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標題: Cdk5/p35蛋白對於雄性激素接受器功能調控及攝護腺癌細胞增生所扮演的角色
The roles of Cdk5/p35 in regulating androgen receptor function and proliferation of prostate cancer cells
作者: 曾群富
Tseng, Chun-Fu
關鍵字: androgen receptor;雄性激素接受器;prostate cancer;攝護腺癌
出版社: 生命科學系所
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Cyclin dependent Kinase 5 (Cdk5) 蛋白在Cdk家族中是很特別的一員,它的激酶活性不需依賴Cyclin蛋白所啟動,並且不涉及細胞週期 (cell cycle) 的調控。Cdk5的激酶活性是需要一種神經特有的活化因子,p35,所啟動。在現今的研究中,Cdk5與p35蛋白活性的功能一般被探討於中樞神經系統與神經退化性疾病。本實驗室為首先發現在攝護腺癌細胞株 (lymph node carcinoma of prostate, LNCaP) 有 Cdk5與p35蛋白的表現,且在Cdk5活性抑制劑Roscovitine抑制其活性的情況下,發現細胞型態有明顯的改變。雄性激素接受器 (androgen receptor) 是種配體依賴的核酸轉錄因子,並在調控攝護腺癌細胞增生中扮演重要的角色。在前人研究中,Cyclin D1是能與雄性激素接受器有蛋白之間的交互作用而p35是種與Cyclin結構相似的蛋白,是故我們利用免疫沉澱法及免疫細胞螢光染色法證明在LNCaP細胞中,Cdk5具有透過p35與雄性激素接受器有蛋白之間交互作用的能力。在處理人工合成之雄性激素R1881刺激下,經由Roscovitine以及Cdk5 siRNA的處理,發現AR-Ser81的磷酸化與Cdk5的蛋白及活性有關。本實驗成果為首度報導雄性激素接受器是Cdk5激酶的受質 (substrate) 。在攝護腺癌最新的研究顯示,Her2/ErbB3的蛋白活性也能經由透過下游訊息所磷酸化AR-Ser81而調節AR的轉錄功能。同時,Cdk5的蛋白活性在神經細胞中Neuregulin所誘導的PI3K/Akt訊息傳遞路徑中能直接影響Her2/ErbB3的活性。因此,我們也利用免疫沉澱法及免疫細胞螢光染色法發現在LNCaP細胞中,Cdk5與Her2/ErbB3有蛋白之間的交互作用。在Cdk5所影響的生物功能上,我們藉由MTT assay發現,在抑制Cdk5活性的情況下,細胞的增生也明顯的受到抑制,並且在處理Cdk5 siRNA的情形下,細胞內PSA表現量有受到抑制。綜合以上的結果,我們認為Cdk5與p35蛋白對於雄性激素接受器功能及攝護腺癌細胞增生中扮演重要的角色,並且為治療攝護腺癌的基礎研究提供一個新的方向。

Cyclin-dependent kinase 5 (Cdk5) is a unique member of Cdk family. Cdk5 is neither activated by Cyclin nor involved in regulating cell cycle. p35 is a neuron-specific activator of Cdk5. The functions of Cdk5 and p35 are explored in central nervous system (CNS) and neurodegenerative diseases in recent decade. Our results demonstrated that Cdk5 and p35 proteins were expressed in prostate cancer cell (lymph node carcinoma of prostate (LNCaP)) and cell morphology was changed by treatment with Cdk5 specific inhibitor, Roscovitine. Androgen receptor (AR) is a ligand-dependent transcription factor that mediates proliferation of prostate cancer. It has been reported that there is protein interaction between Cyclin D1 and AR. Since the similar structure of p35 and Cyclin, the interaction of Cdk5 and AR via p35 was identified by immunoprecipitation and immunocytochemistry. Under the treatment of R1881, we found AR p-Ser81 site was inhibited by treatment with Roscovitine as well as Cdk5 siRNA. On the other hand, up-to-date reports indicate Her2/ ErbB3 could regulate AR transcription function in prostate cancer and regulate AR p-Ser81 site. Cdk5 has also been considered to directly activate Her2/ ErbB3 and involved in neuregulin-dependent PI3K/Akt pathway in neuronal cells. Therefore, the interaction between Cdk5 and Her2/ErbB3 were then identified. Furthermore, Cdk5 inhibition by Roscovitine or siRNA decreased the proliferation and intracellular expression of prostate specific antigen (PSA) of LNCaP cells, which are two important biomarkers for prostate cancer cells. These results suggest that Cdk5 and p35 are novel players in regulating AR function and prostate cancer cell proliferation and potential to become new therapeutic targets in prostate cancer.
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