Research

Plant synthetic biology generally involves the addition of one or more foreign elements to the plants genome to re-program some aspect of its biology. Modifying the genome of a plant is a very time consuming and costly process, taking several years for some crop plants. To overcome this challenge we are designing gene delivery tools based on plant viruses that can deliver genetic cargos systemically, thereby sidestepping transformation and turning years long experiments into a matter of weeks. We plan to build on our early success with ViN vectors by extending it to a range of other plant species, such as rice, sorghum, switchgrass, and hemp. We are also using this platform to rapidly decode the catalytic properties of enzymes of unknown function.

Engineering synthetic signaling systems - Using high-throughput screens and machine learning to design control systems

Molecular tools that can precisely modulate gene expression provide unique opportunities to both study the relationships between genotype and phenotype, as well as correct the pathologies caused by dysregulated gene expression. However, restricting the activity of these tools to the appropriate times or cell types remains a major hurdle to their effectiveness at interrogating biology and their safety in therapeutic settings. We are working to elucidate the design principles for synthetic signaling systems that enable precision control of gene expression and, in doing so, to create fundamental insights into the mechanistic basis of natural signal transduction. Our work spans 3 methods of modulating expression and in each case seeks to overcome a major engineering challenge by generating novel fundamental insights into the transduction mechanism through a combination of high-throughput screening and machine learning. The first are Cas9-based synthetic transcription factors, which enable targeted changes to the transcription of a gene. We plan to restrict their activity via fusion to nuclear receptors that will make their nuclear localization, and hence regulation, conditional on a chemical inducer. We aim to elucidate how the structure of nuclear receptors encodes their nuclear trafficking kinetics and dynamic range, and then use these insights to design controls systems that can rapidly implement strong regulation in response to a non-toxic chemical cue. The second are ribozyme-based tools that regulate expression at the RNA level through splicing or transcleavage. We plan to make their activity contingent on the presence of either native mRNAs, through template dependent splicing, or chemicals, using aptazymes. We aim to understand how changes to the sequence, and resulting structure, of these RNA devices alter their capacity to transduce their triggers into catalysis. The resulting insights will be used to identify a combination of mutations that can overcome the low catalytic efficiencies often associated with these tools. The third are chemicals that activate human G-protein coupled receptors (GPCRs) to modulate expression of the genes they regulate. We plan to identify plant metabolites that act as selective agonists by developing a high-throughput screen that enables massively parallel characterization of GPCR-ligand interactions. We aim to elucidate the design principles for functional expression of human GPCRs in yeast and use the resulting biosensors to reveal the ligand features necessary for selective activation of GPCRs. 

We are recuiting postdocs / staff scientists for this work.

Novel herbicides - Using ribozymes to design safe and effective viral herbicides

Weeds, especially Amaranthus palmeri (Palmer amaranth), represent a major biosecurity threat to agriculture in the United States currently amounting to $33 billion in losses annually. Climate change and herbicide resistant varieties are leading to the spread of this pest to new US states and threatening international grain exports. To address the lack of chemical herbicides capable of addressing this threat we propose to design safe and effective viral herbicides. The environmental stability, self-amplification, systemic spread, and plant growth suppression inherent to plant viruses means that they play an important role in regulating population density in most natural ecosystems. However, their promiscuous capacity to infect plants has limited their application for weed management.  To enable targeted infection of weeds we are designing a novel form of biocontainment that relies on trans-cleaving ribozymes that will restrict the virus’s infectivity to just A. palmeri. We are also ribozyme cargos to load onto these viruses to knock down the expression of essential and herbicide resistance genes in A. palmeri in a sequence specific manner. Finally we are also identifying mutations that enhance systemic infection of A. palmeri. 

Molecular Bonsai - Understanding and engineering dwarfing phenotypes

The capacity to predictably engineer plant organ size could dramatically improve crop productivity and resilience to climate change. A historical example of this is the semi-dwarfed plant varieties that drove the Green-revolution, which arose from mutations in the signaling pathway of the plant hormone gibberellin. We are also using synthetic transcription factors to make targeted changes to the expression of gibberellin signaling genes to elucidate the design rules to engineer semi-dwarfing. For example we are interested in determining the optimal regulatory architecture to generate robust dwarfing and exploring how this varies across tissues and plants. We are also using the resulting lines to generate predicitve models that relate genotype to organ size to enable future in silico design of plant body plans. Finally, gibberellin signaling impacts more than just organ size, we are also develop synthetic transcription factors whose activity can be spatiotemporally restricted to overcome negative pleiotropies. 

