Genomic and metabolomic foundations of human-microbial symbiosis in the gut

描述（由申请人提供）：人类肠道微生物群由细菌的拟杆菌门和厚壁菌门部门主导，并为我们提供了我们不必自己进化的生理特征，包括处理否则难以消化的膳食多糖的能力。这项资助更新的重点是表征微生物群关键物种的基因组和“代谢组学”特征，其营养物质用于衰老/共享行为的操作原则，以及我们从与它们的互利（共生）关系中获得的生理益处。我们使用了无菌（GF）小鼠定殖与多形拟杆菌（B。theta），一种糖分解厌氧菌，进行了首次报道的体内重要人类肠道共生体的全基因组转录谱分析。功能基因组学和基于质谱的代谢组学研究显示了B。theta的适应性觅食行为可以帮助稳定微生物群，以应对膳食多糖的变化。我们现在建议系统地增加这种无菌小鼠模型的复杂性，以检查我们已经测序的人源拟杆菌之间的相互作用（B.θ，B。vulgatus，B. distasonis）、史氏甲烷短杆菌（Methanobrevibacter smithii）（人结肠中的优势产甲烷古菌）和厚壁菌门（Firmicutes）的成员（直肠真杆菌（Eubacterium rectale）、大肠杆菌（E. eligens）。目标1.描述拟杆菌-细菌共生的基因组和代谢基础及其对宿主生物学的影响，如下：（i）生产成品，注释M。史密斯氏杆菌基因组（第一个待测序的甲烷短杆菌属物种）;（ii）定殖GF小鼠，其喂食标准的富含多糖的食物、富含果聚糖的食物或具有B的高脂肪西方饮食。θ和/或M。smithii;（iv）使用激光捕获显微切割的结肠上皮和小鼠基因芯片同时分析宿主转录应答;（v）基于转录组数据集进行代谢组的二氧化硅重建，并使用结果指导假设-基于微生物代谢途径对互惠共生的贡献的生物化学和遗传分析;（vi）使用B评估我们的发现的一般性。vulgatus和B. distasonis。目标2.描述厚壁菌门在简化的人体微生物群3组分模型中对营养共享的贡献，该模型由该部门的一个重要成员，拟杆菌门的一个成员加上产甲烷菌组成。我们将通过用E.直肠（人类结肠微生物群的一个成员，我们目前正在对其基因组进行测序），B。theta和M.史密斯。然后将微生物和宿主的转录和代谢反应作为群落成员和饮食的函数进行评估。基于这些结果，我们将研究含有拟杆菌属的选定组合的聚生体的性质，M. smithii和E.直肠E.精英这些研究应该为监测和操纵微生物群的功能提供新的方法，以促进人类健康。

DESCRIPTION (provided by applicant): The human gut microbiota is dominated by the Bacteroidetes and Firmicutes divisions of Bacteria, and provides us with physiological traits we have not had to evolve on our own, including the ability to process otherwise indigestible dietary polysaccharides. This grant renewal focuses on characterizing the genomic and "metabolomic" features of the microbiota's keystone species, the operating principles that underlie their nutrient for aging/sharing behaviors, and the physiological benefits we accrue from our mutualistic (symbiotic) relationship with them. We have used germ-free (GF) mice colonized with Bacteroides thetaiotaomicron (B. theta), a saccharolytic anaerobe, to conduct the first reported genome-wide transcriptional profiling of a prominent human gut symbiont in vivo. Functional genomic and mass spectrometry-based metabolomic studies showed how B. theta's adaptive foraging behavior could help stabilize the microbiota in the face of changes in dietary polysaccharides. We now propose to systematically add complexity to this gnotobiotic mouse model to examine interactions between human-derived Bacteroidetes we have sequenced (B. theta, B. vulgatus, B. distasonis), Methanobrevibacter smithii (a dominant methanogenic archaeon in the human colon), and members of Firmicutes (Eubacterium rectale, E. eligens). Aim 1. Characterize the genomic and metabolic underpinnings of Bacteroidetes-Archaeal mutualism and its impact on host biology as follows: (i) produce a finished, annotated M. smithii genome (first Methanobrevibacter species to be sequenced); (ii) colonize GF mice fed standard polysaccharide-rich chow, a chow enriched for fructans, or a high fat Western diet with B. theta and/or M. smithii; (iii) perform whole-genome transcriptional profiling of both microbial species in vivo using custom GeneChips; (iv) simultaneously profile host transcriptional responses using laser capture microdissected colonic epithelium and mouse GeneChips; (v) perform in silica reconstructions of the metabolome based on the transcriptome datasets and use the results to direct hypothesis-based biochemical and genetic analyses of the contributions of microbial metabolic pathways to the mutualism; (vi) assess the generality of our findings using B. vulgatus and B. distasonis. Aim 2. Characterize the contributions of the Firmicutes to nutrient sharing in a simplified 3-component model of the human microbiota composed of a prominent member of this division, a member of Bacteroidetes, plus a methanogen. We will create an initial community by colonizing GF mice with E. rectale (a member of the human colonic microbiota whose genome we are currently sequencing), B. theta and M. smithii. Microbial and host transcriptional and metabolic responses will then be assessed as a function of community membership and diet. Based on the results, we will examine the properties of consortia containing selected combinations of Bacteroides spp., M. smithii, and E. rectale/E. eligens. These studies should provide new approaches for monitoring and manipulating the functions of the microbiota to promote human health.