An UMAMIT-GTR transporter cascade controls glucosinolate seed loading in Arabidopsis.

Many plant species translocate maternally synthesized specialized metabolites to the seed to protect the developing embryo and later the germinating seedling before it initiates its own de novo synthesis. While the transport route into the seed is well established for primary metabolites, no model exists for any class of specialized metabolites that move from maternal source tissue(s) to embryo. Glucosinolate seed loading in Arabidopsis depends on plasma membrane localized exporters (USUALLY MULTIPLE AMINO ACIDS MOVE IN AND OUT TRANSPORTERs, UMAMITs) and importers (GLUCOSINOLATE TRANSPORTERs, GTRs), but the critical barriers in the seed loading process remain unknown. Here we dissect the transport route of glucosinolates from their source in the reproductive organ to the embryo by re-introducing the transporters at specific apoplastic barriers in their respective mutant backgrounds. We find that UMAMIT exporters and GTR importers form a transporter cascade that is both essential and sufficient for moving glucosinolates across at least four plasma membrane barriers along the route. We propose a model in which UMAMITs export glucosinolates out of the biosynthetic cells to the apoplast, from where GTRs import them into the phloem stream, which moves them to the unloading zone in the chalazal seed coat. From here, the UMAMITs export them out of maternal tissue and ultimately, the GTRs import them into the embryo symplasm, where the seed-specific glucosinolate profile is established by enzymatic modifications. Moreover, we propose that methylsulfinylalkyl glucosinolates are the predominant mobile form in seed loading. Elucidation of the seed loading process of glucosinolates identifies barrier-specific targets for transport engineering strategies to eliminate or over-accumulate a specialized metabolite in seeds with minimal interruption of other cellular processes.

Export of defensive glucosinolates is key for their accumulation in seeds.

Plant membrane transporters controlling metabolite distribution contribute key agronomic traits1-6. To eliminate anti-nutritional factors in edible parts of crops, the mutation of importers can block the accumulation of these factors in sink tissues7. However, this often results in a substantially altered distribution pattern within the plant8-12, whereas engineering of exporters may prevent such changes in distribution. In brassicaceous oilseed crops, anti-nutritional glucosinolate defence compounds are translocated to the seeds. However, the molecular targets for export engineering of glucosinolates remain unclear. Here we identify and characterize members of the USUALLY MULTIPLE AMINO ACIDS MOVE IN AND OUT TRANSPORTER (UMAMIT) family-UMAMIT29, UMAMIT30 and UMAMIT31-in Arabidopsis thaliana as glucosinolate exporters with a uniport mechanism. Loss-of-function umamit29 umamit30 umamit31 triple mutants have a very low level of seed glucosinolates, demonstrating a key role for these transporters in translocating glucosinolates into seeds. We propose a model in which the UMAMIT uniporters facilitate glucosinolate efflux from biosynthetic cells along the electrochemical gradient into the apoplast, where the high-affinity H+-coupled glucosinolate importers GLUCOSINOLATE TRANSPORTERS (GTRs) load them into the phloem for translocation to the seeds. Our findings validate the theory that two differently energized transporter types are required for cellular nutrient homeostasis13. The UMAMIT exporters are new molecular targets to improve nutritional value of seeds of brassicaceous oilseed crops without altering the distribution of the defence compounds in the whole plant.