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GTP-dependent segregation of H-ras from lipid rafts is required for biological activity

Nature Cell Biology volume 3, pages 368–375 (2001)Cite this article

Abstract

Different sites of plasma membrane attachment may underlie functional differences between isoforms of Ras. Here we show that palmitoylation and farnesylation targets H-ras to lipid rafts and caveolae, but that the interaction of H-ras with these membrane subdomains is dynamic. GTP-loading redistributes H-ras from rafts into bulk plasma membrane by a mechanism that requires the adjacent hypervariable region of H-ras. Release of H-ras-GTP from rafts is necessary for efficient activation of Raf. By contrast, K-ras is located outside rafts irrespective of bound nucleotide. Our studies identify a novel protein determinant that is required for H-ras function, and show that the GTP/GDP state of H-ras determines its lateral segregation on the plasma membrane.

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Figure 1: Plasma-membrane localization of GFP–tH and GFP–tK.
Figure 2: Raft association of GFP–tH visualized by crosslinking GPI-anchored PLAP.
Figure 3: Distribution of Ras constructs on flotation gradients.
Figure 4: GTP loading in vivo and in vitro is sufficient to drive H-ras out of lipid rafts.
Figure 5: Plasma-membrane distribution of full-length Ras.
Figure 6: Quantification of Ras distribution on the plasma membrane.
Figure 7: Raf activation in plasma-membrane microdomains.

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Acknowledgements

We thank S. Kornfeld for helpful advice and M. Lindsay for technical assistance. This work was supported by grants from the National Health and Medical Research Council of Australia to R.G.P. and J.F.H. J.F.H. is also supported by the Royal Children's Hospital Foundation of Queensland. The Institute of Molecular Bioscience is a Special Research Centre of the Australian Research Council.

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Authors and Affiliations

  1. Department of Pathology, Laboratory of Experimental Oncology, University of Queensland Medical School, Herston Road, Queensland, 4006, Australia

    Ian A. Prior, Angus Harding, Jun Yan, Judith Sluimer & John F. Hancock

  2. Centre for Microscopy and Microanalysis and Department of Physiology and Pharmacology, Institute for Molecular Bioscience, University of Queensland, Queensland, 4072, Australia

    Robert G. Parton

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Correspondence to John F. Hancock.

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Prior, I., Harding, A., Yan, J. et al. GTP-dependent segregation of H-ras from lipid rafts is required for biological activity. Nat Cell Biol 3, 368–375 (2001). https://doi.org/10.1038/35070050

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