LEAPS-MPS: Determining the Mechanisms by Which Alarmone Signaling in Clostridioides Difficile Differs From Tthat in other Bacteria

在数学和物理科学理事会以及化学系的支持下，老道明大学的Erin B. Purcell教授和她的团队将研究被称为“警报器”的核苷酸信号分子。“不同类型的细菌通过合成不同的化学物质来应对环境压力，例如四磷酸盐和五磷酸盐警报器，以调节生存机制。”直到最近，较小的三磷酸盐警报器被认为是较大的信号分子的降解产物。然而，最近发现一些细菌除了直接合成四磷酸和五磷酸信号外，还直接合成三磷酸警报器。迄今为止，厌氧芽孢形成细菌艰难梭菌是唯一的细菌报道合成三磷酸警报器专门。珀塞尔实验室的研究生和本科生学员打算确定这种生物的警报器合成和利用与其他细菌有何不同。该项目有望扩大对警报信号的理解，并建立仅由三磷酸警报信号的信号范式。该项目将为培养基础生物化学技术的研究生和本科生提供机会。研究生研究人员，大部分来自代表性不足的群体，也将获得作为本科生受训人员的共同导师的经验，以培养他们作为未来科学领袖和榜样的领导技能。该项目的目标是确定这种生物独特的警报器合成的结构基础，以及不完全表征的三磷酸警报器pGpp所起的调节作用。Purcell小组特别感兴趣的是发现梭状芽胞酮合成酶的活性位点残基参与水解其核苷酸底物上的磷酸键以产生pGpp。该项目的成功实施还将确定梭状芽孢杆菌警报器水解酶和警报器结合效应物是否能够识别附近细菌产生的较大的警报器，从而潜在地利用警报器进行细胞间和种间通信以及细胞内信号传导。该项目有可能鉴定艰难梭菌中由警报信号调节的基因和过程，以确定它是否以保守的方式使用其非规范警报信号，或者该生物体中警报信号通路的输出是否也是独特的。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

With the support of the Mathematical and Physical Sciences Directorate and the Division of Chemistry, Professor Erin B. Purcell and her group at Old Dominion University will study nucleotide signal molecules known as ‘alarmones.’ Bacteria across diverse types respond to environmental stress by synthesizing different chemical species, such as tetraphosphate and pentaphosphate alarmones, to regulate survival mechanisms. Until recently, smaller triphosphate alarmones were thought to be degradation products of the larger signal molecules. However, it has recently been discovered that some bacteria synthesize triphosphate alarmones directly in addition to the tetra- and pentaphosphate signals. To date, the anaerobic spore-forming bacterium Clostridioides difficile is the only bacterium reported to synthesize triphosphate alarmones exclusively. Graduate and undergraduate trainees in the Purcell lab intend to determine how this organism’s alarmone synthesis and utilization differ from those in other bacteria. This project is expected to expand understanding of alarmone signaling and establish a paradigm for signaling by the triphosphate alarmone alone. This project will provide opportunities to train graduate and undergraduate researchers in foundational biochemistry techniques. Graduate researchers, mostly from underrepresented groups, will also gain experience as co-mentors of undergraduate trainees to nurture their leadership skills as future scientific leaders and role models.The objectives of this project will be to determine the structural basis of this organism’s unique alarmone synthesis and the regulatory role played by the incompletely characterized triphosphate alarmone, pGpp. The Purcell group is especially interested in discovering which active site residues in the clostridial alarmone synthetase enzymes are involved in hydrolyzing phosphate bonds on its nucleotide substrates to generate pGpp. Successful implementation of this project will also determine whether clostridial alarmone hydrolases and alarmone-binding effectors can recognize the larger alarmones produced by nearby bacteria, potentially utilizing alarmones for intercellular and interspecies communication as well as intracellular signaling. This project has the potential to identify the genes and processes regulated by alarmone signaling in C. difficile to determine whether it uses its non-canonical alarmone in a conserved manner or whether the outputs of alarmone signaling pathways in this organism are also unique.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.