Enhancing carbon fixation in plants - Using addative recalcitrance to stablize photosynthetically fixed carbon

We are developing genetic circuits to flip the plants metabolism from turning the carbon that it fixes via photosynthesis from D-sugars that microbes can easily break down, to L-sugars that they cannot. This resistance to breakdown is because L-sugars, which are the mirror image of D-sugars in terms of shape, are very rarely produced in nature and so are not recognized by active site of microbial enzymes, as these microbes have not evolved to eat them. This will allow carbon, once fixed as an L-sugar, to stay fixed and not be released back into the air via microbial decomposition once the plant dies. Control systems are needed to make sure the plant can initialy pursue its natural metabolism to grow enough leaves, roots, and stems to ensure efficient carbon fixation by keep the L-sugar biosynthesis genes tightly off. These control systems are designed to, upon activation via spraying with an agro chemical, strongly activate the expression of the L-sugar biosynthesis genes and repress the genes that channel carbon into D-sugars. We are also exploring other mechanisms of addative recalitrance such as mineralization, the production of L-sugar polymers, and the generation of protein crystals. 

Synthetic lichen - Exploring the morphological basis of symbiosis

Symbiosis between microbes plays a major role in maintaining the consortia that colonize the world’s biomes. Lichens are a product of microbial symbiosis that incorporates aspects of both metabolism and morphology, making them a great model to study these processes. The metabolic aspect of this relationship is thought to occur via a mutualistic exchange of carbon and sometimes nitrogen that is fixed by the photobiont (photosynthesizing partner), often cyanobacteria or algae, for minerals and/or micronutrients from the mycobiont (the filamentous fungal partner). In addition to metabolism, there is also a significant morphological aspect to the lichen symbiosis where in the mycobiont creates a compartment within its mycelial structure for the photobiont to grow. This morphology has been thought to contribute to the resilience lichen by enabling a level of desiccation and UV tolerance the free-living photobiont might not possess. The balance between the metabolic contributions of the photobiont and the morphological contributions of the mycobiont might explain the ecological stability and success of lichen symbiosis. However, the morphological contribution of the mycobiont to lichen symbiosis remains largely unexplored. This is in part because it is very challenging to study these relationships in natural lichens due their slow growth, heterogeneity in non-primary members, and the lack of tools to engineer the members of natural lichenaceous consortia. We are using synthetic biology to create a synthetic lichen from free-living filamentous fungi and cyanobacteria. To do this we are building a cell-cell signaling system based on the plant growth hormone auxin that lets cyanobacteria signal to fungi, and installing genetic circuits in Aspergillus fungi to drive the formation of a lichen-like thallus in response to this signal.  

Lichen-inspired living materials - Creating sustainable mycomaterials with tunable material properties

Engineered living materials (ELMs) have unique properties thanks to their embedding of living cells within a substrate such as self-assembly, sensing, and repair but have been challenging to scale to the centimeter and meter scales. In contrast, materials based on the mycelium of filamentous fungi (mycomaterials) have been successfully deployed for meter scale construction but currently lack the tunable material properties and bioactivity that are accessible to ELM. Additionally, all current approaches require carbon inputs, which reduce their sustainability. We are developing lichen-inspired mycomaterials to address these challenges. We achieve zero input growth of the fungi (Aspergillus niger) by establishing a co-culture with a cyanobacteria (Synechococcus elongatus) that is engineered to secrete up to 80% of its fixed carbon as sucrose. To tune material properties within the mycelium we have engineered a version of the Silicatein α enzyme from sea sponges (Tethya aurantia and Latrunculia oparinae) to be displayed on the surface of hyphae. We show that this enables mineralization of silica oxide on the hyphal surface and changes the strength and elasticity of the mycelium. To facilitate tuning of these systems we are developing synthetic transcription factors that are responsive to the phytohormone auxin to titrate expression of the Silicatein. Finally, we are also using these tools to explore whether changes in the morphology of the mycelium, specifically the degree of hyphal fusion, are able to alter material properties. Together, this work will enable the creation of sustainable and tunable mycomaterials.

Decoding the molecular basis of flavor - Engineering the flavor and fragrance of tomato

The flavors and fragrances of crops like tomato can bring us joy, alter our moods, and are a critical part of our cultural identities. Improving this trait is an important way to encourage people to adopt plant-based diets and thereby reduce the impact of food production on climate change. Flavor and fragrance of tomato plants are a product of primary and secondary metabolites synthesized by a complex knot of biosynthetic pathways. These molecules are bound by receptors, called GPCRs, in the mouth and nose, whose combinatorial activation is thought to lead to the perception of specific tastes and smells. We are using high throughput screens to identify which of these GPCRs are activated by tomato compounds. We are then developing yeast based biosensors expressing these GPCRs as part of a synthetic signaling cascade to study what sorts of genome edits will lead to improved flavor. We are also working to create tomato lines that express these synthetic signaling cascades linked to pigment biosynthesis, to create plants able to perceive how much of a flavor or fragrance they are producing and report it by changing color, as a tool for engineering flavor and fragrance. In the future we hope to expand the use of this strategy to engineer both the flavor of other crops and other aspects of plant secondary metabolism. 

A talk that outlines how our lab thinks about engineering multicellular organisms.