| Rhizobia can form nitrogen-fixing organelles, called symbiosomes, inside the cells of legume root nodules, surrounded by a membrane of host plant origin. However, it is unknown how such plant-originating lipids are transported to the symbiosomes. In this study, we identified AsE246, a lipid transport protein that is specifically expressed in Astragalus sinicus root nodules.It was found that AsE246can bind lipids in vitro. More importantly, AsE246can bind the plant-synthesized membrane lipid digalactosyldiacylglycerol (DGDG) in vivo. In vitro fluorescent assays on the fatty acid1-pyrenedodecanoic acid demonstrated that the recombinant protein AsE246had the capacity to bind lipid. DsRed-AsE246fusion protein was found localized to the plasma membrane in onion epidermal cells. An immuno-fluorescence experiment showed that AsE246was colocalized with symbiosomes from an early stage of symbiosis between A. sinicus and Mesorhizobium huakuii7653R, even in the infection threads. Further immuno-electron microscopy (EM) result confirmed that AsE246was localized on the symbiosome membranes, indicating that AsE246participated in the transportation of related lipids and was involved in the biogenesis of the symbiosome membrane. Overexpression of AsE246resulted in increased numbers of root nodules. Knockdown expression of AsE246via RNAi resulted in the reduction of root nodule numbers, DGDG content and nitrogen fixation activity, and the abnormal development of symbiosomes, including the accumulation of poly-p-hydroxybutyrate, shrinkage in the symbiosome membrane and enlargement of the peribacteroid space. Observation of the symbiotic infection events suggested that AsE246affected nodule organogenesis associated with infection thread formation.Taken together, these results indicate that AsE246contributes to lipids transport to the symbiosome membrane, and this transport is required for effective legume-rhizobium symbiosis. |
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