Translational regulation offers a powerful biological control axis with the potential to enable programmable control over synthetic mRNAs. Here, we introduce inducible Deaminases Acting on RNA (iDARs): deaminase domains (DDs) with conditional RNA-editing activities. Using a domain-insertion strategy, we designed autoinhibited enzymes that can be converted into active RNA editors in response to triggers based on small molecules (chemiDARs), intracellular antigens (antiDARs), protease cleavage (lysiDAR), and optical excitation (optiDAR). Coupling these domains with novel stop codon containing RNA substrates enabled conditional protein translation or transcript degradation. Mutational tuning of inositol hexaphosphate (IP6)-binding pockets produced tightly regulated deaminases with minimal basal activity, facilitating dose-dependent readthrough translation in response to low-nanomolar drug concentration, with dynamic ranges exceeding 100-fold. By encoding iDARs alongside their substrates, we developed 'self-editing' polycistronic transcripts capable of directing translation of encoded proteins in a trigger-dependent manner following delivery to cells as in vitro transcribed mRNAs. Overall, iDARs provide a generalizable framework for generating controllable deaminases, enabling the design of post-transcriptional circuits that link biochemical sensing to readouts based on de novo translation or mRNA decay.

Selective encapsulation of target enzymes is an increasingly well studied field with a host of potential applications for biotechnology. Natively, many bacteria utilize bacterial microcompartments (BMC) for enzyme encapsulation to enhance catalysis. BMCs are protein shells that enable selective localization of targeted metabolic enzymes and may improve catalytic rates by co-localizing pathway enzymes and/or serve to sequester toxic or volatile intermediates. The microcompartment shell of Haliangium ochraceum (HO) is a notable BMC chassis because of its modularity and versatility; it is easily expressed and assembled outside its native host and can accept a wide array of cargo. Recently, it was demonstrated that assembly of HO BMC shells can be easily achieved in vitro. Following up on our previous work on in vivo assembly of HO-BMCs with triose phosphate isomerase (TPI) as model enzyme cargo, here we have demonstrated the advantages of in vitro assembly (IVA) for targeted enzyme encapsulation. We achieved variable loading of BMC shells with targeted amounts of TPI and demonstrated enhanced thermal stability of encapsulated TPI versus free TPI up to 62°C.

Indole-3-acetic acid (IAA) is a tryptophan-derived gut microbial metabolite with reported anti-inflammatory activities, but the organisms and anaerobic pathways that support robust production remain unclear. Screening 206 human gut bacterial isolates by LC-MS revealed that IAA production is rare: only five strains exceeded the limit of quantitation, and high-capacity production was confined to the acetogens Blautia hydrogenotrophica and Intestinibacter bartlettii. Across growth conditions, IAA was a minor product that rose alongside carbohydrate-sensitive, OFOR-linked catabolism of multiple amino acids, generating abundant branched-chain and aromatic organic acids. In gnotobiotic mice mono-colonized with I. bartlettii, these metabolites were produced in vivo but showed distinct host handling, with branched-chain fatty acids largely extracted between portal and peripheral plasma, whereas aromatic acids and their glycine conjugates appeared in plasma and urine. Genomic analyses and heterologous enzyme assays identified expanded repertoires of 2-oxoacid:ferredoxin oxidoreductases (OFORs) with activities spanning pyruvate/oxaloacetate, branched-chain, and aromatic 2-oxoacids, including indolepyruvate conversion to indoleacetyl-CoA, a putative intermediate en route to IAA. Finally, position-specific 13C tracing showed that CO2 released during amino acid oxidation is reassimilated into acetate via reductive acetogenesis, indicating that gut acetogens can maintain redox balance without fermenting partner strains. Together, these findings show that high IAA output is restricted to select gut acetogens and linked to a broader OFOR-driven anaerobic metabolism that generates additional metabolites that are absorbed by the host.

Artificial biomolecular condensates have emerged as powerful tools to control cellular behaviors. Here we introduce a method to build artificial condensates within living mammalian cells through the design of modular RNA motifs formed by a single, short strand of RNA. These condensates emerge spontaneously, creating RNA-rich compartments that remain separated from their surrounding environment. The RNA sequences include stem-loop domains that fold as the RNA is transcribed, and then condense in the nucleus and cytoplasm through loop-loop interactions. These sequences can be optimized and diversified, enabling the generation of distinct, non-mixing condensate populations and the programmable control of their subcellular localization. The RNA motifs can also be modified to recruit small molecules, proteins, and RNA molecules in a sequence-specific manner to the RNA-rich phase. By introducing additional RNAs that link two distinct types of condensates, we can create droplets with multiple subcompartments, whose organization can be controlled by tuning the stoichiometry of different RNA sequences. These artificial condensates provide a versatile platform for studying and manipulating molecular functions inside living cells.

