The Lipotubuloids of Ornithogalum umbellatum L. Contain Hyperstable Microtubules

The epidermal cells of bracts, petals and sepals of Ornithogalum umbellatum L. (Star-of-Bethlehem, Asparagaceae) contain lipotubuloids, complex aggregates of lipid droplets (LDs) enmeshed by bundles of microtubules (MTs). We investigated lipotubuloid organization and stability through the transient expression of GFP fusion proteins targeted to different subcellular structures and with immunofluorescence and transmission electron microscopy (TEM). Live cell imaging confirmed that lipotubuloids contain LDs, organelles including endomembranes, mitochondria and peroxisomes, a tonoplast-defined vacuole, and that they move through actin microfilament-based streaming. Intriguingly, the different microscopy modes used showed different patterns of MT organization in the lipotubuloid. While MT sheets and bundles were visible by TEM, few MTs were seen with fusion proteins and immunofluorescence. Oryzalin-based MT depolymerization experiments suggest a possible resolution for this paradox: TEM showed that lipotubuloid MTs resisted depolymerization, even after 20 h in oryzalin, while MT polymerization was visible in lipotubuloids with fusion proteins during oryzalin wash-out. These results suggest that the Ornithogalum lipotubuloids contain hyperstable MTs, possibly formed with microtubule-associated proteins (MAPs) that normally occlude fusion protein and antibody binding sites.