Ecology and functions of small SCIFF proteins in the gut microbiome

尽管肠道微生物组及其对宿主健康的影响引起了人们的极大兴趣，但在分子水平上对微生物-微生物和微生物-宿主相互作用的研究仍处于起步阶段。我们先前研究了猪肠道培养细菌中的生物合成基因簇（BGC），猪是生物医学研究和农业中的重要动物。虽然在分析的38个新分类群中，原核生物的BGC总数相对较低，但含有SCIFF型（45个残基中有6个半胱氨酸）小蛋白质的核糖体肽BGC富集，表明产生了新的sactipeptide，在24个菌株中检测到（63%的患病率），而在公开的基因组中<1%。这种广泛的发生和已知的抗菌肽的抗菌潜力表明编码的分子在塑造肠道生态系统中的重要作用。在这个项目中，我们联合收割机结合我们在厌氧培养，宏基因组学和生物分子天然产物化学方面的专业知识，（i）研究小SCIFF蛋白的加工，（ii）结构表征其成熟产物，（iii）功能研究其在肠道微生物组中的作用。这些产品将通过合成生物学技术进行研究，用于途径拦截，重组和异源表达，并结合最先进的化学分析进行分离和表征。来自猪肠的培养菌株及其简化群落将用于分批和连续培养系统中，以评估调节靶小蛋白产生的管腔因子。同时，将使用鸟枪分子分析在猪中测试这些小蛋白对体内天然微生物群落的影响。将与Z项目合作分析体外和体内实验的样品。总之，这个跨学科的项目将提供详细的功能性见解，以了解与肠道中微生物-宿主相互作用相关的新型小蛋白的生物学。它加强了联盟内现有的专业知识，这要归功于处理微生物群落、新模型系统、合成生物学和化学分析方法的独特组合。

Despite the major interest for intestinal microbiomes and their impact on host health, research on microbe-microbe and microbe-host interactions at the molecular level is in its infancy. We previously studied biosynthetic gene clusters (BGCs) in cultured bacteria from the intestine of pigs, which are important animals in both biomedical research and agriculture. Whilst the overall number of BGCs in the 38 novel taxa analyzed was relatively low for prokaryotic organisms, there was an enrichment for ribosomal peptide BGCs containing SCIFF-type (six cysteines in forty-five residues) small proteins pointing at the production of novel sactipeptides, which were detected in 24 strains (63% prevalence) compared with <1% in publicly available genomes. This widespread occurrence and the antibacterial potential of known sactipeptides suggest an important role of the encoded molecules in shaping the intestinal ecosystem. In this project, we combine our expertise in anaerobic cultivation, metagenomics, and biomolecular natural products chemistry to (i) study the processing of the small SCIFF proteins, (ii) structurally characterize their matured products, and (iii) functionally study their roles within gut microbiomes. The products will be investigated via synthetic biology techniques for pathway interception, recombination and heterologous expression combined with state-of-the-art chemical analytics for isolation and characterization. Cultured strains from the pig intestine and simplified communities thereof will be used in batch and continuous culture systems to assess luminal factors that regulate production of the target small proteins. In parallel, the impact of these small proteins on native microbial communities in vivo will be tested in pigs using shotgun molecular analyses. Samples from both in vitro and in vivo experiments will be analyzed in collaboration with the Z-projects. In summary, this interdisciplinary project will provide detailed functional insights into the biology of novel small proteins of relevance for microbe-host interactions in the gut. It strengthens current expertise within the consortium thanks to a unique combination of approaches dealing with microbial communities, new model systems, synthetic biology, and chemical analytics.