Volatile organic compounds (VOC) emitted by soil bacteria influence interactions with other soil microbes and with plant roots. While their potential as plant-growth promoters is well recognized, their role in promoting plant resilience to abiotic stress and the underlying molecular mechanisms remains poorly understood. Here, we investigate the role of Pseudomonas VOCs in enhancing plant resilience to drought stress. Arabidopsis thaliana plants were exposed to VOCs emitted by Pseudomonas strains under both control and osmotic-stress conditions. VOC exposure generally enhanced plant growth, and this effect was even more pronounced under both drought and salt stress. Transcriptomic analysis revealed that VOC exposure modulates key stress-responsive pathways, including those related to abscisic acid biosynthesis and signalling, sugar transport, iron uptake, aliphatic glucosinolate biosynthesis, and plant defences. Using Arabidopsis mutants, we identified abscisic acid and aliphatic glucosinolates as important components in mediating the plant response to VOCs. SWEET11/12 sugar transporters and ABA signaling genes were downregulated by VOCs exposure, in order to allow for a positive regulation of lateral root numbers (in case of SWEET genes) and plant growth in general under drought stress. In summary, using metabolomics, transcriptomics and functional analysis, we showed a negative cross-talk between the effects of VOCs on plant growth and glucosinolate production, whereas a positive interaction was observed between the biosynthesis of coumarins and VOCs. Notably, VOCs also improved drought tolerance in soil-grown Brassica oleracea plants. We showed that VOC treatment altered the root-associated microbiome under drought, leading to a community composition more similar to that of well-watered plants. Our results show that Pseudomonas emitted VOCs can promote plant growth under drought conditions, linked to root transcriptional reprogramming and direct or indirect microbiome modulation.

Agriculture is under pressure to provide food for a growing population and the feedstock required to drive the bioeconomy. Methods to breed and genetically modify plants are inadequate to keep pace. When engineering crops, traits are painstakingly introduced into plants one-at-a-time, combine unpredictably, and are continuously expressed. Synthetic biology is changing these paradigms with new genome construction tools, computer aided design (CAD), and artificial intelligence (AI). “Smart plants” contain circuits that respond to environmental change, alter morphology, or respond to threats. Further, the plant and associated microbes (fungi, bacteria, archaea) are now being viewed by genetic engineers as a holistic system. Historically, plant health has been enhanced by many natural and laboratory-evolved soil microbes marketed to enhance growth or provide nutrients, or pest/stress resistance. Synthetic biology has expanded the number of species that can be engineered, increased the complexity of engineered functions, controlled environmental release, and can assemble stable consortia. New CAD tools will manage genetic engineering projects spanning multiple plant genomes (nucleus, chloroplast, mitochondrion) and the thousands of genomes of associated bacteria/fungi. This review covers advanced genetic engineering techniques to drive the next agricultural revolution, as well as push plant engineering into new realms for manufacturing, infrastructure, sensing, and remediation.

Fold switching, where a protein region interconverts between entirely distinct three-dimensional structures, is emerging as vital for certain protein functions. Here, we report a remarkable example in the F7 pyocin, a phage tail-like bactericidal nanomachine. Cryogenic electron microscopy and tomography reveal that a 163-residue segment of the central tail fiber undergoes a dramatic transition—from a trimeric α-helical coiled-coil to a triangular β-prism—upon binding to the bacterial cell surface. This massive fold switch remodels the tail tip, ejects the internal tape measure protein, and drives membrane puncture. Site-directed mutations that selectively destabilize the β-prism conformation completely abolish bactericidal activity without impairing particle assembly, implying that the energy released during this transition powers penetration. AlphaFold-based analyses further predict similar large-scale coiled-coil to β-prism switches in diverse non-contractile phage tails. This discovery reveals a sophisticated, ATP-independent strategy for microbial warfare and opens exciting possibilities for engineering next-generation bacteriocins to combat multidrug-resistant pathogens.

Microbial metabolites play a critical role in regulating ecosystems, including the human body and its microbiota. However, understanding the physiologically relevant role of these molecules, especially through liquid chromatography tandem mass spectrometry (LC-MS/MS)-based untargeted metabolomics, poses significant challenges and often requires manual parsing of a large amount of literature, databases, and webpages. To address this gap, we established the Collaborative Microbial Metabolite Center knowledgebase (CMMC-KB), a platform that fosters collaborative efforts within the scientific community to curate knowledge about microbial metabolites. The CMMC-KB aims to collect comprehensive information about microbial molecules originating from microbial biosynthesis, drug metabolism, exposure-related molecules, food, host-derived molecules, and, whenever available, their known activities. Molecules from other sources, including host-produced, dietary, and pharmaceutical compounds, are also included. By enabling direct integration of this knowledgebase with downstream analytical tools, including molecular networking, we can deepen insights into microbiota and their metabolites, ultimately advancing our understanding of microbial ecosystems.

Cell lysis to release intracellular targets is a vital step in many bacterial sensing platforms and is often achieved through chemical or physical approaches. However, these conventional methods can have certain limitations such as cost, required equipment, safety, or risk of target damage. Cell lysis induced by bacteriophages, which are bacteria-infecting viruses, has some notable advantages, including safety and the self-amplifying properties of phage. Bacteriophages also induce species-selective infection, enabling the targeted lysis of a specific bacterial species in mixed cultures. Despite this, bacteriophage-induced lysis has to date been relatively poorly adopted in the bacterial biosensing field. In this Perspective, we outline the potential benefits of bacteriophage lysis in biosensors, while also exploring the reasons that it has not been more widely adopted. We also identify future research directions to facilitate increased incorporation of bacteriophages into bacterial detection platforms, including improving the characterization, availability, and stability of phage strains.