PROMOTER SPECIFICITY OF TRANSCRIPTION FACTORS

描述：相关的酵母转录激活因子SWI5和ACE2 具有相关的锌指/DNA结合域，并与相同的序列结合。 然而，SWI5激活了HO(交配型转换内切酶)和ACE2 激活CTS1(几丁质酶)。这两种激活剂都在细胞周期的G1期起作用。 它们的特殊性的基础是什么？在此之前，斯蒂尔曼博士 已经有了几个发现。第一，CTS1中ACE2的结合位点 鉴定了启动子并将ACE2和SWI5与CTS1和HO结合 在体外进行比较。这导致了SWI5和ACE2结合到 具有相同亲和力的相同序列。 有趣的是，第三个基因SIC1(cdk抑制物)被鉴定出来，它是 被SWI5和ACE2激活。其次，SWI5绑定到HO UAS in 与另一种激活剂PHO2在体外举行音乐会。在体内，PHO2不是 野生型HO转录所必需的，但当SWI- HO中的结合位点因突变而减弱。PHO2协同工作 使用不同的酵母激活剂对PHO5和HIS4等基因进行激活。它会播放 不参与CTS1的表达，不与ACE2协同作用。第三，通过 通过制作SWI5-ACE2嵌合体，鉴定了这些蛋白质中的大区域 它们不同于DNA结合区，决定着靶基因 专一性。第四，CTS1启动子中的负性元件大致是 已定义移除后允许SWI5激活的选项。有人建议， ACE2的一种活性必须是中和这一元素。此元素还 将抑制启动子融合中的异源Cyc1激活剂。可能的 该元件的反式作用抑制物是通过突变鉴定的 允许SWI5激活CTS1的基因(NCE)。第五。两个PHO2中的区域 和HO合作所需的SWI5。PHO2突变 都是缺失，SWI5突变包括单一氨基酸的变化， 它们聚集在一个地区。第六，两个相距较远的SWI5的突变 HO的结合位点被用来暴露PHO2要求，导致 想象一下，SWI5-PHO2复合体介导了长程相互作用 在这两个地点之间。 在下一阶段，PI建议1.进一步描述 PHO2-SWI5相互作用。2.描述CTS1中的负调控 推动者。3.表征HO启动子中负调控的作用 专一性。4.描述ACE2如何赋予启动子特异性 激活。5.描述受 SWI5和ACE2。

DESCRIPTION: The related yeast transcriptional activators SWI5 and ACE2 have related zinc finger/DNA-binding domains, and bind to the same sequence. Yet SWI5 activates HO (endonuclease for mating type switching) and ACE2 activates CTS1 (chitinase). Both activators work in G1 of the cell cycle. What is the basis of their specificity? In the prior period, Dr. Stillman has made several findings. First, binding sites for ACE2 in the CTS1 promoter were identified and binding of ACE2 and SWI5 to CTS1 and HO sites compared in vitro. This led to the conclusion that SWI5 and ACE2 bind to the same sequences with the same affinities. Interestingly, a third gene SIC1 (cdk inhibitor) was identified which is activated by both SWI5 and ACE2. Second, SWI5 binds to the HO UAS in concert with another activator, PHO2, in vitro. In vivo, PHO2 is not required for transcription of wild type HO, but is important when the SWI- binding sites in HO have been weakened by mutation. PHO2 works in concert with different yeast activators for genes such as PHO5 and HIS4. It plays no role in expression of CTS1 and does not cooperate with ACE2. Third, by making SWI5-ACE2 chimeras, large regions in these proteins were identified which are distinct from the DNA-binding domains and dictate target gene specificity. Fourth, a negative element in the CTS1 promoter was roughly defined which when removed allows SWI5 to activate. It is suggested that one activity of ACE2 must be to counteract this element. This element also will inhibit the heterologous CYC1 activators in promoter fusions. Possible trans-acting repressors for this element were identified by mutations in genes (NCE) that allow SWI5 to activate CTS1. Fifth. regions in both PHO2 and SWI5 required for cooperation at HO were identified. The PHO2 mutations were deletions and the SWI5 mutations included single amino acid changes, which clustered in one region. Sixth, mutations in the two distant SWI5 binding sites of HO were used to expose a PHO2 requirement, leading to a picture that a SWI5-PHO2 complex mediates the long range interactions between these two sites. In the next period, the PI proposes to 1. further characterize the PHO2-SWI5 interaction. 2. Characterize the negative regulation in the CTS1 promoter. 3. Characterize the role of negative regulation in HO promoter specificity. 4. Characterize how ACE2 confers promoter specific activation. 5. Characterize the regulation of other genes regulated by SWI5 and ACE2.