title: The vegetative vacuole proteome of Arabidopsis thaliana reveals predicted and unexpected proteins.
abstract: Vacuoles play central roles in plant growth, development, and stress responses. To better understand vacuole function and biogenesis we have characterized the vegetative vacuolar proteome from Arabidopsis thaliana. Vacuoles were isolated from protoplasts derived from rosette leaf tissue. Total purified vacuolar proteins were then subjected either to multidimensional liquid chromatography/tandem mass spectrometry or to one-dimensional SDS-PAGE coupled with nano-liquid chromatography/tandem mass spectrometry (nano-LC MS/MS). To ensure maximum coverage of the proteome, a tonoplast-enriched fraction was also analyzed separately by one-dimensional SDS-PAGE followed by nano-LC MS/MS. Cumulatively, 402 proteins were identified. The sensitivity of our analyses is indicated by the high coverage of membrane proteins. Eleven of the twelve known vacuolar-ATPase subunits were identified. Here, we present evidence of four tonoplast-localized soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs), representing each of the four groups of SNARE proteins necessary for membrane fusion. In addition, potential cargo of the N- and C-terminal propeptide sorting pathways, association of the vacuole with the cytoskeleton, and the vacuolar localization of 89 proteins of unknown function are identified. A detailed analysis of these proteins and their roles in vacuole function and biogenesis is presented.
authors.affiliation: Center for Plant Cell Biology, Botany and Plant Sciences, University of California, Riverside, California 92521, USA.
authors.forename: Clay
authors.initials: C
authors.lastname: Carter
authors.forename: Songqin
authors.initials: S
authors.lastname: Pan
authors.forename: Jan
authors.initials: J
authors.lastname: Zouhar
authors.forename: Emily L
authors.initials: EL
authors.lastname: Avila
authors.forename: Thomas
authors.initials: T
authors.lastname: Girke
authors.forename: Natasha V
authors.initials: NV
authors.lastname: Raikhel
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000737:chemistry / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D003599:Cytoskeleton / Q000737:chemistry / Q000378:metabolism; D008561:Membrane Fusion / Q000502:physiology; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D010455:Peptides / Q000378:metabolism; D018515:Plant Leaves / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary / Q000502:physiology; D021122:Protein Subunits / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D021381:Protein Transport / Q000502:physiology; D020543:Proteome / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D011523:Protoplasts / Q000737:chemistry / Q000378:metabolism; D050600:SNARE Proteins; D015398:Signal Transduction / Q000502:physiology; D025262:Vacuolar Proton-Translocating ATPases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D014617:Vacuoles / Q000737:chemistry / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D029681:Arabidopsis Proteins; D008565:Membrane Proteins; D010455:Peptides; D021122:Protein Subunits; D020543:Proteome; D050600:SNARE Proteins; D033921:Vesicular Transport Proteins; D025262:Vacuolar Proton-Translocating ATPases