The large GTPase Sey1/atlastin mediates lipid droplet- and FadL-dependent intracellular fatty acid metabolism of Legionella pneumophila .

The amoeba-resistant bacterium Legionella pneumophila causes Legionnaires' disease and employs a type IV secretion system (T4SS) to replicate in the unique, ER-associated Legionella -containing vacuole (LCV). The large fusion GTPase Sey1/atlastin is implicated in ER dynamics, ER-derived lipid droplet (LD) formation, and LCV maturation. Here we employ cryo-electron tomography, confocal microscopy, proteomics, and isotopologue profiling to analyze LCV-LDs interactions in the genetically tractable amoeba Dictyostelium discoideum . Dually fluorescence-labeled D. discoideum producing LCV and LD markers revealed that Sey1 as well as the L. pneumophila T4SS and the Ran GTPase activator LegG1 promote LCV-LDs interactions. In vitro reconstitution using purified LCVs and LDs from parental or D sey1 mutant D. discoideum indicated that Sey1 and GTP promote this process. Sey1 and the L. pneumophila fatty acid transporter FadL are implicated in palmitate catabolism and palmitate-dependent intracellular growth. Taken together, our results reveal that Sey1 and LegG1 mediate LD- and FadL-dependent fatty acid metabolism of intracellular L. pneumophila .