Signalling and regulation of autophagy-related pathways during cellular differentiation of African trypanosomes.

非洲锥虫细胞分化过程中自噬相关途径的信号传导和调节。

• 批准号: MR/W026996/1
• 负责人: Mathieu Cayla
• 金额: $ 156.51万
• 依托单位: University of York
• 依托单位国家: 英国
• 项目类别: Fellowship
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FW026996%2F1
• 关键词: Signalling; regulation; autophagy; related; pathways

For a long time, lysosomes have been seen as waste management organelles. Over the last two decades, however, this view has evolved. Lysosomes are now considered as central organelles for the maintenance of homeostasis (a state of steady internal, physical and chemical conditions maintained by the cell for proper functioning), cellular survival and differentiation. Autophagy (from greek "Auto" self and "phagos" eating) is an essential process by which cells target intracellular components, macromolecules, organelles or intracellular pathogens to the lysosome for degradation and recycling. Autophagy is essential during cell growth, differentiation, or in response to stressors such as nutrient limitation. Moreover, defects or loss of regulation of autophagy can contribute to cancer or to the progression of neurodegenerative diseases. In eukaryotes, protein kinases and kinase enzyme complexes create protein phosphorylations. These reversible regulatory modifications are essential to control autophagy and therefore promote cell survival and differentiation. Recent studies have identified an unconventional secretory pathway called lysosome exocytosis that is dependent on the autophagy machinery. Lysosome exocytosis is a fundamental mechanism for the release of proteins that lysosomes could not degrade, thus avoiding potential toxic effect for the cell. Additionally, it has been implicated in the breach of host cellular membrane to allow invasion by the worm C. elegans. Indeed, the worm releases its lysosome contents directly into the extracellular environment to perforate the membrane of the host cells, allowing their invasion by the worm within the infected host.Studying how protein kinases regulate autophagy and lysosome exocytosis by phosphorylation events is of major importance to understand how insect-transmitted parasites invade and survive in human or animal hosts. These organisms persist in changing environments that demand finely-tuned and rapid adaptation for survival. A tractable model to explore the diversity of eukaryotic signalling networks are kinetoplastid parasites. Kinetoplastid parasites are of great clinical relevance affecting millions of people and animals with diseases such as Leishmaniasis, Chagas disease and Sleeping Sickness, leading to critical social-economic implications. Kinetoplastid are exquisitely sensitive to their environment and require autophagy to adapt drastic changes during host transitions.I propose to use the kinetoplastid parasite Trypanosoma brucei (causative agent of African sleeping sickness and livestock 'nagana') to characterise signalling pathways that regulate autophagy and lysosome exocytosis during lifecycle differentiation. In this project, I will develop an unbiased approach that will combine the optimisation of a high-throughput live imaging system, a gene silencing screen targetting all protein kinases encoded in the genome and mass spectrometry. Results obtained will reveal the molecular composition and regulations by phosphorylation of autophagy and lysosome exocytosis in T. brucei. The function of these regulatory components will then be evaluated for their roles in differentiation during the parasite lifecycle. The results of this study will not only provide a better understanding of how kinetoplastid parasites adapt to the different hosts encountered during their complex lifecycles and persist in vivo, but will lay the foundations for the use of T. brucei as model to study the regulation of, and crosstalk between, the autophagy and exocytosis pathways.

长期以来，溶酶体被视为废物管理细胞器。然而，在过去的二十年里，这种观点已经发生了变化。溶酶体现在被认为是维持稳态（细胞维持正常功能的稳定内部，物理和化学条件的状态），细胞存活和分化的中心细胞器。自噬（英语：Autophagy）是细胞将细胞内成分、大分子、细胞器或细胞内病原体靶向溶酶体进行降解和再循环的一个重要过程。自噬在细胞生长、分化过程中或对营养限制等应激源的反应中是必不可少的。此外，自噬调控的缺陷或丧失可导致癌症或神经退行性疾病的进展。在真核生物中，蛋白质激酶和激酶复合物产生蛋白质磷酸化。这些可逆的调节修饰对于控制自噬并因此促进细胞存活和分化是必不可少的。最近的研究已经确定了一种非常规的分泌途径，称为溶酶体胞吐作用，它依赖于自噬机制。溶酶体胞吐作用是释放溶酶体不能降解的蛋白质的基本机制，从而避免对细胞的潜在毒性作用。此外，它还涉及宿主细胞膜的破裂，从而允许蠕虫C的入侵。优美的事实上，蠕虫将其溶酶体内容物直接释放到细胞外环境中以覆盖宿主细胞的膜，从而允许蠕虫在受感染的宿主中入侵。研究蛋白激酶如何通过磷酸化事件调节自噬和溶酶体胞吐对于理解虫传寄生虫如何入侵并在人类或动物宿主中存活具有重要意义。这些有机体在不断变化的环境中生存，需要微调和快速适应生存。动质体寄生虫是探索真核生物信号网络多样性的一个易处理的模型。动体寄生虫具有重要的临床意义，影响数百万人和动物的疾病，如利什曼病，恰加斯病和昏睡病，导致严重的社会经济影响。动质体是精致的敏感，他们的环境，并需要自噬，以适应宿主transitions.I期间的剧烈变化，建议使用动质体寄生虫布氏锥虫（非洲昏睡病和牲畜的病原体“Nagana”）来调节自噬和溶酶体胞吐在生命周期分化的信号通路。在这个项目中，我将开发一种无偏见的方法，将结合联合收割机的高通量实时成像系统，基因沉默筛选靶向基因组中编码的所有蛋白激酶和质谱的优化。这些结果将揭示T.布鲁塞。然后将评估这些调节组分在寄生虫生命周期中分化的作用。这项研究的结果不仅将更好地了解动质体寄生虫在其复杂的生命周期中如何适应不同的宿主并在体内持续存在，而且将为T.布鲁氏杆菌作为模型来研究自噬和胞吐途径的调节以及它们之间的相互作用。

项目成果

Differentiation granules, a dynamic regulator of T. brucei development.

分化颗粒，布氏锥虫发育的动态调节剂。

• DOI: 10.21203/rs.3.rs-3442788/v1
• 发表时间: 2023
• 作者: Cayla M