探討酵母菌Ndt80蛋白對減數分裂期DNA複製的可能調控功能

Abstract

在減數分裂中，一次DNA複製之後連續進行二次細胞核分裂。然而，在二次分裂期之間如何控制DNA不再進行複製目前並不是很清楚。在出芽酵母菌中，Ndt80是一個在減數分裂特定表現蛋白，它是啟動染色體分離以及孢子形成的一個必要轉錄因子。我們發現，若在營養生長細胞中異位表現Ndt80，則細胞出芽會受到抑制而且無法進行DNA複製，這顯示在減數分裂中Ndt80也可能具有抑制DNA複製的能力。我們認為Ndt80可能參與減數分裂二次分裂期間不再發生一次DNA複製的調控。 為了證明Ndt80是否在減數分裂中具有抑制DNA複製的能力，我們使細胞在減數分裂DNA複製期(pre-meiotic DNA replication)之前提早表現NDT80。在起始減數分裂時利用IME1啟動子將NDT80提早表現，細胞仍然可以進行DNA複製，但大多數的細胞會停滯在第一次減數分裂期。若將NDT80提早表現在減數分裂前的培養(pre-meiotic incubation)時，此時大部分細胞處於後複製複合體期(post-RC state)，則部分細胞的DNA複製受到抑制。然而，若將NDT80表現在G1時期的細胞，則看不到DNA複製被抑制的現象。由這些結果可以得知，Ndt80在減數分裂中也許能透過防止前複製複合體(pre-RC)形成而抑制DNA複製，但是當pre-RC存在時卻無法抑制DNA複製。

Meiosis is characterized by a single round of DNA replication followed by two rounds of chromosome segregation. However, the mechanism to prevent DNA replication between the two rounds of division is not clear. In the budding yeast, the meiosis-specific transcription factor Ndt80 is pivotal to nuclear divisions and spore formation. Interestingly, ectopic expression of NDT80 in vegetative cells blocked budding and DNA replication. It is possible that Ndt80 also has the ability to inhibit DNA replication during meiosis. Therefore, we propose that Ndt80 might be involved in repressing another round of DNA replication between meiosis I and meiosis II. To test whether Ndt80 has the ability to inhibit replication in meiosis, we tried to precociously express NDT80 before the pre-meiotic DNA replication. We found cells that precociously expressed of NDT80 through the IME1 promoter at the initiation of meiosis could execute pre-meiotic DNA synthesis but were arrested at meiosis I. When NDT80 was expressed during the pre-meiotic incubation, in which most cells were at the post-RC stage, pre-meiotic DNA replication was partially repressed. However, the repression effect was not observed when NDT80 was expressed at the time that most cells were at G1 phase. These results suggest that Ndt80 might repress DNA replication by preventing pre-RC formation, but it could not repress DNA replication at the existing pre-RCs.