Acetylation of SUMO paralogs as a determinant of SUMO-mediated protein networks

蛋白质通过磷酸化、乙酰化或泛素化对蛋白质进行翻译后修饰(PTM)是控制细胞信号网络的核心机制。一般原理是通过特定的相互作用区域选择性地识别修饰的蛋白质。泛素相关的相扑蛋白的修饰也调节特定的蛋白质-蛋白质相互作用，从而调节各种细胞信号过程。相扑的附着通常会指定一种蛋白质，供包含相扑相互作用基序(SIM)的结合伙伴识别。突出的例子是靶向相加作用的RanGAP1到RanBP2或依赖相扑/SIM的PML核体组装(NBS)。经典SIMs的特征是一段疏水残基，这些残基与相扑表面的疏水口袋相互作用。在SIM模块的一个子集中，酸性残基位于疏水核心的两侧，并通过与修饰剂上的基本界面的静电相互作用增强与相扑的结合。尽管在相扑/SIM相互作用中有这些相对详细的分子见解，但这些相互作用的特异性和动力学是如何调节的还不清楚。我们最近揭示了PTMS对这一进程的重要性。我们最初发现，在含有SIM的蛋白质子集中，与疏水区域相邻的丝氨酸或苏氨酸残基被磷酸化。这些磷酸SIMs中磷酸化残基的额外负电荷增强了与相扑基本界面的结合。最近，我们将这一基本界面内赖氨酸残基的位置特异性乙酰化定义为控制相扑/SIM相互作用的一种新的调节原理。我们可以证明K37位的SUMO1或K33位的SUMO2的乙酰化仍然允许与RanBP2结合，但排除了与PIAS家族成员PML或Daxx的相互作用。因此，相扑影响了PML NBS的组装，特别是阻止了SIM依赖的辅抑制子Daxx被招募到这些结构中。重要的是，在生化实验中，我们可以进一步证明K33乙酰化的SUMO2是含有溴域(BRD)的蛋白p300的结合平台。在这项提案中，我们的目标是描绘乙酰化如何控制相扑介导的蛋白质网络。我们的目标是识别受乙酰化相扑修饰调控的特定底物，并试图了解这如何影响选定的细胞途径，特别是对基因表达的控制。最后，我们想要解开相扑的乙酰化是如何调节的。在资助期内，我们预计将达到以下里程碑：1.破译相扑副词上的特定乙酰码，它决定了相扑/SIM和乙酰-相扑/BRD相互作用的选择性。定义乙酰化相扑密码的靶点，并描绘其对基因表达的影响。确定控制相扑对虾乙酰化的调节成分和生理刺激。

The post-translational modification (PTM) of proteins through phosphorylation, acetylation or ubiquitylation is a central mechanism for the control of cellular signalling networks. A general principle is the selective recognition of the modified protein by a specific interaction domain. Modification by the ubiquitin-related SUMO protein also modulates specific protein-protein interactions thereby regulating various cellular signalling processes. Attachment of SUMO typically earmarks a protein for the recognition by a binding partner that contains a SUMO interaction motif (SIM). Prominent examples are the targeting of sumoylated RanGAP1 to RanBP2 or the SUMO/SIM-dependent assembly of PML nuclear bodies (NBs). Canonical SIMs are characterized by a stretch of hydrophobic residues, which interacts with a hydrophobic pocket on the surface of SUMO. In a subset of SIM modules acidic residues flank the hydrophobic core and enhance the binding to SUMO via electrostatic interactions with a basic interface on the modifier. Despite these relatively detailed molecular insights in SUMO/SIM interactions it is unclear how specificity and dynamics of these interactions are regulated. We recently unravelled the importance of PTMs for this process. We initially found that in a subset of SIM-containing proteins serine or threonine residues that juxtapose the hydrophobic region are phosphorylated. The additional negative charges of phosphorylated residues in these phosphoSIMs enhance the binding to the basic interface of SUMO. Very recently we defined site-specific acetylation of lysine residues within this basic interface as a novel regulatory principle in the control of SUMO/SIM interactions. We could show that acetylation of SUMO1 at K37 or SUMO2 at K33 still allows binding to RanBP2, but precludes the interaction with PIAS family members, PML or Daxx. Accordingly, SUMO affected the assembly of PML NBs and in particular prevented the SIM-dependent recruitment of the co-repressor Daxx to these structures. Importantly, in biochemical experiments we could further demonstrate that K33 acetylated SUMO2 serves as a binding platform for the Bromodomain (BRD) containing protein p300. In this proposal we aim to delineate how acetylation controls SUMO-mediated protein networks. We aim to identify specific substrates that are regulated by acetyl-SUMO modification and seek to get insight how this affects selected cellular pathways, in particular the control of gene expression. Finally, we want to unravel how the acetylation of SUMO is regulated.Within the funding period we anticipate to reach the following milestones:1. Decipher a specific acetyl-code on SUMO paralogs that determines the selectivity of SUMO/SIM and acetyl-SUMO/BRD interactions.2. Define targets of the acetyl-SUMO-code and delineate its impact on gene expression.3. Identify the regulatory components and physiological stimuli that control acetylation of SUMO paralogs.