Regulation of the DNA-degrading enzyme TREX1 by intramembrane proteolysis and its implications for human auto-immune diseases as well as a model for ER proteostasis

膜内蛋白水解对 DNA 降解酶 TREX1 的调节及其对人类自身免疫性疾病的影响以及 ER 蛋白稳态模型

• 批准号: 518795826
• 负责人: Professor Dr. Bernd Schröder
• 金额: --
• 依托单位: Institut für Physiologische Chemie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/518795826?language=en
• 关键词: Regulation; DNA; degrading; enzyme; TREX1

Mutations in the Three prime repair exonuclease 1 (TREX1) gene lead to several auto-inflammatory diseases including Aicardi-Goutières syndrome (AGS). TREX1 is a DNA-degrading enzyme, which can clear DNA from the cytosol. From a cell’s perspective, DNA in the cytosol can indicate intrusion of a pathogen. Therefore, DNA-sensing mechanisms like the cGAS-STING pathway have evolved triggering defence mechanisms like production of interferons. However, overactivation of these pathways by accumulating DNA, e.g. in the absence of TREX1, leads to immunopathology.TREX1 is synthesised as a tail-anchored protein which is attached to the ER membrane by a C-terminal transmembrane domain (TMD). This is in contradiction to several reports detecting TREX1 also in the nucleus. We have identified TREX1 as novel substrate of the ER-localised intramembrane protease Signal peptide peptidase (SPP). Cleavage by SPP releases a soluble form of TREX1 into the cytosol. We found that TREX1 can exist constitutively in the cleaved soluble as well as a membrane-bound form. However, TREX1 cleavage can also result in its rapid further degradation so that SPP also regulates total cellular TREX1 levels. These findings suggest a major regulatory impact of SPP on TREX1. As SPP mediates the transition between both forms, the analysis of SPP-deficient cells and mice provides a system where generation of soluble TREX1 is prevented. We plan to analyse where and under which conditions TREX1 cleavage takes place, how this influences the stability of the cleavage product and how this is connected to nuclear translocation. A central question is how the release of TREX1 from the membrane impacts on its capability to degrade cytosolic DNA resulting from different sources, including retrotransposons. Thinking this further, we will also study how absence or presence of cytosolic TREX1 modulates the initiation of interferon responses by exogenously introduced, but also pathogen-derived DNA or DNA released following irradiation like upon radiotherapy. This will also be analysed in a novel mouse model solely expressing soluble TREX1. Strong support that TREX1 proteolysis is a biologically relevant process is provided by an AGS-causing mutation in the TREX1 TMD, which significantly enhances cleavability of TREX1. We will investigate how the modulated processing of this and further TREX1 TMD mutations is connected to disease pathogenesis. With regard to the protease SPP, many aspects of substrate selection and further handling of cleavage products remain open to date. We consider TREX1 as a suitable model substrate to approach these SPP-related mechanistic questions to better understand SPP as part of ER proteostasis pathways. Altogether, this project will provide essential insights into cell biological mechanisms regulating nucleic acid-driven immune responses as well as how a dysregulation of these processes can lead to human disease.

三个主要修复核酸外切酶1（TREX 1）基因的突变导致几种自身炎症性疾病，包括Aicardi-Goutières综合征（AGS）。TREX 1是一种DNA降解酶，可以从细胞质中清除DNA。从细胞的角度来看，细胞质中的DNA可以指示病原体的入侵。因此，像cGAS-STING通路这样的DNA传感机制已经进化成触发防御机制，如干扰素的产生。然而，通过积累DNA过度激活这些途径，例如在不存在TREX 1的情况下，导致免疫病理学TREX 1被合成为通过C-末端跨膜结构域（TMD）连接到ER膜的尾锚定蛋白。这与在细胞核中也检测到TREX 1的几份报告相矛盾。我们已经确定TREX 1作为ER定位的膜内蛋白酶信号肽肽酶（SPP）的新底物。SPP的切割将可溶形式的TREX 1释放到胞质溶胶中。我们发现TREX 1可以组成性地存在于切割的可溶性以及膜结合的形式。然而，TREX 1切割也可导致其快速进一步降解，使得SPP也调节总细胞TREX 1水平。这些发现表明SPP对TREX 1的主要调节影响。由于SPP介导两种形式之间的转换，因此SPP缺陷细胞和小鼠的分析提供了一种防止可溶性TREX 1生成的系统。我们计划分析TREX 1切割在哪里以及在什么条件下发生，这如何影响切割产物的稳定性，以及这如何与核转位有关。一个中心问题是TREX 1从膜上的释放如何影响其降解由不同来源（包括逆转录转座子）产生的胞质DNA的能力。进一步考虑这一点，我们还将研究胞质TREX 1的存在或不存在如何通过外源性引入来调节干扰素应答的启动，以及病原体衍生的DNA或辐射后释放的DNA，如放射治疗。这也将在仅表达可溶性TREX 1的新型小鼠模型中进行分析。TREX 1蛋白水解是一个生物学相关过程的强有力支持是由TREX 1 TMD中的AGS引起的突变提供的，该突变显著增强了TREX 1的可切割性。我们将研究如何调制处理这个和进一步TREX 1 TMD突变是连接到疾病的发病机制。关于蛋白酶SPP，迄今为止，底物选择和切割产物进一步处理的许多方面仍然悬而未决。我们认为TREX 1作为一个合适的模型底物，以接近这些SPP相关的机制问题，以更好地了解SPP作为ER蛋白质稳态途径的一部分。总而言之，该项目将为调节核酸驱动的免疫反应的细胞生物学机制以及这些过程的失调如何导致人类疾病提供重要见解。