PADI4-Cit - Precision bifunctional peptide tools to understand protein citrullination

蛋白质瓜氨酸化是一种翻译后修饰(PTM)，涉及多种生物学功能，包括表观基因调控和先天免疫反应。其异常调节参与了多种致病机制的进展。尽管有这些重要的作用，我们对瓜氨酸化的理解仍处于初级阶段。哺乳动物的瓜氨酸化反应是由多肽精氨酸脱亚胺酶(PADI)家族的五种杂合同工酶催化的，这些同工酶表现出重叠的底物特异性和一系列的组织表达模式。这种复杂的情况使得对瓜氨酸化的影响的精确理解具有挑战性，目前还没有在单一底物水平上探索瓜氨酸化的一般策略。为了剖析这个复杂的系统，我的目标是开发双功能化学工具，将PADI酶招募到特定的底物和基因上，从而能够以目前无法达到的精度和特异性水平研究瓜氨酸化对蛋白质功能和基因表达的影响。我将重点介绍PADI4，PADI4是PADIS的主要核成员，产生的方法可以在底物水平上精确地理解单个瓜氨酸化事件在基于细胞的背景下的生化影响。我的新的双功能工具将被用来解决两个相关的目标：了解瓜氨酸对单个蛋白质底物的影响，以及2)基因调控。这项工作的结果应该有助于阐明这种相对较少了解的PTM，为瓜氨酸化对蛋白质功能的影响提供新的见解，并为未来应用于研究其他底物和PADI提供一个蓝图，有可能为治疗干预提供新的途径。

Protein citrullination is a posttranslational modification (PTM) involved in a wide range of biological functions including epigeneticgene regulation and the innate immune response. Its dysregulation is implicated in the progress of diverse pathogeneses. Despitethese important roles, our understanding of citrullination remains in its infancy. Citrullination in mammals is catalysed by fivepromiscuous isozymes from the peptidylarginine deiminase (PADI) family, which display overlapping substrate specificities and arange of tissue expression patterns. This complex landscape makes precise understanding of the effects of citrullination challenging,with no general strategies currently available to explore citrullination at the single substrate level. To dissect this complicated system,I aim to develop bifunctional chemical tools to recruit PADI enzymes to specific substrates and genes, allowing investigation of theeffects of citrullination on protein function and gene expression with a level of precision and specificity not currently possible. I willfocus on PADI4, the primarily nuclear member of the PADIs, producing methods to precisely understand the biochemical effects ofsingle citrullination events in a cell-based context at the substrate level. My new bifunctional tools will be used to address two relatedgoals: understanding the effect of citrullination on 1) individual protein substrates, and 2) gene regulation. Results from this workshould shed light on this relatively poorly understood PTM, providing new insight into the effects of citrullination on protein functionand a blueprint for future applications to study other substrates and PADIs, with the potential to provide new avenues for therapeuticintervention.