Structural transitions and cellular remodelling in spore germination

孢子萌发中的结构转变和细胞重塑

• 批准号: BB/W015072/1
• 负责人: Per Bullough
• 金额: $ 113.04万
• 依托单位: University of Sheffield
• 依托单位国家: 英国
• 项目类别: Research Grant
• 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=BB%2FW015072%2F1
• 关键词: Structural; transitions; cellular; remodelling; spore

Bacteria such as those causing botulism and anthrax survive harsh conditions and spread disease as spores. However for these spores to cause disease symptoms they must germinate into so-called vegetative cells. Toxins produced by the vegetative cells are most often the main factor in making infected animal or human hosts sick. We wish to understand how this germination takes place and how the active vegetative cell emerges from the dormant spore. This is a remarkable metamorphosis where one intricate type of cell structure is completely transformed into a radically different structure- it is analogous to the metamorphosis of a seed into a seedling and arguably as complex! The process is also interesting because vegetative cells are much more vulnerable to attack, for example by antibiotics or disinfectants, than spores are. Thus if we could work out how to 'germinate to exterminate', we could develop new weapons against a number of diseases and food-spoilage organisms. We will study germination in two selected organisms- Clostridium sporogenes (C. sporogenes) and Clostridioides difficile (C. difficile). C. sporogenes is non-pathogenic but very closely related to C. botulinum which causes botulism, a potentially lethal paralysis. Spores are extremely heat resistant so eradication from food requires costly heat treatment that lowers nutritional value & makes the product less appealing, yet there is increasing demand for minimally processed, safe foods. C. botulinum is also listed as a potential bio-weapon (UK Anti-terrorism, Crime & Security Act 2001). The advantage of using the harmless C. sporogenes is that we can learn a lot about C. botulinum without the need for highly specialised containment facilities. C. difficile is the leading cause of antibiotic-associated diarrhoea, especially in hospital patients and the elderly. Disease occurs when ingested spores germinate in response to bile acids in the gut. The toxins are the major cause of disease & in many cases, death. Ungerminated spores remaining in the gut can be the source of recurrent infections even after treatment.Our multidisciplinary project will apply state of the art techniques of structural and molecular biology to investigate the way these spores germinate. A crucial aspect of this work will involve mapping the proteins that make up spores in 3D molecular detail. To achieve this, we will exploit exciting new developments in microscopy - such as cryo-electron tomography - that allow us to visualise spores and cells in unprecedented detail. We will create 3D images of the spore where we can even identify and locate individual molecules. We will then go on to visualise the structures of spores as they transform into active cells, as might happen when contaminated food is ingested. The project will exploit new developments in imaging methods that allow us to see the structures more clearly. It will involve combinations of molecular genetics, use and development of electron light and scanning probe imaging approaches and computational image processing. Ultimately we aim to make a 3D movie that explains the mechanical and biochemical changes that take place when the active vegetative cell emerges out of the spore.

细菌（例如引起肉毒杆菌和炭疽的细菌）在恶劣的条件下幸存下来，并将疾病作为孢子传播。但是，要使这些孢子引起疾病症状，它们必须发芽到所谓的营养细胞中。植物细胞产生的毒素通常是使感染动物或人类宿主生病的主要因素。我们希望了解这种发芽是如何发生的，以及活性营养细胞如何从休眠的孢子中出现。这是一种显着的变形，其中一种复杂的细胞结构完全转化为根本不同的结构 - 它类似于种子的变形为幼苗，可以说是复杂的！这一过程也很有趣，因为与孢子相比，营养细胞更容易受到攻击，例如抗生素或消毒剂。因此，如果我们可以弄清楚如何“发芽以消灭”，我们就可以针对多种疾病和食物杂货生物开发新武器。我们将研究两个选定的生物体 - 孢子梭菌（C. sporogenes）和梭状芽胞杆菌艰难梭菌（艰难梭菌）。 C.孢子虫是非致病性的，但与肉毒杆菌c. c. c. c. c. c. c. c. c. c. c. c. c. c. c. c. c. c. c. to c. botulim c. botulasism c. c. c. c. c. c. opogenes c. c. c. c. c. c. c. opogenes。孢子具有极高的耐热性，因此消除食物需要昂贵的热处理，以降低营养价值并使产品吸引人的吸引力降低，但对最少加工的安全食品的需求增加。肉毒杆菌也被列为潜在的生物武器（英国反恐怖主义，2001年犯罪与安全法）。使用无害的C.孢子虫的优点是，我们可以在不需要高度专业化的围护设施的情况下学到很多有关肉毒杆菌的知识。艰难梭菌是抗生素相关腹泻的主要原因，尤其是在医院患者和老年人中。当对肠道中的胆汁酸发芽时摄入孢子发芽时，就会发生疾病。毒素是疾病的主要原因，在许多情况下，死亡。即使在治疗后，肠道中剩余的无孢子也可能是复发感染的根源。我们的多学科项目将采用结构和分子生物学的最先进技术来研究这些孢子的发芽方式。这项工作的关键方面将涉及映射以3D分子细节组成孢子的蛋白质。为了实现这一目标，我们将利用显微镜中令人兴奋的新发展（例如冷冻电子断层扫描），使我们能够以前所未有的细节可视化孢子和细胞。我们将创建孢子的3D图像，甚至可以识别和定位单个分子。然后，我们将继续可视化孢子变成活性细胞的结构，就像摄入受污染的食物时可能发生的那样。该项目将利用成像方法中的新发展，使我们能够更清楚地看到结构。它将涉及分子遗传学的组合，电子光的使用和开发以及扫描探针成像方法以及计算图像处理。最终，我们的目标是制作一部3D电影，以解释活跃的营养细胞从孢子中出现时发生的机械和生化变化。