Interactome of the plant-specific ESCRT-III component AtVPS2.2 in Arabidopsis thaliana.
The endosomal sorting complexes required for transport (ESCRT) guides transmembrane proteins to domains that bud away from the cytoplasm. The ESCRT machinery consists of four complexes. ESCRT complexes 0-II are important for cargo recognition and concentration via ubiquitin binding. Most of the membrane bending function is mediated by the large multimeric ESCRT-III complex and associated proteins. Here we present the first in vivo proteome analysis of a member of the ESCRT-III complex which is unique to the plant kingdom. We show with LC-MS/MS, yeast-two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) that coimmunoprecipitated proteins from Arabidopsis thaliana roots expressing a functional GFP-tagged VACUOLAR PROTEIN SORTING 2.2 (AtVPS2.2) protein are members of the ESCRT-III complex and associated proteins. Therefore we propose that at least in plants the large ESCRT-III membrane scaffolding complex consists of a mixture of SNF7, VPS2 and the associated VPS46 and VPS60 proteins. Apart from transmembrane proteins, numerous membrane-associated but also nuclear and extracellular proteins have been identified, indicating that AtVPS2.2 might be involved in processes beyond the classical ESCRT role. This study is the first in vivo proteome analysis with a tagged ESCRT-III component demonstrating the feasibility of this approach and provides numerous starting points for the investigation of the biological process in which AtVPS2.2 is involved.
Department of Applied Genetics and Cell Biology, BOKU-University of Natural Resources and Life Sciences, Muthgasse 18, A-1190 Vienna, Austria.
Verena
V
Ibl
Edina
E
Csaszar
Nicole
N
Schlager
Susanne
S
Neubert
Christoph
C
Spitzer
Marie-Theres
MT
Hauser
D017360:Arabidopsis / Q000166:cytology / Q000254:growth & development / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000378:metabolism*; D021961:Cell Membrane Structures / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D034281:Dynamins / Q000378:metabolism; D056827:Endosomal Sorting Complexes Required for Transport / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D047468:Immunoprecipitation; D010739:Phospholipase D / Q000378:metabolism; D018517:Plant Roots / Q000166:cytology / Q000254:growth & development / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D020543:Proteome / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D033921:Vesicular Transport Proteins / Q000378:metabolism*
D029681:Arabidopsis Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D020543:Proteome; D011993:Recombinant Fusion Proteins; C571713:VPS2.2 protein, Arabidopsis; D033921:Vesicular Transport Proteins; D049452:Green Fluorescent Proteins; D010739:Phospholipase D; D034281:Dynamins