Tracing the role of D14L as a regulator of arbuscular mycorrhizal symbiosis

Arbuscular mycorrhizal (AM) symbiosis between plants and AM fungi is conserved across the land plants and is the default nutrient uptake strategy for plants in nature. Plant recognition and accommodation of AM fungi requires signalling through two evolutionarily ancient pathways: the common symbiosis signalling pathway (CSSP) and the DWARF14-LIKE (D14L) pathway. D14L has diverse roles in plant development, including inhibition of mesocotyl elongation during seedling development and symbiotic perception of AM fungi in the soil. During D14L signalling in rice, the negative repressor SMAX1 is degraded and CSSP components are transcriptionally upregulated, indicating that the symbiotic function of D14L is to modulate the CSSP. Meanwhile, the transcription factor NSP2, which is implicated in AM symbiosis and is a target of the phosphate starvation response, has been found to operate upstream of D14L in Medicago. In this study, the functional conservation of D14L and its symbiotic role relative to the CSSP and NSP2 were defined in rice. To investigate the evolutionary history of D14L, trans-species complementation of the rice *d14l* mutant by *D14L* homologues from a range of AM and nonmycorrhizal plants was performed. This analysis confirmed the highly conserved nature of the D14L protein and revealed that D14L does not specify the mycorrhizal status of angiosperms. To assess the role of NSP2 and closely related NSP1 in AM symbiosis in rice, CRISPR mutants were generated and investigated for defects in fungal colonisation and mesocotyl development. It was found that both NSP1 and NSP2 are essential for symbiosis and mesocotyl development. Furthermore, the mesocotyl phenotype of *nsp* mutants could be rescued by application of D14L signalling chemicals, providing evidence that both NSP1 and NSP2 are upstream of D14L in rice. To examine the downstream components of D14L signalling, complementation lines of rice *d14l* with the CSSP component CCaMK were generated. It was found that a gain-of-function CCaMK could complement the *d14l* mutant, revealing that D14L signalling regulates the CSSP for AM symbiosis. These results place the D14L signalling pathway at the centre of NSP and CSSP function. Transcriptome analysis of gain-of-function CCaMK lines demonstrated that CCaMK upregulates the expression of potential presymbiosis genes and genes relevant to conditioning a plant for symbiosis. Overall, a model is proposed in which D14L signalling functions as a master switch that can integrate the plant nutrient status into a decision to rapidly launch or block symbiosis

Conditioning plants for arbuscular mycorrhizal symbiosis through DWARF14-LIKE signalling.

The evolutionarily ancient α/β hydrolase DWARF14-LIKE (D14L) is indispensable for the perception of beneficial arbuscular mycorrhizal (AM) fungi in the rhizosphere, and for a range of developmental processes. Variants of D14L recognise natural strigolactones and the smoke constituent karrikin, both classified as butenolides, and additional unknown ligand(s), critical for symbiosis and development. Recent advances in the understanding of downstream effects of D14L signalling include biochemical evidence for the degradation of the repressor SMAX1. Indeed, genetic removal of rice SMAX1 leads to the de-repression of symbiosis programmes and to the simultaneous increase in strigolactone production. As strigolactones are key to attraction of the fungus in the rhizosphere, the D14L signalling pathway appears to coordinate fungal stimulation and root symbiotic competency. Here, we discuss the possible integrative roles of D14L signalling in conditioning plants for AM symbiosis.Bill & Melinda Gates Foundatio

Systematic mapping of drug metabolism by the human gut microbiome

The human gut microbiome harbors hundreds of bacterial species with diverse biochemical capabilities, making it one of nature’s highest density, highest diversity bioreactors. Several drugs have been previously shown to be directly metabolized by the gut microbiome, but the extent of this phenomenon has not been systematically explored. Here, we develop a systematic screen for mapping the ability of the complex human gut microbiome to biochemically transform small molecules (MDM-Screen), and apply it to a library of 575 clinically used oral drugs. We show that 13% of the analyzed drugs, spanning 28 pharmacological classes, are metabolized by a single microbiome sample. In a proof-of-principle example, we show that microbiome-derived metabolism occurs in vivo, identify the genes responsible for it, and provide a possible link between its consequences and clinically observed features of drug bioavailability and toxicity. Our findings reveal a previously underappreciated role for the gut microbiome in drug metabolism, and provide a comprehensive framework for characterizing this important class of drug-microbiome interactions

Nutrient regulation of lipochitooligosaccharide recognition in plants via NSP1 and NSP2.

Many plants associate with arbuscular mycorrhizal fungi for nutrient acquisition, while legumes also associate with nitrogen-fixing rhizobial bacteria. Both associations rely on symbiosis signaling and here we show that cereals can perceive lipochitooligosaccharides (LCOs) for activation of symbiosis signaling, surprisingly including Nod factors produced by nitrogen-fixing bacteria. However, legumes show stringent perception of specifically decorated LCOs, that is absent in cereals. LCO perception in plants is activated by nutrient starvation, through transcriptional regulation of Nodulation Signaling Pathway (NSP)1 and NSP2. These transcription factors induce expression of an LCO receptor and act through the control of strigolactone biosynthesis and the karrikin-like receptor DWARF14-LIKE. We conclude that LCO production and perception is coordinately regulated by nutrient starvation to promote engagement with mycorrhizal fungi. Our work has implications for the use of both mycorrhizal and rhizobial associations for sustainable productivity in cereals

Reproducibility of fluorescent expression from engineered biological constructs in E. coli

We present results of the first large-scale interlaboratory study carried out in synthetic biology, as part of the 2014 and 2015 International Genetically Engineered Machine (iGEM) competitions. Participants at 88 institutions around the world measured fluorescence from three engineered constitutive constructs in E. coli. Few participants were able to measure absolute fluorescence, so data was analyzed in terms of ratios. Precision was strongly related to fluorescent strength, ranging from 1.54-fold standard deviation for the ratio between strong promoters to 5.75-fold for the ratio between the strongest and weakest promoter, and while host strain did not affect expression ratios, choice of instrument did. This result shows that high quantitative precision and reproducibility of results is possible, while at the same time indicating areas needing improved laboratory practices.Peer reviewe