu.tw/bitstream/246246/178895/1/ntu-98-R95b43027-1.pdf | - |
item.fulltext | with fulltext | - |
item.languageiso639-1 | en_US | - |
item.grantfulltext | open | - |
顯示於: | 生化科學研究所
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