title: A complex and mobile structure forms a distinct subregion within the continuous vacuolar membrane in young cotyledons of Arabidopsis.
abstract: The plant vacuole is a multifunctional organelle which is essential for growth and development. To visualize the dynamics of plant vacuolar membranes, gamma-TIP (tonoplast intrinsic protein) was fused to GFP and expressed in Arabidopsis thaliana. The marker molecule was targeted to the vacuolar membranes in most tissues, as expected. In rapidly expanding cells, some additional spherical structures were often observed within the lumen of vacuoles, which emitted strong fluorescence. To confirm their normal presence, we examined wild-type Arabidopsis cotyledons by transmission electron microscopy. The metal-contact rapid-freezing method revealed that the vacuolar lumen of epidermal cells contained many cytoplasmic projections, which often formed spherical structures (1-3 microm diameter) consisting of double membranes. Thus we concluded that these structures are authentic and named them 'bulbs'. Three-dimensional reconstruction from serial electron microscopic images demonstrates that bulbs are very intricately folded, but are continuous with the limiting vacuolar membrane. The fluorescence intensity of bulbs is about threefold higher than that of vacuolar membrane. GFP-AtRab75c, another marker of the vacuole, did not give fluorescent signals of bulbs in transgenic plants, but the existence of bulbs was still confirmed by electron microscopy. These results suggest that bulbs define a subregion in the continuous vacuolar membrane, where some proteins are concentrated and others segregated.
authors.affiliation: Molecular Membrane Biology Laboratory, RIKEN, Wako, Saitama 351-0198, Japan.
authors.forename: Chieko
authors.initials: C
authors.lastname: Saito
authors.forename: Takashi
authors.initials: T
authors.lastname: Ueda
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Abe
authors.forename: Yoh
authors.initials: Y
authors.lastname: Wada
authors.forename: Tsuneyoshi
authors.initials: T
authors.lastname: Kuroiwa
authors.forename: Akiko
authors.initials: A
authors.lastname: Hisada
authors.forename: Masaki
authors.initials: M
authors.lastname: Furuya
authors.forename: Akihiko
authors.initials: A
authors.lastname: Nakano
mesh_terms: D017360:Arabidopsis / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D018548:Cotyledon / Q000254:growth & development / Q000502:physiology*; D049452:Green Fluorescent Proteins; D007425:Intracellular Membranes / Q000502:physiology* / Q000648:ultrastructure; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D019441:Plant Epidermis / Q000254:growth & development / Q000502:physiology* / Q000648:ultrastructure; D010940:Plant Proteins / Q000378:metabolism; D030821:Plants, Genetically Modified; D014617:Vacuoles / Q000502:physiology* / Q000648:ultrastructure
chemical_list: D008164:Luminescent Proteins; D008565:Membrane Proteins; D010940:Plant Proteins; C077491:tonoplast intrinsic protein, plant; D049452:Green Fluorescent Proteins