Characterization of Parp14-Trim25 directed mitochondrial ISGylation and its antagonism by viral deISGylases

Parp14-Trim25 定向线粒体 ISG 化的表征及其病毒去 ISG 酶的拮抗作用

• 批准号: BB/Y000307/1
• 负责人: Sumana Sanyal
• 金额: $ 63.34万
• 依托单位: University of Oxford
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2024
• 资助国家: 英国
• 起止时间: 2024 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FY000307%2F1
• 关键词: Characterization; Parp14; Trim25; directed; mitochondrial

Interferon stimulated gene 15 (ISG15) is a ubiquitin like post-translational modifier, and expressed in abundance upon immune challenges such as infection, and other cellular stresses such as mitochondrial disruption, ageing and DNA damage. ISG15 can modify substrates in a process referred to as ISGylation. Many viruses in turn have co-evolved the ability to hydrolyse ISG15 modifications via the reverse reaction referred to as deISGylation, allowing them to subvert host cell function and innate immune responses.Much of the innate immune signalling takes place at the mitochondrial membranes, which function as platforms or signalling hubs in collaboration with the endoplasmic reticulum membrane. These processes consist of various signalling branches with specificity for different pathogen classes. The role of ISGylation in antiviral signalling at the mitochondria and subsequent virus restriction are poorly understood processes.Our efforts to expand the knowledge on the role of ISGylation, recently led to the identification of a new antiviral branch defined by the Poly ADP-ribosyltransferase 14 (Parp14) enzyme together with an E3 ligase Trim25. We showed that this enzyme drives ISGylation of as yet unidentified substrates at the mitochondria. Many of these substrates in turn are deISGylated by a viral protease (SARS-CoV-2 PLpro). How Parp14-Trim25 functions to regulate ISGylation at the mitochondria, and its functional implications are not known. Similarly, the full repertoire of substrates of Parp14-Trim25 and how its ADP-ribosyltransferase enzymatic activity impacts ISGylation remain important issues to address. While not much is known about Parp14-Trim25, deletion of Parp14 leads to increased susceptibility to virus infection and aberrant innate immune signalling. Importantly, viruses encoding potent deISGylating activities are also known to contain ADP-ribosylhydrolase activities in macrodomains and have effective immune suppression abilities. Whether these hydrolytic activities function in concert to evade host immunity is currently unknown.This proposal combines genetic, biochemical and proteomics approaches in cultured and hIPS-derived macrophages to systematically address these open questions. Our work will provide comprehensive insight into the novel Parp14-regulated antiviral branch and shed light on its physiological role in a relevant cellular system. The findings will inform on the mechanisms of antiviral immunity and other stress conditions, and those disrupted by many RNA viruses to ultimately offer hints toward novel avenues for therapeutic intervention.

干扰素刺激基因 15 (ISG15) 是一种类似泛素的翻译后修饰剂，在感染等免疫挑战以及线粒体破坏、衰老和 DNA 损伤等其他细胞应激时大量表达。 ISG15 可以在称为 ISGylation 的过程中修饰底物。许多病毒反过来又共同进化出通过称为去ISG化的逆反应水解ISG15修饰的能力，从而使它们能够破坏宿主细胞功能和先天免疫反应。大部分先天免疫信号发生在线粒体膜上，线粒体膜发挥作用作为与内质网膜协作的平台或信号中枢。这些过程由针对不同病原体类别具有特异性的各种信号传导分支组成。 ISGylation 在线粒体抗病毒信号传导中的作用以及随后的病毒限制过程尚不清楚。我们努力扩大对 ISGylation 作用的了解，最近发现了由聚 ADP-核糖基转移酶 14 定义的新抗病毒分支（ Parp14) 酶与 E3 连接酶 Trim25 一起。我们发现这种酶驱动线粒体中尚未鉴定的底物的 ISGylation。许多这些底物又被病毒蛋白酶 (SARS-CoV-2 PLpro) 去糖基化。 Parp14-Trim25 如何发挥作用来调节线粒体的 ISGylation，及其功能意义尚不清楚。同样，Parp14-Trim25 底物的完整库及其 ADP-核糖基转移酶酶活性如何影响 ISGylation 仍然是需要解决的重要问题。虽然对 Parp14-Trim25 知之甚少，但 Parp14 的缺失会导致对病毒感染和异常先天免疫信号的易感性增加。重要的是，编码有效去ISG化活性的病毒还已知在大结构域中含有ADP-核糖基水解酶活性，并具有有效的免疫抑制能力。目前尚不清楚这些水解活性是否协同作用以逃避宿主免疫。该提案结合了培养的巨噬细胞和 hIPS 衍生的巨噬细胞的遗传、生化和蛋白质组学方法，系统地解决了这些悬而未决的问题。我们的工作将提供对新型 Parp14 调节的抗病毒分支的全面了解，并阐明其在相关细胞系统中的生理作用。这些发现将揭示抗病毒免疫和其他应激条件的机制，以及被许多 RNA 病毒破坏的机制，最终为治疗干预的新途径提供线索。