It is generally believed that exchange of secondary metabolite biosynthetic gene clusters (BGCs) among closely related bacteria is an important driver of BGC evolution and diversification. Applying this idea may help researchers efficiently connect many BGCs to their products and characterize the products' roles in various environments. However, existing genetic tools support only a small fraction of these efforts. Here, we present the development of chassis-independent recombinase-assisted genome engineering (CRAGE), which enables single-step integration of large, complex BGC constructs directly into the chromosomes of diverse bacteria with high accuracy and efficiency. To demonstrate the efficacy of CRAGE, we expressed three known and six previously identified but experimentally elusive non-ribosomal peptide synthetase (NRPS) and NRPS-polyketide synthase (PKS) hybrid BGCs from Photorhabdus luminescens in 25 diverse γ-Proteobacteria species. Successful activation of six BGCs identified 22 products for which diversity and yield were greater when the BGCs were expressed in strains closely related to the native strain than when they were expressed in either native or more distantly related strains. Activation of these BGCs demonstrates the feasibility of exploiting their underlying catalytic activity and plasticity, and provides evidence that systematic approaches based on CRAGE will be useful for discovering and identifying previously uncharacterized metabolites.

Summary There is a dynamic reciprocity between plants and their environment: soil physiochemical properties influence plant morphology and metabolism, and root morphology and exudates shape the environment surrounding roots. Here, we investigate the reproducibility of plant trait changes in response to three growth environments. We utilized fabricated ecosystem (Eco FAB ) devices to grow the model grass Brachypodium distachyon in three distinct media across four laboratories: phosphate‐sufficient and ‐deficient mineral media allowed assessment of the effects of phosphate starvation, and a complex, sterile soil extract represented a more natural environment with yet uncharacterized effects on plant growth and metabolism. Tissue weight and phosphate content, total root length, and root tissue and exudate metabolic profiles were consistent across laboratories and distinct between experimental treatments. Plants grown in soil extract were morphologically and metabolically distinct, with root hairs four times longer than with other growth conditions. Further, plants depleted half of the metabolites investigated from the soil extract. To interact with their environment, plants not only adapt morphology and release complex metabolite mixtures, but also selectively deplete a range of soil‐derived metabolites. The Eco FAB s utilized here generated high interlaboratory reproducibility, demonstrating their value in standardized investigations of plant traits.

Soybean [Glycine max (L.) Merr.] seeds accumulate sucrose, raffinose family oligosaccharides (RFO), phytin, and small amounts of galactopinitols and fagopyritols during normal seed maturation. RFO and phytin are indigestible by non‐ruminant animals and contribute to decreased feed efficiency, reduced mineral adsorption, and phosphorous pollution in manure. Low raffinose, stachyose, and phytin seed may have imbibitional chilling sensitivity and reduced field emergence. The objective was to characterize the profiles of soluble carbohydrates in cotyledons, axis, and seed coats of low raffinose and stachyose (LRS) seeds expressing the mutant stc1 phenotype; in low raffinose, stachyose, and phytin (LRSP1, LRSP2) seeds expressing the mutant mips phenotype; and in normal raffinose, stachyose, and phytin (CHECK) seeds expressing the Stc1 and Mips phenotype during 17 stages of soybean seed development, maturation, and desiccation. Seventy percent of RFO accumulated after maximum seed dry weight during seed desiccation. LRS, LRSP1, and LRSP2 seeds had low RFO, but LRS seeds accumulated higher galactinol and di‐ and tri‐galactosides of myo‐inositol, d‐chiro‐inositol, and d‐pinitol earlier during seed maturation than CHECK, LRSP1, and LRSP2. LRSP1 and LRSP2 seed had low RFO and low galactosyl cyclitols during maturation and were reported to have imbibitional chilling sensitivity and reduced field emergence.

Microorganisms play vital roles in modulating organic matter decomposition and nutrient cycling in soil ecosystems. The enzyme latch paradigm posits microbial degradation of polyphenols is hindered in anoxic peat leading to polyphenol accumulation, and consequently diminished microbial activity. This model assumes that polyphenols are microbially unavailable under anoxia, a supposition that has not been thoroughly investigated in any soil type. Here, we use anoxic soil reactors amended with and without a chemically defined polyphenol to test this hypothesis, employing metabolomics and genome-resolved metaproteomics to interrogate soil microbial polyphenol metabolism. Challenging the idea that polyphenols are not bioavailable under anoxia, we provide metabolite evidence that polyphenols are depolymerized, resulting in monomer accumulation, followed by the generation of small phenolic degradation products. Further, we show that soil microbiome function is maintained, and possibly enhanced, with polyphenol addition. In summary, this study provides chemical and enzymatic evidence that some soil microbiota can degrade polyphenols under anoxia and subvert the assumed polyphenol lock on soil microbial metabolism.