Functional Multigenomic Screening of Human-Associated Bacteria for NF-κB-Inducing Bioactive Effectors
Functional Multigenomic Screening of Human-Associated Bacteria for NF-κB-Inducing Bioactive Effectors
复制标题
DOI：
10.1128/mbio.02587-19
复制
发表时间：
2019-11-01
期刊：
影响因子：
6.4
通讯作者：
Brady, Sean F.
中科院分区：
文献类型：
--
作者：
Estrela, Andreia B.;Nakashige, Toshiki G.;Brady, Sean F.
微生物群对人类宿主的影响至少部分是由其产生的小分子和蛋白质效应器驱动的。在这里，我们报告使用功能性多基因组筛选，以确定微生物编码的效应。在这项研究中，基因组DNA从116人类相关的细菌进行了克隆，并得到的多基因组文库进行了筛选，使用核因子-κ B报告（NF-κ B）测定。功能性多基因组学建立在功能性宏基因组学的概念基础上，但利用了人类相关细菌培养和测序的日益进步。发现赋予大肠杆菌NF-κ B诱导活性的效应基因编码四种一般类别的蛋白质：细胞壁水解酶、膜转运蛋白、脂多糖生物合成酶和功能未知的蛋白质。多基因组文库的紧凑性，这是由于能够正常化输入DNA的比例，应简化筛选库使用不同的异源主机和报告基因检测，增加发现新的效应基因的速率。重要人类相关的细菌被认为是编码生物活性的小分子和蛋白质，在人类健康和疾病中发挥着密切的作用。在这里，我们报告的创建和功能筛选的多基因组库构建使用基因组DNA从116个细菌在不同的网站在整个人体。筛选各个克隆中能够赋予大肠杆菌NF kappa B诱导活性的基因。NF-κ B是一种有用的报告分子，用于与免疫、发病机制和炎症相关的一系列细胞过程。与宏基因组文库的筛选相比，当构建多基因组文库时，归一化输入DNA比率的能力应有助于更有效地检查微生物的不同生物活性。多基因组筛选利用了人类微生物群培养和测序中不断增长的可用资源，并为更深入地研究肠道细菌与人类宿主相互作用的机制提供了起点。
The effect of the microbiota on its human host is driven, at least in part, by small-molecule and protein effectors it produces. Here, we report on the use of functional multigenomic screening to identify microbiota-encoded effectors. In this study, genomic DNA from 116 human-associated bacteria was cloned en masse, and the resulting multigenomic library was screened using a nuclear factor-kappa B reporter (NF-kappa B) assay. Functional multigenomics builds on the concept of functional metagenomics but takes advantage of increasing advances in cultivating and sequencing human-associated bacteria. Effector genes found to confer NF-kappa B-inducing activity to Escherichia coli encode proteins in four general categories: cell wall hydrolases, membrane transporters, lipopolysaccharide biosynthetic enzymes, and proteins of unknown function. The compact nature of multigenomic libraries, which results from the ability to normalize input DNA ratios, should simplify screening of libraries using diverse heterologous hosts and reporter assays, increasing the rate of discovery of novel effector genes.IMPORTANCE Human-associated bacteria are thought to encode bioactive small molecules and proteins that play an intimate role in human health and disease. Here, we report on the creation and functional screening of a multigenomic library constructed using genomic DNA from 116 bacteria found at diverse sites across the human body. Individual clones were screened for genes capable of conferring NF kappa B-inducing activity to Escherichia coli. NF-kappa B is a useful reporter for a range of cellular processes related to immunity, pathogenesis, and inflammation. Compared to the screening of metagenomic libraries, the ability to normalize input DNA ratios when constructing a multigenomic library should facilitate the more efficient examination of commensal bacteria for diverse bioactivities. Multigenomic screening takes advantage of the growing available resources in culturing and sequencing the human microbiota and generates starting points for more in-depth studies on the mechanisms by which commensal bacteria interact with their human host.