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Annexin 2 Binding to Phosphatidylinositol 4,5-Bisphosphate on Endocytic Vesicles Is Regulated by the Stress Response Pathway
- Source :
- Journal of Biological Chemistry. 279:14157-14164
- Publication Year :
- 2004
- Publisher :
- Elsevier BV, 2004.
-
Abstract
- Annexin 2 is a Ca(2+)-binding protein that has an essential role in actin-dependent macropinosome motility. We show here that macropinosome rocketing can be induced by hyperosmotic shock, either alone or synergistically when combined with phorbol ester or pervanadate. Rocketing was blocked by inhibitors of phosphatidylinositol-3-kinase(s), p38 mitogen-activated protein (MAP) kinase, and calcium, suggesting the involvement of phosphoinositide signaling. Since various phosphoinositides are enriched on inwardly mobile vesicles, we examined whether or not annexin 2 binds to any of this class of phospholipid. In liposome sedimentation assays, we show that recombinant annexin 2 binds to phosphatidylinositol 4,5-bisphosphate (PtdIns-4,5P(2)) but not to other poly- and mono-phosphoinositides. The affinity of annexin 2 for PtdIns-4,5P(2) (K(D) approximately 5 microm) is comparable with those reported for a variety of PtdIns-4,5P(2)-binding proteins and is enhanced in the presence of Ca(2+). Although annexin 1 also bound to PtdIns-4,5P(2), annexin 5 did not, indicating that this is not a generic annexin property. To test whether annexin 2 binds to PtdIns-4,5P(2) in vivo, we microinjected rat basophilic leukemia cells stably expressing annexin 2-green fluorescent protein (GFP) with fluorescently tagged antibodies to PtdIns-4,5P(2). Annexin 2-GFP and anti-PtdIns-4,5P(2) IgG co-localize at sites of pinosome formation, and annexin 2-GFP relocalizes to intracellular membranes in Ptk cells microinjected with Arf6Q67L, which has been shown to stimulate PtdIns-4,5P(2) synthesis on pinosomes through activation of phosphatidylinositol 5 kinase. These results establish a novel phospholipid-binding specificity for annexin 2 consistent with a role in mediating the interaction between the macropinosome surface and the polymerized actin tail.
- Subjects :
- Phosphatidylinositol 4,5-Diphosphate
Biology
Biochemistry
Article
Green fluorescent protein
chemistry.chemical_compound
Osmotic Pressure
Annexin
Cell Line, Tumor
Animals
Phosphatidylinositol
Macropinosome
Transport Vesicles
Molecular Biology
Annexin A2
Cytoskeleton
Kinase
Cell Biology
Molecular biology
Actins
Rats
Cell biology
Endocytic vesicle
Leukemia, Basophilic, Acute
chemistry
Phosphatidylinositol 4,5-bisphosphate
Pinocytosis
Signal Transduction
Subjects
Details
- ISSN :
- 00219258
- Volume :
- 279
- Database :
- OpenAIRE
- Journal :
- Journal of Biological Chemistry
- Accession number :
- edsair.doi.dedup.....a9d105726a60bd6ec58b5ad4208e5afb