Tracking lipid droplet dynamics using BODIPY C12 in Phaeodactylum tricornutum

Oleaginous microalgae are central to fundamental research on microalgae and biotechnology for single-cell oil production. Although many tools to manipulate physiological and molecular mechanisms have been developed for microalgae, cellular biology tools that enable visualization of lipid droplet (LD) dynamics in vivo are lacking. In this study, we demonstrate the use of the fluorescent fatty acid conjugate BODIPY C12 (BP C12) to study LD dynamics in two ecotypes of the model diatom Phaeodactylum tricornutum Pt1 and Pt4, as well as fatty acid trafficking in vivo, through real-time visualization of LD formation and breakdown by epifluorescence microscopy and FACS. Cells supplied with BP C12 under nutrient-replete conditions exhibited incorporation of BP C12 signal inside the cells in tiny vesicles. Under nitrogen starvation, the BP C12 signal was localized to the preformed LDs. During recovery from nitrogen starvation, we observed the breakdown of the LDs harbouring the BP C12 signal. Thin-layer chromatography showed that most BP C12 remained unesterified, but the signal was found in both polar and neutral lipids, suggesting its suitability for lipid-metabolic studies in P. tricornutum. Use of BP C12 also elucidated substrate utilization in the two ecotypes. These findings pave the way for the use of BP C12 in various assays with P. tricornutum.