酵母菌YPL014W受到Mcm1調控並參與細胞週期之功能分析

Abstract

細胞週期，是指能持續分裂的真核細胞從一次有絲分裂結束後生長，再到下一次分裂結束的循環過程。細胞週期是所有生物繁殖複製所必需的機制，而出芽酵母菌則是用來研究細胞週期調控制很重要的模式生物。真核生物的細胞週期可分為G1、S、G2、M四個時期，其中G1時期的長度受到外在環境及訊息的影響非常大，細胞是否能夠通過G1要進入到S時期此一主要的調控點是由Cdk及G1 週期素(Cyclin)活性所調控。由先前的研究中顯示Ypl014w可能與Cdk1、Cln2等細胞週期調蛋白形成複合體，因此本研究的目的在於探討此蛋白質在酵母菌細胞週期中所執行的功能為何。我們的實驗結果顯示YPL014W受到一轉錄因子Mcm1的調控而有週期性的表現，並且透過免疫沉澱的方法證實其參與在Cln2複合體中。此基因的剔除對細胞週期的進行沒有顯著的影響，但是過度表現此蛋白質會造成細胞週期停滯在G1時期。本實驗證實Ypl014w的確參與在細胞週期的調控之中並且受到Mcm1的調控,而其詳細的機制及調控的方式我們將會更進一步的研究透徹。

The G1-S transition in the cell cycle is largely regulated by the activity of the G1 cyclin-Cdc28 cyclin-dependent protein kinase complex. Among G1 cyclins, Cln1 and Cln2 appear to make a specific contribution to the initiation of the cell cycle, while Cln3 functions for the timely expression of CLN1, CLN2 and other G1-specific genes. The Cln3-Cdc28 kinase activates the Swi4-Swi6 transcription complex which induces CLN1 and CLN2 transcription in late G1 and drives the transition to S phase. Previous results suggest that Ypl014w p is a component of the G1 cyclin/Cdc28 complex. However, it is still elusive how this gene is cell cycle-regulated and whether it affects the activity of the G1 cyclin-CDK complex. The preliminary data showed that YPL014W is activated by the transcription factor, Mcm1p. Chromatin immunoprecipitation experiments demonstrate a direct binding of Mcm1p at the promoter of YPL014W. In this study, we show that YPL014W is expressed in a cell cycle-regulated pattern. In addition, the interaction between Ypl014w and Cln2 was confirmed by co-immunoprecipitation. Moreover we demonstrated the Ypl014w overexpression cause the cell cycle arrest at G1 phase. These observations support the view that the Mcm1p-coordinated rise of YPL014W level is requires for the cell cycle regulation , the mechanism involved thus need to be further investigated.