Transcriptional activation domain of p300/CBP-associated factor(P/CAF)

在真核生物中，编码第二类基因的转录涉及一般因子和RNA聚合酶II全酶在启动子上有序地聚集到基础转录预起始复合体中。转录激活剂与核心启动子外的特定DNA序列结合在一起，有效地促进转录。序列特异性激活剂与其他不直接结合DNA的因子一起工作。这些因子被称为共激活子，通过它们与DNA结合的转录因子的相互作用被招募到启动子。P/CAF最初被鉴定为p300/CBP结合蛋白，因为它与酵母组蛋白乙酰转移酶(HAT)的序列相似，即GCN5。此外，P/CAF与其他共激活因子如ACIR和SRC-1也是HAT相互作用。在本研究中，我重点研究了P/CAF在转录调控中的作用，并研究了P/CAF的转录激活结构域。为了研究P/CAF的激活功能，我在酵母中以GALA DNA结合域的形式表达了P/CAF的不同区域。P/CAF确实在酵母中作为转录激活蛋白发挥作用，氨基酸366-654区域是激活功能所必需的。由于该区域与HAT结构域相同，P/CaF的激活功能可能与组蛋白的乙酰化有关，而P/CaF与CBP的相互作用不依赖于激活结构域。用表面等离子体共振分析法研究了共激活剂在体外形成的复合体。

In eukaryores, transcription of mRNA-encoding classII genes involves the ordered recruitment of general factors and the RNA polymerase II holoenzyme into the basal transcription pre-initiation complex on promoter.The transcription is efficiently enhanced by transcriptional activators which bind to specific DNA sequences outside of the core promoter.Sequence-specific activators work in conjunction with other factors which do not bind DNA directly.These factors are termed co-activators and are recruited to promoters via their interaction with the DNA-bound transcription factors.P/CAF was originally identified as a p300/CBP-binding protein by virtue of its sequence similarity to a yeast histone acetyltransferase (HAT), namely yGCN5.In addition, P/CAF interacts with other co-activators such as ACIR and SRC-1 which are also HAT.In this study, I focused on the function of P/CAF in the transcriptional regulation and investigated the transcriptional activation domain of P/CAF.In order to examine the activation function of P/CAF, I expressed various regions of P/CAF as GALA DNA binding domain in yeast.P/CAF indeed worked as a transcriptional activator protein in yeast and the region of amino acids 366-654 was necessary for activation function.As this region is identical to the HAT domain, the activation function of P/CAF might be related to the acetylation of histones.On the other hand, the interaction of P/CAF with CBP was independent on the activation domain. The complex formation of co-activators in vitro was also examined using surface plasmon resonance anaysis.