Investigations of an invertebrate secreted immune effector that influences microbial communities of the gut

动物的肠道被各种各样的微生物定植，最近的研究表明，肠道的微生物群落影响着宿主的各种生理过程，从大脑功能到发育和肥胖。因此，了解肠道微生物群落和宿主之间的相互作用是非常重要的。研究人员将使用海洋滤食性生物体--罗布斯塔(Ciona enterinalisA型)来研究动物如何利用其免疫系统的元素来调节与喜欢生活在肠道中的微生物的健康互动。由于Ciona是透明的，它代表了一种独特的模式生物，在其中可以直接看到肠道微生物群落。Ciona也是学生在课堂上以及圣彼得堡科学节或女孩海洋夏令营等公共宣传季节性项目中的一个很好的教学工具。研究人员将利用实验室中在受控条件下饲养的动物来帮助学生了解不同的微生物是如何在肠道中定居的，并说明动物使用的自然机制来影响这些微生物在肠道中的位置和如何定居。Ciona robusta(Ciona enterinalisA型)是一种无脊椎动物滤食性原索动物，将被用来研究宿主如何利用先天免疫元素与肠道微生物群谈判相互作用。该提案的重点是可变的免疫球蛋白样几丁质结合蛋白(VCBPs)。VCBP是双功能的；N-末端的Ig结构域识别细菌，C-末端的结构域与几丁质结合。VCBPs是了解细菌、真菌和肠道微生物组病毒之间复杂对话的关键。这些蛋白由脊椎动物肠道上皮细胞大量表达和分泌，具有与脊椎动物IgA相似的功能，包括调节细菌黏附和在粘膜表面形成生物膜。初步数据显示，当VCBP-C与一些溶原体结合时，可以观察到前驱体的诱导，这一过程决定了生物膜形成的结果，并可能影响新释放的噬菌体可能针对的其他细菌的生长。来自乔纳肠道的各种细菌和真菌分离株已经被培养出来，现在将利用无菌动物在体外和体内进行研究。该项目通过体外和体内实验相结合，将检测VCBP-C与肠道培养分离株(细菌和真菌)之间的相互作用；然后我们将检测体外(细菌)生物膜形成和无菌动物的定植的转录本，以帮助表征定植的遗传学。作为多功能蛋白质，VCBPs可能是微生物群细菌和真菌群落之间相互作用的桥梁；由于VCBPs影响前驱体的诱导，初步观察表明，前驱体具有广泛的宿主范围，因此这些病毒的激活可能会重塑微生物组的组成。Ciona模型提供了一个独特的机会来研究先天性免疫如何不仅防止病原体，而且调节微生物生态以实现和维持动态平衡。Ciona将继续作为一个平台，进一步探索免疫进化不仅是为了“防御”，而且是为了“调节肠道微生物生态”这一假说。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。

The gut of animals is colonized by a rich assortment of microbes and recent research has demonstrated that microbial communities of the gut influence all kinds of physiological processes of the host, from brain function, to development, and obesity. Hence, understanding the interplay of microbial gut communities and the host is highly important. The investigators will use Ciona robusta (Ciona intestinalis type A), a marine filter-feeding organism, to investigate how animals use elements of their immune system to regulate healthy interactions with microbes that like to live in the gut. As Ciona is transparent, it represents a unique model organism in which gut microbial communities can be visualized directly. Ciona also is a great teaching tool for students in the classroom and for public outreach in the PIs seasonal programs like the St. Petersburg Science Festival or the Oceanography Camp for Girls. The investigators will utilize animals reared in the lab under controlled conditions to help students understand how distinct microbes colonize the gut, and illustrate natural mechanisms used by animals to influence where and how these microbes settle within the gut. Ciona robusta (Ciona intestinalis type A), an invertebrate filter-feeding protochordate, will be used to investigate how the host utilizes elements of innate immunity to negotiate interactions with the gut microbiome. The proposal focuses on variable immunoglobulin-like chitin-binding proteins (VCBPs). VCBPs are bifunctional; the N-terminal Ig domains recognize bacteria and the C-terminal domain binds chitin. VCBPs are a key to understanding complex dialogues between bacteria, fungi and viruses of the gut microbiome. These proteins are expressed and secreted in high abundance by the gut epithelium in Ciona, and exhibit functions that resemble vertebrate IgA, including the modulation of bacterial adherence and formation of biofilms on mucosal surfaces. Preliminary data reveal that when VCBP-C binds to some lysogens, induction of prophages is observed, a process that shapes the outcome of biofilm formation and may impact the growth of other bacteria that may be targeted by the newly released phages. Various bacterial and fungal isolates, derived from the gut of Ciona, have been cultured and will now be studied both in vitro and in vivo utilizing germ-free animals. This project, via a combination of in vitro and in vivo experiments, will examine the interaction between VCBP-C and cultured isolates (bacterial and fungal) of the gut; we will then examine transcriptomes of in vitro (bacterial) biofilm formation and colonization of germ-free animals to help characterize the genetics of colonization. As multi-functional proteins, the VCBPs likely bridge interactions between bacterial and fungal communities of the microbiome; and because VCBPs influence the induction of prophages, which in preliminary observations have been shown to possess broad host ranges, it is likely that activation of these viruses can re-shape the composition of the microbiome. The Ciona model presents a unique opportunity to study how innate immunity not only protects against pathogens but also modulates microbial ecology to achieve and maintain homeostasis. Ciona will continue to serve as a platform to further explore the hypothesis that immunity evolved not just to "defend" but to "modulate microbial ecology" of the gut lumen.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.