Campylobacter pathogenesis: the Unfolded Protein Response (UPR), inflammation and human disease

弯曲杆菌发病机制：未折叠蛋白反应 (UPR)、炎症和人类疾病

• 批准号: 2881697
• 金额: --
• 依托单位: London School of Hygiene & Tropical Medicine
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2881697
• 关键词: Campylobacter; pathogenesis; Unfolded; Protein; Response

Infectious diarrhoea is a global problem with Campylobacter being the most common bacterial cause. Despite its importance, the mechanisms by which Campylobacter infection promotes inflammation and disease in humans remain unclear. Campylobacter infection is the most common bacterial cause of human diarrhoeal disease. The species Campylobacter jejuni is responsible for over 80% of human cases with symptoms typically including bloody diarrhoea, fever and abdominal pains. A key precursor of diarrhoea is inflammation triggered when C. jejuni invades human intestinal epithelial cells (IECs), leading to tissue damage and disease. The cell has developed a system for maintaining transcriptional fidelity of the ER - the unfolded protein response (UPR). Although it mainly acts upon imbalances in the homeostasis of unfolded proteins, several bacterial species like Campylobacter have been shown to activate this pathway to increase intracellular survival. Unpublished data from Gong et al. demonstrate campylobacter mediated UPR activation through the PERK and IRE1 arms. The paper however stopped short of identifying the molecular drivers that activate the different arms of the UPR. Reactive oxygen species (ROS) are a group of oxygen-based chemical intermediaries with an uneven number of electrons. Normally, ROS production is mediated by the activation of the nitrous oxide (NOX) pathway and is counteracted by several defence mechanisms, such as antioxidant release and a balance, the 'Redox' state, is maintained. Interestingly, several manuscripts have established an antimicrobial role of ROS. Previous work demonstrate that Campylobacter can modulate ROS production pathway components to aid intracellular survival and proliferation: C. jejuni differentially regulates intracellular and extracellular ROS production in human T84 and Caco-2 cells. C. jejuni downregulates the transcription and translation of nicotinamide adenine dinucleotide phosphate (NAPDH) oxidase (NOX1), a key ROS-generating enzyme in IECs and antioxidant defence genes CAT and SOD1. Furthermore, inhibition of NOX1 by diphenylene iodonium (DPI) and siRNA reduced C. jejuni ability to interact, invade, and survive within T84 and Caco-2 cells. The UPR and NOX pathways are activated to aid intracellular bacteria clearance and form part of the autonomous cell response but are susceptible to bacterial influence. The project will aim to investigate the interplay between the UPR and ROS in Campylobacter mediated infection - before, during and after activation. Campylobacter mutants lacking certain virulence factors will be developed and their ability to invade host cells, as well as to activate the UPR and NOX pathways will be assessed. Transcriptional analysis of both bacterial and host cells during invasion assays will provide a better understanding of the genomic changes of both agents during the invasion cycle, and bacterial mutants will be used will be used to determine changes to the transcription of virulence factors, as well as host cell defence mechanisms. Particular attention will be given to changes in the transcriptome corresponding to increased inflammatory molecules such as interleukins. Different strains of C jejuni and various animal cell lines will be used to investigate the discrepancies in disease states observed in literature.The biochemical basis of both signalling pathways will be investigated, and techniques such as Mass-spec, ELISA and pulldown assays will be used to identify individual components of the pathways of UPR and ROS pathway activation.Once the core aspects of the project have been completed, the project become more expansive; It will combine 'Omic technologies to gain a better systemic understanding of the UPR and ROS systems when reacting to C jejuni infection and will relate the data to inflammatory processes. Other aims of the project include the development of a 3D cell culture model to better recapitulate in vivo environments.

传染性腹泻是弯曲杆菌是最常见的细菌原因的全球问题。尽管它的重要性，但弯曲杆菌感染促进人类炎症和疾病的机制尚不清楚。弯曲杆菌感染是人类腹泻病的最常见细菌原因。弯曲杆菌空肠的物种负责超过80％的人类病例患有症状，包括血液腹泻，发烧和腹部疼痛。腹泻的关键前体是炎症的炎症，当空肠梭菌侵入人类肠上皮细胞（IEC）时，导致组织损伤和疾病。该细胞已经开发了一种系统，用于维持ER的转录保真度 - 展开的蛋白质反应（UPR）。尽管它主要作用于展开的蛋白质稳态失衡，但已经证明了几种细菌种类，例如弯曲杆菌，可以激活这种途径，以增加细胞内存活率。 Gong等人的未发表数据。通过PERK和IRE1臂展示了弯曲杆菌介导的UPR激活。但是，该纸无法识别激活UPR不同臂的分子驱动器。活性氧（ROS）是一组基于氧的化学中介机构，具有不均匀的电子。通常，ROS的产生是通过一氧化二氮（NOX）途径的激活来介导的，并通过几种防御机制来抵消，例如抗氧化剂释放和平衡，即“氧化还原”状态。有趣的是，一些手稿已经确立了ROS的抗菌作用。先前的工作表明，弯曲杆菌可以调节ROS生产途径成分以帮助细胞内存活和增殖：J。Jejuni差异地调节了人T84和CACO-2细胞中细胞内和细胞外ROS的产生。 C. jejuni下调了烟酰胺腺苷二核苷酸磷酸（NAPDH）氧化酶（NOX1）的转录和翻译，这是IECS和抗氧化剂防御基因CAT和SOD1中一种关键的ROS-ROS生成酶。此外，二苯基碘（DPI）对NOX1的抑制作用和siRNA降低了J. Jejuni在T84和CACO-2细胞中相互作用，入侵和生存的能力。 UPR和NOX途径被激活以帮助细胞内细菌清除率，并构成自主细胞反应的一部分，但易受细菌影响。该项目将旨在研究弯曲杆菌介导的感染中UPR和ROS之间的相互作用 - 激活之前和之后。将开发缺乏某些毒力因子的弯曲杆菌突变体，并将评估其侵入宿主细胞的能力，并激活UPR和NOX途径。在入侵测定过程中对细菌和宿主细胞的转录分析将更好地了解两种药物在入侵周期中的基因组变化，并且将使用细菌突变体来确定毒力因子转录的变化以及宿主细胞防御机制。将特别注意与炎症分子（如白细胞介素）相对应的转录组的变化。 Different strains of C jejuni and various animal cell lines will be used to investigate the discrepancies in disease states observed in literature.The biochemical basis of both signalling pathways will be investigated, and techniques such as Mass-spec, ELISA and pulldown assays will be used to identify individual components of the pathways of UPR and ROS pathway activation.Once the core aspects of the project have been completed, the project become more expansive;它将结合使用“ OMIC技术”，以便在与C jejuni感染反应时对UPR和ROS系统有更好的系统性理解，并将数据与炎症过程相关联。该项目的其他目的包括开发3D细胞培养模型，以更好地概括体内环境。