EZH2藉由PIP5K1C誘導鈣離子訊息傳遞調控人類間質幹細胞的神經分化

EZH2 Regulates Neuronal Differentiation of Human Mesenchymal Stem Cells through PIP5K1C-Dependent Calcium Signaling

陳伶慈(Ling-Tzu Chen)

指導教授 ： 謝嘉玲 ； 余永倫 ； 鄒瑞煌

亞洲大學/健康學院/生物科技學系碩士班/碩士(2010年)

摘要

EZH2是PcG蛋白群的一員，其功能在調節幹細胞的自我更新與分化。在PI(4,5)P2的合成過程PIP5K1C扮演著重要的角色。磷脂酶C（phospholipase C）會將PI(4,5)P2水解成細胞內傳訊者IP3和DAG，其中IP3是細胞內鈣離子的活化者。在神經分化過程中，細胞內鈣離子濃度改變是必須且關鍵的。然而，EZH2是否調控細胞內鈣離子訊息傳遞，以及它是如何調控人類間質幹細胞（hMSCs）的神經分化，其機制目前尚未清楚。在本研究中，偵測神經標記的表現結果顯示，我們成功地誘導hMSCs分化成神經系列（lineage）的細胞。將EZH2抑制後，細胞內鈣離子的濃度很顯著上升約14倍，也使PIP5K1C的表現量上升。在hMSCs細胞中，EZH2會跟PIP5K1C的啟動子（promoter）結合，而在神經分化後EZH2會離開。抑制PIP5K1C會降低EZH2默化細胞內鈣離子釋出，也會抑制hMSCs的神經分化，這顯示EZH2能藉由PIP5K1C調控細胞內鈣離子濃度，進而影響hMSCs的神經分化。至此，我們最早提出此證據顯示，在增生的hMSCs細胞中，EZH2負調控細胞內鈣離子。在神經分化的細胞中，EZH2離開PIP5K1C的啟動子後，活化PIP5K1C的表現，並誘導細胞內鈣離子訊息傳遞，使hMSCs分化成神經系列的細胞。

關鍵字

人類間質幹細胞 ； EZH2 ； PIP5K1C ； 神經分化

並列摘要

Enhancer of zeste homolog 2 (EZH2), a polycomb group (PcG) protein, regulates stem cells renewal and differentiation. Phosphatidylinositol-4-phosphate 5-kinase, type I gamma (PIP5K1C) plays an important role in synthesis of phosphatidylinositol 4, 5-bisphosphate [PI(4,5)P2]. Phospholipase C (PLC) hydrolyzes PI(4,5)P2 into inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG), in which IP3 is an intracellular messenger to activate calcium signaling. Change in intracellular calcium concentration is required and critical in neuronal differentiation. However, whether EZH2 modulates intracellular calcium signaling and how it regulates neuronal differentiation of human mesenchymal stem cells (hMSCs) are still unclear. In the present study, we successfully induce hMSCs to differentiate into neuronal lineage by detecting the expression of neuronal markers. Knockdown of EZH2 dramatically increases the level of intracellular calcium by 14-fold, and also evokes the expression of PIP5K1C. EZH2 binds the promoter of PIP5K1C in hMSCs and then dissociates after neuronal differentiation. Knockdown of PIP5K1C significantly reduces intracellular calcium release in EZH2-silenced cells, and disrupts neuronal differentiation of hMSCs, indicating that EZH2 regulates intracellular calcium through PIP5K1C, and thus affects neuronal differentiation of hMSCs. Here, we provide the first evidence to show in proliferating hMSCs, EZH2 negatively regulates intracellular calcium. After induction of neuronal differentiation, EZH2 departs from the promoter of PIP5K1C, activates its expression, and evokes intracellular calcium signaling, leading to differentiate hMSCs into neuronal lineages.

並列關鍵字

hMSCs ； EZH2 ； PIP5K1C ； neuronal differentiation

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