Effects of acetylation on the function of a transcriptional co-activator

转录共激活因子p300具有内在的组蛋白乙酰转移酶（HAT）活性，是一系列细胞过程所必需的。严格控制p300对于确保精确的组蛋白乙酰化和基因激活至关重要。我们已经确定p300受翻译后修饰和细胞分布的动态调节。此外，我们是第一个证明p300受细胞质蛋白酶体系统的调节。我们的研究为细胞运输和空间再分布如何控制p300的蛋白质稳定性和转录活性提供了新的见解。有趣的是，p300在体内是一个真正的乙酰化底物，我们已经将乙酰化位点定位到它的c端区域。我们还发现E1A病毒蛋白，p300的抑制因子，增强p300的自乙酰化，同时抑制p300 HAT活性。基于这些观察，我们假设可逆乙酰化在p300活性的控制中起关键作用，进而影响p300在重要细胞过程中的功能。我们将首先研究乙酰化对p300功能和转换的影响。我们将识别和表征p300中特定的乙酰化位点。我们还将生成模拟乙酰化位点的突变体，并产生针对特定乙酰化赖氨酸的抗体，以研究乙酰化对p300蛋白稳定性和转录活性的影响。我们的目标是确定乙酰化介导的p300调控的分子基础及其对p300在转录激活方面作为HAT、支架或染色质桥的功能的影响。我们的长期目标是确定这些调节机制如何影响p300依赖性基因的表达，从细胞内运输，蛋白质周转到转录激活，并确定p300在网络生物学方式中的调节的分子基础。

The transcriptional co-activator p300 contains an intrinsic histone acetyltransferase (HAT) activity and is required for an array of cellular processes. Tight control of p300 is critical to ensure precise histone acetylation and gene activation. We have established that p300 is dynamically regulated by post-translational modifications and cellular distributions. In addition, we are the first to show that p300 is regulated by the cytoplasmic proteasome system. Our studies have provided novel insight on how cellular trafficking and spatial redistributions control the protein stability and transcriptional activity of p300. Interestingly, p300 is a bona fide acetylation substrate in vivo, and we have mapped the acetylation sites to its C-terminal region. We also found that E1A viral protein, a repressor of p300, enhances p300 autoacetylation while repressing p300 HAT activity. Based on these observations, we hypothesize that reversible acetylation plays critical roles in the control of p300 activity, which in turn affects p300 function in important cellular processes. We will first study the effects of acetylation on p300 function and turnover. We will identify and characterize the specific acetylation sites in p300. We will also generate mutants that mimic the acetylation sites and produce antibodies against the specific acetylated-lysine to study the impact of acetylation on protein stability and transcriptional activity of p300. Our goal is to determine the molecular basis for acetylation-mediated p300 regulation and their consequences on the function of p300 as a HAT, a scaffold, or a bridge on chromatin with respect to transcriptional activation. Our long term goals are to determine how these regulatory mechanisms affect p300-dependent gene expression from intracellular trafficking, protein turnover to transcriptional activation, and to determine the molecular basis for p300 regulation in a network biology fashion.