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Sterol-dependent endocytosis mediates post-cytokinetic acquisition of PIN2 auxin efflux carrier polarity


The polarization of yeast and animal cells relies on membrane sterols for polar targeting of proteins to the plasma membrane, their polar endocytic recycling and restricted lateral diffusion1,2,3,4. However, little is known about sterol function in plant-cell polarity5. Directional root growth along the gravity vector requires polar transport of the plant hormone auxin. In Arabidopsis, asymmetric plasma membrane localization of the PIN–FORMED2 (PIN2) auxin transporter directs root gravitropism6,7,8,9,10. Although the composition of membrane sterols influences gravitropism and localization of two other PIN proteins11, it remains unknown how sterols contribute mechanistically to PIN polarity. Here, we show that correct membrane sterol composition is essential for the acquisition of PIN2 polarity. Polar PIN2 localization is defective in the sterol-biosynthesis mutant cyclopropylsterol isomerase1-1 (cpi1-1) which displays altered sterol composition, PIN2 endocytosis, and root gravitropism. At the end of cytokinesis, PIN2 localizes initially to both newly formed membranes but subsequently disappears from one. By contrast, PIN2 frequently remains at both daughter membranes in endocytosis-defective cpi1-1 cells. Hence, sterol composition affects post-cytokinetic acquisition of PIN2 polarity by endocytosis, suggesting a mechanism for sterol action on establishment of asymmetric protein localization.

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Figure 1: Isolation and characterization of the cpi1-1 mutant.
Figure 2: CPI1 and PIN2 protein localization in Arabidopsis root tips.
Figure 3: Myc–CPI1 immunogold localization to ER including cell plate-associated ER membranes.
Figure 4: PIN2 polarity acquisition after cytokinesis requires CPI1 function.
Figure 5: PIN2 endocytosis requires CPI1 function.

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We gratefully acknowledge M. Bennett, D. Ehrhardt, T. Guilfoyle, J. Haseloff, R. Heidstra, R. Hellens, I. Moore, P. Mullineaux, G. Jürgens, B. Scheres, R. Swarup and J. Xu for sharing published research materials used in this study. We also acknowledge the Nottingham Arabidopsis Stock Centre for distributing mutant lines including SALK T-DNA insertion mutants, provided by J. Alonso and J. Ecker, and the John Innes Centre for EXOTIC Gene Trap lines. We thank I.-B. Carlsson and K. Lundgren for technical assistance with sterol measurements and K. Schumacher for providing seedlings for electron microscopy. We thank L. Bako, R. Bhalerao, U. Fischer, E. Johnson, A. Marchant, and G. Samuelsson for discussions and comments on the manuscript. This work was supported by a grant from the Swedish Research Council for Environment, Agricultural Sciences and Spatial Planning (Formas) to M.G., in part by a postdoctoral stipend from the Carl Tryggers Foundation to Y.B., an EU Marie-Curie International Incoming Postdoctoral Fellowship to Y.I., and the Swedish Foundation for Strategic Research (SSF).

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Correspondence to Markus Grebe.

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Supplementary figures S1, S2, S3, S4, S5, Supplementary table S1 and Supplementary Methods (PDF 1528 kb)

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Men, S., Boutté, Y., Ikeda, Y. et al. Sterol-dependent endocytosis mediates post-cytokinetic acquisition of PIN2 auxin efflux carrier polarity. Nat Cell Biol 10, 237–244 (2008).

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