INVESTIGATING THE ROLE OF RSC4 ACETYLATION

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 对于每个真核生物来说，一个复杂的问题是如何调节染色质重塑的过程，这是介导整个基因组DNA可及性的关键功能。使用UCSF质谱设备，我们发现RSC重塑复合物的一个重要亚单位Rsc 4在其N-末端的K25上被Gcn 5乙酰化。 在标准生长条件下纯化RSC表明大部分Rsc 4是乙酰化的。 野生型和突变型（K25 R）RSC复合物的体外比较揭示了类似的能力，重塑核小体，结合DNA，并识别乙酰化。 由于乙酰化似乎不会改变RSC的酶性质，我们正在采取几种体内方法来了解乙酰化是重要的生物过程，并了解乙酰化的功能后果。 具体来说，我们将使用质谱法来确定乙酰化被调节的条件，并结合基因突变，确定负责这种调节的基因。 这些研究是第一个表征乙酰化的染色质重塑复合物的背景下的功能。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. A complex problem for every eukaryotic organism is how to regulate the process of chromatin remodeling, a critical function to mediate DNA accessibility throughout the genome. Using the UCSF Mass Spec Facility, we have found that Rsc4, an essential sub unit of the RSC remodeling complex, is acetylated, by Gcn5, on K25 at its N-terminus. Purification of RSC under standard growth conditions suggests that most of Rsc4 is acetylated. In vitro comparisons of WT and mutant (K25R) RSC complexes reveal similar capabilities to remodel nucleosomes, bind DNA, and recognize acetylation. Since acetylation does not appear to alter the enzymatic properties of RSC, we are taking several in vivo approaches to understand the biological processes in which acetylation is important and to understand the functional consequences of the acetylation. Specifically, we will use mass spectrometry to identify conditions where acetylation is regulated and, combined with genetic mutations, identify the genes responsible for this regulation. These studies are the first to characterize the function of acetylation in the context of a chromatin remodeling complex.