Defining the Biochemical Function and Therapeutic Utility of Unique PARP14 and PARP15 ADP-Ribosylation Sites

定义独特的 PARP14 和 PARP15 ADP-核糖基化位点的生化功能和治疗效用

• 批准号: 10046261
• 负责人: Ian O'Brien Carter-O'Connell
• 金额: $ 38.85万
• 依托单位: SANTA CLARA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10046261
• 关键词: ADP ribosylation; Active Sites; Adenosine Diphosphate Ribose; Amino Acids; Antiviral Agents; Behavior; Biochemical; Biological Assay; Biology; Cell physiology; Cells; Cellular biology; Chemicals; DNA Repair; Data; Development; Developmental Biology; Disease; Enzymes; Event; Family; Family member; Human; Immunity; Immunologic Factors; In Vitro; Knowledge; Label; Libraries; Life; Link; Malignant Neoplasms; Medicine; Modeling; Modification; Monitor; N-substituted Glycines; Pathway interactions; Peptides; Peptoids; Phosphotransferases; Physiological; Play; Poly(ADP-ribose) Polymerases; Post-Translational Modification Site; Post-Translational Protein Processing; Proteins; Proteome; RNA; Regulation; Role; Series; Signal Transduction; Site; Structure-Activity Relationship; Techniques; Therapeutic; Therapeutic Intervention; Variant; Viral; Virus Diseases; Virus Replication; Work; base; cancer therapy; design; inhibitor/antagonist; interdisciplinary approach; mimetics; novel; novel therapeutics; paralogous gene; protein aminoacid sequence; side effect; therapeutic target; tumor progression

SUMMARY ADP-ribosylation is a widespread and ubiquitous post-translational modification across all kingdoms of life. Although it was the first PTM described, the biochemical selectivity and cellular consequences of ADP- ribosylation is very poorly understood. The objective of this project is to define the pathways linking unique ADP- ribosylation modifications to their downstream function in the cell. We will use this information to develop first-in- class peptide-based inhibitors to target malfunctioning ADP-ribose signaling networks. In humans, 17 different poly-ADP-ribose polymerases (PARPs) catalyze ADP-ribosylation. PARP enzymes have been implicated in a number of physiological functions (e.g. DNA repair, RNA regulation, cell development) and a broad array of diseases, most notably viral infection. While previous efforts have validated the roles for PARPs 13, 14, and 15 in viral regulation, their mechanism of action remains unclear. In large part, this lack of clarity arises from our inability to connect a single site of ADP-ribosylation to its downstream function. Work in the kinase field has demonstrated the power of connecting enzyme-specific post-translational modification sites (PTMs) to their specific cellular function, fundamentally altering our understanding of signal transduction and revealing new avenues for therapeutic intervention in disease. By leveraging the expertise from my lab and that of my collaborators, our work will use a multidisciplinary approach to determine the functional consequences of unique ADP-ribosylation events in the cell and develop novel PARP-selective peptide-based inhibitors to block specific modifications. In the first aim, we will define the sites of modification for both PARP14 and PARP15 on themselves (auto-modification) and on other targets (trans-modification). We have worked extensively with these PARP family members to define their specific protein targets and we have a robust viral model to interrogate how specific site alterations change pathway behavior. Our work in this aim will provide the first links between specific ADP-ribosylation sites and their effects in the cell and it will be adaptable to the other PARPs. In the second aim, a PARP14-specific target peptide sequence that we have developed will serve as a platform to identify selective PARP14 inhibitors. We will synthesize peptide and peptoid (poly-N-substituted glycine) derivatives of the initial target peptide and identify structure activity relationships (SAR analysis) with the resulting library. This work will expand our knowledge of the biochemical mechanisms governing target selection and will provide a novel avenue for targeting PARPs to treat disease. The work from this proposal will be broadly applicable to the study of the remaining PARP family members and will have applications in medicine, cell biology, developmental biology, and the study of the post-translationally modified proteome.

概括 ADP-核糖基化是所有生命领域广泛且普遍存在的翻译后修饰。 尽管这是第一个描述的 PTM，但 ADP- 的生化选择性和细胞后果 人们对核糖基化知之甚少。该项目的目标是定义连接独特的 ADP-的途径 核糖基化修饰对其在细胞中的下游功能。我们将利用这些信息来开发先进的 类基于肽的抑制剂，针对功能障碍的 ADP-核糖信号网络。在人类中，有 17 种不同的 聚 ADP 核糖聚合酶 (PARP) 催化 ADP 核糖基化。 PARP 酶与 许多生理功能（例如 DNA 修复、RNA 调节、细胞发育）和广泛的 疾病，尤其是病毒感染。虽然之前的工作已经验证了 PARP 13、14 和 15 的作用 在病毒调控方面，它们的作用机制仍不清楚。在很大程度上，这种缺乏清晰度是由于我们 无法将单个 ADP-核糖基化位点与其下游功能连接起来。激酶领域的工作有 证明了将酶特异性翻译后修饰位点 (PTM) 连接到其 特定的细胞功能，从根本上改变我们对信号转导的理解并揭示新的 疾病治疗干预的途径。通过利用我的实验室和我的同事的专业知识 合作者，我们的工作将使用多学科方法来确定功能后果 细胞中独特的 ADP-核糖基化事件，并开发新型 PARP 选择性肽抑制剂 阻止特定修改。第一个目标是定义 PARP14 和 PARP14 的修饰位点。 PARP15 对自身（自动修改）和其他目标（反式修改）。我们已经工作过 与这些 PARP 家族成员广泛合作，以确定他们的特定蛋白质靶标，我们拥有强大的病毒 模型来询问特定位点的改变如何改变通路行为。我们为实现这一目标所做的工作将提供 特定 ADP-核糖基化位点及其在细胞中的作用之间的第一个联系，并且它将适应其他 PARP。在第二个目标中，我们开发的 PARP14 特异性靶肽序列将作为 鉴定选择性 PARP14 抑制剂的平台。我们将合成肽和类肽（聚-N-取代 甘氨酸）初始目标肽的衍生物，并鉴定结构活性关系（SAR 分析） 生成的库。这项工作将扩展我们对控制目标选择的生化机制的了解 并将为靶向 PARP 来治疗疾病提供新途径。该提案的工作将广泛 适用于其他 PARP 家族成员的研究，并将在医学、细胞等领域得到应用 生物学、发育生物学和翻译后修饰蛋白质组的研究。