The Arabidopsis P₄-ATPase ALA3 requires a ß-subunit to function in phospholipid translocation and secretory vesicle formation

Lisbeth R. Poulsen¹, Rosa L. López-Marqués¹, Stephen C. McDowell², Juha Okkeri³, Dirk Licht³, Alexander Schulz¹, Thomas Pomorski³,  Jeffrey F. Harper², and Michael G. Palmgren¹

¹Centre for Membrane Pumps in Cells and Disease - PUMPKIN, Danish National Research Foundation, Department of Plant Biology, University of Copenhagen, DK-1871 Frederiksberg C, Denmark

²Biochemistry Department MS200, University of Nevada Reno, NV 89557, USA

³Humboldt-University Berlin, Faculty of Mathematics and Natural Science I, Institute of Biology, 10115 Berlin, Germany

Correspondence to L.R.P.: lrpo@life.ku.dk

Vesicle budding in e.g. endocytosis in eukaryotes is dependent on the activity of lipid translocases (P₄-ATPases) that have been implicated in generating bilayer lipid asymmetry and in inducing membrane curvature, which is a requirement for vesicle formation. We show that Aminophospholipid ATPase3 (ALA3), a member of the P₄-ATPase subfamily in the plant Arabidopsis thaliana, localizes to the Golgi apparatus and that genetic lesions of ALA3 result in impaired growth of roots and shoots. The root growth defect is accompanied by the failure of the root to release border cells of the root cap. Electron micrograph data suggest that functioning and shedding of border cells are dependent on ALA3, as ala3 mutants are devoid of the characteristic proliferation of slime vesicles at the trans-Golgi containing polysaccharides and enzymes for secretion. In yeast complementation experiments, ALA3 gains functionality following interaction with members of a novel family of plant membrane-bound proteins, ALIS1 to ALIS5 for ALA- Interacting Subunit. Based on co-localization studies in planta, interaction and transport studies in yeast, at least one member of this family, ALIS1, is likely to serve as an essential ß-subunit of ALA3. We propose that the ALA3/ALIS1 protein complex forms an important part of the Golgi machinery required for secretory processes during plant development.

We are currently aiming at optimizing expression of the ALA3/ALIS1 complex in yeast and plants in order to characterize its biochemical properties and to obtain structural data, which can help understanding the interaction between the two subunits and the role of the ß-subunit in the mechanism of lipid transport across membranes.

Presenting author: Lisbeth Rosager Poulsen

Category:

This abstract is submitted as category 3.