Collaborative Research: OSIB: Chemical signaling in a host-microbe symbiosis

尽管人们认识到共生微生物的重要性，但人们对宿主和微生物之间使用的化学语言却知之甚少。目前，在细菌用来适应宿主环境的表达化学方面存在着知识缺口，识别相关的化学物质将大大有助于我们了解细菌如何能够在利基环境中定居和维持自己，或者在相同的生态位上击败其他微生物。我们的建议是高度跨学科的，我们的团队也是如此，他们结合了前沿遗传学、尖端分析测量科学和药物化学，以确定发光细菌鱼弧菌是如何使用一种小化合物来更好地定植其鱿鱼宿主的，同时也发现了其他尚未确定的分子。因此，我们的系统提供了一个切入点，以确定可能在其他有益和致病的定植系统中相关的化学信号，并在可访问的模型中阐明相关的靶标。我们将利用我们易处理的、对环境感兴趣的模型系统，与Expanding Your Horizons合作，在拉丁裔人口较多的大蒙特利湾地区，为5-10年级的年轻女孩创建一个动手推广研讨会。人们对识别和表征植物、动物和人类宿主-微生物群相互作用中宿主-微生物交流的小分子非常感兴趣。真棘鱼、短尾鱿鱼和鱼弧菌发光细菌之间的天然简单联系提供了一种共生关系，可以在现场进行询问，以发现功能化合物。宿主的细菌定植可通过复杂的细菌遗传和基因组方法获得，感染部位可以在活体动物中成像。我们的合作开发了新的方法来识别和研究这种共生中的小分子。这项工作的结果是鉴定了一种名为二酮基哌嗪(DKP)的有机小分子，它可能有助于形成共生生物膜和发光表型。我们使用成像质谱学来鉴定更多的生物膜相关分子，并通过结构解析和合成化学方法合成了大量的DKP。在这里，我们将利用我们在成像质谱学、结构分离和鉴定、正向遗传筛选和合成生物有机化学方面的跨学科专业知识来鉴定这种DKP的功能，并确定这种分子的生物合成来源，以探索它是否用于其他微生物-宿主系统。这一模式系统将使我们能够解决目前在理解共生细菌在宿主特定组织定居期间产生的小分子方面的知识差距。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Despite an appreciation of the importance of symbiotic microbes, the chemical language used between hosts and microorganisms is poorly understood. There is a current gap in knowledge when it comes to the expressed chemistry that bacteria utilize to adapt to host environments, and the identification of the chemical matter responsible would add significantly to our understanding of how bacteria are able to colonize and maintain themselves in niche environments or out-compete other microorganisms for the same niche. Our proposal is highly interdisciplinary, as is our team, incorporating forward genetics, cutting-edge analytical measurement science, and medicinal chemistry to identify how the glowing bacterium, Vibrio fischeri, uses a small chemical compound to better colonize its squid host while also discovering other molecules that have yet to be identified. Our system thus provides an entry point to identify chemical signals that may be relevant in other beneficial and pathogenic colonization systems and to elucidate the relevant targets in an accessible model. We will leverage our tractable, environmentally intriguing model system to create a hands-on outreach workshop with young girls in grades 5-10 in the greater Monterey Bay area, which has a high Latinx population, in collaboration with Expanding Your Horizons.There is significant interest in identifying and characterizing small molecules that underlie host-microbe communication in plant, animal, and human host-microbiome interactions. The naturally simple association between Euprymna scolopes bobtail squid and Vibrio fischeri luminescent bacteria provides a symbiotic relationship that can be interrogated in situ to discover functional compounds. Bacterial colonization of the host is accessible to sophisticated bacterial genetic and genomic approaches, and the site of infection can be imaged in live animals. Our collaborative efforts have developed novel methods to identify and study small molecules in this symbiosis. This work has resulted in the identification of a small organic molecule, termed diketopiperazine (DKP), as a potential contributor to symbiotic biofilm and luminescence phenotypes from V. fischeri. We have used imaging mass spectrometry to identify additional biofilm-associated molecules, and, using structure elucidation and synthetic chemistry, we have synthesized large amounts of the DKP. Here, we will leverage our interdisciplinary expertise in imaging mass spectrometry, structure isolation and elucidation, forward genetic screening, and synthetic bioorganic chemistry to identify the function of this DKP and identify the biosynthetic origin of this molecule to explore whether it is used in other microbial-host systems. This model system will allow us to address current knowledge gaps in our understanding of the small molecules that symbiotic bacteria produce during colonization of host specific tissues.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.