In a synthetic biology approach, we have successfully introduced into chloroplasts the entire biochemical pathway for artemisinin, the most effective antimalarial compound which currently is not accessible to many patients in the poorest counties in Africa and Asia. To facilitate the engineering of the whole pathway, we developed a new synthetic biology approach termed COSTREL (COmbinatorial Supertransformation of Transplastomic REcipient Lines’). First, we transferred the genes for the core pathway into the tobacco chloroplast genome. The resulting plants were then combinatorially supertransformed with genes for additional enzymes, including all accessory enzymes known to affect the artemisinin pathway. By screening large populations of COSTREL lines, we isolated plants that produce more than 120 mg artemisinic acid per kg biomass. This work provides a novel and highly efficient strategy for engineering complex biochemical pathways into plants and optimizing the metabolic output. It also demonstrates how a complex pathway in secondary metabolism can be transplanted from a medicinal herb into a high-biomass crop and, moreover, showed that the pathway can be relocated from the cytosol of specialized cells (trichomes of Artimisia annua, the natural source of artemisinin) into leaf chloroplasts (of tobacco). Publication of this work received enormous media attention and was also highlighted in several multidisciplinary journals, including Nature and Science. Additional pathways engineered into chloroplasts include the pathway for the stress-protective compound dhurrin and the high-value ketocarotenoid astaxanthin. Implementation of the pathway for astaxanthin, a non-natural photosynthetic pigment, has revealed that it can completely replace the pigments naturally present in the photosynthetic apparatus, thus representing an important breakthrough in engineering of artificial photosystems with simpler pigment composition. Additionally, a synthetic biology approach has been developed to facilitate the large-scale engineering of signal transduction pathways in plants, leading to plant varieties with improved resistance to abiotic stresses (drought, salt stress, osmotic stress). Finally, a major breakthrough has been achieved with the development of a chloroplast transformation technology for the model plant Arabidopsis.
To develop a technology for mitochondrial genome engineering in plants, large sets of vectors for mitochondrial transformation were constructed. Large-scale mitochondrial transformation and selection experiments with all vectors have been conducted and analysis of candidate lines is underway. In addition, we recently achieved a breakthrough in mitochondrial genome engineering by developing of a new technology for site-directed mutagenesis of the plant mitochondrial genome. Mutant plants have been isolated and comprehensively characterized genetically and biochemically (publication in preparation).
Exploiting our recent discovery that entire genomes can be horizontally transferred between plant species across graft junctions, we aim to create novel (synthetic) crop species as well as new ornamental and medicinal plants. To facilitate horizontal genome transfer between species in the nightshade family (Solanaceae), we have developed transgenic lines with different selectable marker genes for nearly 20 different species. Large-scale grafting experiments and selection for horizontal genome transfer have been performed, and several candidate events have been isolated. In parallel, we elucidated the cellular mechanisms underlying horizontal genome transfer.
