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Phosphatidylinositol-3-OH kinases are Rab5 effectors

Nature Cell Biology volume 1, pages 249–252 (1999)Cite this article

Phosphatidylinositol-3-OH kinases (PI(3)Ks) constitute a family of lipid-modifying enzymes that are involved in signal transduction, cytoskeletal organization and membrane transport1. Three different classes of PI(3)K have been described, which differ in their specificities for phosphatidylinositol, phosphatidylinositol-4-phosphate and phosphatidylinositol-4,5-bisphosphate substrates, as well as in their regulation. PI(3)Ks are needed for receptor trafficking in the endocytic pathway in mammalian cells and for transport from the Golgi to the vacuole in yeast (reviewed in ref. 1). The docking and fusion of early endosomes, which is regulated by the small GTPase Rab5 (ref. 2), requires PI(3)K activity3. In addition, PI(3)Ks regulate the recruitment to membranes of EEA1, an effector of the small GTPase Rab5 and a core component of the docking and fusion machinery4,5 that binds phosphatidylinositol-3-phosphate (PtdIns(3)P) through a FYVE-finger motif6. We have recently identified over 20 cytosolic proteins that interact with the active form of Rab5 (ref. 5). Here we report the identification of two distinct PI(3)Ks, hVPS34 and p85α–p110β, among these cytosolic proteins. We suggest a new mechanism by which Rab5 can modify its membrane environment by coupling the local production of phosphoinositides to the selective recruitment of Rab5 effector proteins.

Given the presence of a class-I-PI(3)K regulatory subunit in the Rab5-GTPγS eluate, we would expect the catalytic subunit to be either p110α or p110β (ref. 1). By using isoform-specific antibodies we established that p110β, but not p110α, was present in the eluate (Fig. 1c). We detected no interaction of p110β with fusion proteins consisting of glutathione-S-transferase (GST) fused to Rab4-GTPγS or Rab11-GTPγS. We also tested a class-II PI(3)K (PI3K-C2α), but failed to detect binding of this enzyme to Rab5-GTPγS or Rab5-GDP. These results indicate that Rab5 specifically and GTP-dependently interacts with the class-I PI(3)K p85α–p110β.

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Figure 1: The PI(3)Ks p85α–p110β and p150–hVPS34 bind specifically to a Rab5-GTPγS affinity column.
Figure 2: Role of PI(3)Ks in endosome fusion and recruitment of EEA1.

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Acknowledgements

We thank J. Downward for p85α complementary DNA; U. Maier for the p110β construct; R. Lippe for the complex consisting of Rab5 and guanine-nucleotide-dissociation factor; and A. Giner for technical assistance. S.C. and M.M. are recipients of Max Planck and Human Frontier Science Program fellowships, respectively. This work was supported by the Max Planck Gesellschaft and by grants from the Human Frontier Science Program (RG-432/96), EU TMR (ERB-CT96-0020) and Biomed (BMH4-97-2410) (M.Z.).

Correspondence and requests for materials should be addressed to M.Z.

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Authors and Affiliations

  1. Max Planck Institute for Molecular Cell Biology and Genetics, Pfotenhauerstrasse, Dresden, 01307, Germany

    Savvas Christoforidis, Marta Miaczynska, Liyun Zhao & Marino Zerial

  2. European Molecular Biology Laboratory (EMBL), Meyerhofstrasse 1, Heidelberg, 69117, Germany

    Savvas Christoforidis, Marta Miaczynska, Keith Ashman, Matthias Wilm & Marino Zerial

  3. Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York, 10461, USA

    Shu-Chin Yip & Jonathan M. Backer

  4. Ludwig Institute of Cancer Research, University College London, Riding House Street, London, W1P 8BT, UK

    Michael D. Waterfield

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Christoforidis, S., Miaczynska, M., Ashman, K. et al. Phosphatidylinositol-3-OH kinases are Rab5 effectors. Nat Cell Biol 1, 249–252 (1999). https://doi.org/10.1038/12075

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