Microparticles enhance the formation of seven major classes of natural products in native and metabolically engineered actinobacteria through accelerated morphological development.

Actinobacteria provide a rich spectrum of bioactive natural products and therefore display an invaluable source towards commercially valuable pharmaceuticals and agrochemicals. Here, we studied the use of inorganic talc microparticles (hydrous magnesium silicate, 3MgO·4SiO2·H2O, 10 µm) as a general supplement to enhance natural product formation in this important class of bacteria. Added to cultures of recombinant Streptomyces lividans, talc (10 g L-1) enhanced production of the macrocyclic peptide antibiotic bottromycin A2 and its methylated derivative Met-bottromycin A2 up to 43%. Hereby, the microparticles fundamentally affected metabolism. With talc, S. lividans grew to 40% smaller pellets and, using RNA sequencing, revealed accelerated morphogenesis and aging, indicated by early upregulation of developmental regulator genes such as ssgA, ssgB, wblA, sigN and bldN. Furthermore, the microparticles re-balanced the expression of individual bottromycin cluster genes, resulting in a higher macrocyclization efficiency at the level of BotAH and correspondingly lower levels of non-cyclized shunt by-products, driving the production of mature bottromycin. Testing a variety of Streptomyces species, talc addition resulted in up to 13-fold higher titers for the RiPPs bottromycin and cinnamycin, the alkaloid undecylprodigiosin, the polyketide pamamycin, the tetracycline-type oxytetracycline, and the anthramycin-analogues usabamycins. Moreover, talc addition boosted production in other actinobacteria, outside of the genus of Streptomyces: vancomycin (Amycolatopsis japonicum DSM 44213), teicoplanin (Actinoplanes teichomyceticus ATCC 31121), and the angucyclinone-type antibiotic simocyclinone (Kitasatospora sp. DSM 102431). For teicoplanin, the microparticles were even crucial to activate production. Taken together, the use of talc was beneficial in 75% of all tested cases and optimized natural and heterologous hosts forming the substance of interest with clusters under native and synthetic control. Given its simplicity and broad benefits, microparticle-supplementation appears as an enabling technology in natural product research of these most important microbes. Whole transcriptomes of a bottromycin production strain derived from three time points and two treatments (+/- talc microparticles) using 3 biological replicates each.

放线菌门（Actinobacteria）可产生丰富多样的生物活性天然产物，因此是极具商业价值的药物与农用化学品来源。本研究探讨了将无机滑石微粒（talc microparticles，水合硅酸镁，3MgO·4SiO2·H2O，粒径10 µm）作为通用添加剂，以提升这类重要细菌的天然产物合成能力的方案。将浓度为10 g·L⁻¹的滑石添加到重组变铅青链霉菌（Streptomyces lividans）的培养体系中，可使大环肽类抗生素波卓霉素A2（bottromycin A2）及其甲基化衍生物Met-波卓霉素A2的产量提升最高达43%。在此过程中，滑石微粒从根本上重塑了菌体的代谢模式：添加滑石后，变铅青链霉菌形成的菌团体积减小40%；通过RNA测序（RNA sequencing）分析发现，菌体的形态发生与衰老进程加速，具体表现为发育调控基因（如ssgA、ssgB、wblA、sigN及bldN）的早期上调表达。此外，滑石微粒还重新平衡了波卓霉素合成基因簇中各基因的表达水平，使得BotAH介导的大环化效率提升，同时减少了非环化分流副产物的生成，最终促进成熟波卓霉素的合成。对多种链霉菌属（Streptomyces）菌种进行测试后发现，添加滑石可使核糖体合成并经翻译后修饰的肽（RiPPs）类化合物波卓霉素与辛那霉素（cinnamycin）、生物碱类的十一烷基灵菌红素（undecylprodigiosin）、聚酮类的帕马霉素（pamamycin）、四环素类的土霉素（oxytetracycline）以及蒽环霉素类似物乌沙霉素（usabamycins）的效价提升最高达13倍。不仅如此，添加滑石还能提升链霉菌属以外的其他放线菌的产物合成能力：日本拟无枝酸菌（Amycolatopsis japonicum DSM 44213）生产的万古霉素（vancomycin）、替考游动放线菌（Actinoplanes teichomyceticus ATCC 31121）生产的替考拉宁（teicoplanin），以及北里孢菌属（Kitasatospora sp. DSM 102431）生产的安格拉环酮类抗生素西莫环素（simocyclinone）。其中对于替考拉宁而言，滑石微粒甚至是激活其合成的关键因素。综上，在所有测试案例中，添加滑石的方案在75%的情况下均能发挥有益作用，可优化天然宿主与异源宿主的目标产物合成，无论产物合成基因簇受天然调控还是人工合成调控。鉴于该方法操作简便且适用范围广泛，微粒添加策略有望成为这类重要微生物天然产物研究中的一项赋能技术。本研究包含一株波卓霉素生产菌株的完整转录组数据：该菌株在三个时间点、两种培养条件（添加/不添加滑石微粒）下分别培养，每组设置3次生物学重复。