In vivo analysis of 3-phosphoinositide dynamics during Dictyostelium phagocytosis and chemotaxis

Phagocytosis and chemotaxis are receptor-mediated processes that require extensive rearrangements of the actin cytoskeleton, and are controlled by lipid second messengers such as phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P3] and phosphatidylinositol 3,4-bisphosphate [PtdIns(3,4)P2]. We used a panel of pleckstrin homology (PH) domains with distinct binding specificities for PtdIns(3,4,5)P3 and PtdIns(3,4)P2 to study the spatiotemporal dynamics of these phosphoinositides in vivo. During phagocytosis and macropinocytosis PtdIns(3,4,5)P3 levels transiently increased at sites of engulfment, followed by a rapid PtdIns(3,4)P2 production round the phagosome/macropinosome upon its internalisation, suggesting that PtdIns(3,4,5)P3 is degraded to PtdIns(3,4)P2. PTEN null mutants, which are defective in phagocytosis, showed normal rates of PtdIns(3,4,5)P3degradation, but unexpectedly an accelerated PtdIns(3,4)P2 degradation. During chemotaxis to cAMP only PtdIns(3,4,5)P3 was formed in the plasma membrane, and no PtdIns(3,4)P2 was detectable, showing that all PtdIns(3,4,5)P3 was degraded by PTEN to PtdIns(4,5)P2. Furthermore, we showed that different PtdIns(3,4,5)P3 binding PH domains gave distinct spatial and temporal readouts of the same underlying PtdIns(3,4,5)P3 signal, enabling distinct biological responses to one signal.

[1]  G. Gerisch,et al.  Coronin involved in phagocytosis: Dynamics of particle-induced relocalization visualized by a green fluorescent protein tag , 1995, Cell.

[2]  K. Zhou,et al.  A phosphatidylinositol (PI) kinase gene family in Dictyostelium discoideum: biological roles of putative mammalian p110 and yeast Vps34p PI 3-kinase homologs during growth and development , 1995, Molecular and cellular biology.

[3]  J. Bush,et al.  Involvement of the vacuolar proton-translocating ATPase in multiple steps of the endo-lysosomal system and in the contractile vacuole system of Dictyostelium discoideum. , 1996, Journal of cell science.

[4]  J. Swanson,et al.  A role for phosphoinositide 3-kinase in the completion of macropinocytosis and phagocytosis by macrophages , 1996, The Journal of cell biology.

[5]  J. Bush,et al.  Inactivation of Two Dictyostelium discoideum Genes, DdPIK1 and DdPIK2, Encoding Proteins Related to Mammalian Phosphatidylinositide 3-kinases, Results in Defects in Endocytosis, Lysosome to Postlysosome Transport, and Actin Cytoskeleton Organization , 1997, The Journal of cell biology.

[6]  M. Andjelkovic,et al.  High Affinity Binding of Inositol Phosphates and Phosphoinositides to the Pleckstrin Homology Domain of RAC/Protein Kinase B and Their Influence on Kinase Activity* , 1997, The Journal of Biological Chemistry.

[7]  J. Holik,et al.  Signaling by Phosphoinositide-3,4,5-Trisphosphate Through Proteins Containing Pleckstrin and Sec7 Homology Domains , 1997, Science.

[8]  D. Murphy,et al.  G Protein Signaling Events Are Activated at the Leading Edge of Chemotactic Cells , 1998, Cell.

[9]  S. Bozzaro,et al.  G Protein β Subunit–null Mutants Are Impaired in Phagocytosis and Chemotaxis Due to Inappropriate Regulation of the Actin Cytoskeleton , 1998, The Journal of cell biology.

[10]  J. Bush,et al.  Overexpression of a novel rho family GTPase, RacC, induces unusual actin-based structures and positively affects phagocytosis in Dictyostelium discoideum. , 1998, Molecular biology of the cell.

[11]  K. Zhou,et al.  Disruption of Dictyostelium PI3K genes reduces [32P]phosphatidylinositol 3,4 bisphosphate and [32P]phosphatidylinositol trisphosphate levels, alters F-actin distribution and impairs pinocytosis. , 1998, Journal of cell science.

[12]  D. Knecht,et al.  Use of a fusion protein between GFP and an actin-binding domain to visualize transient filamentous-actin structures , 1998, Current Biology.

[13]  Ching-shih Chen,et al.  A Novel Integrin-activated Pathway Forms PKB/Akt- stimulatory Phosphatidylinositol 3,4-Bisphosphate via Phosphatidylinositol 3-Phosphate in Platelets* , 1998, The Journal of Biological Chemistry.

[14]  Hui Ma,et al.  Chemoattractant‐mediated transient activation and membrane localization of Akt/PKB is required for efficient chemotaxis to cAMP in Dictyostelium , 1999, The EMBO journal.

[15]  J. van der Kaay,et al.  Distinct Phosphatidylinositol 3-Kinase Lipid Products Accumulate upon Oxidative and Osmotic Stress and Lead to Different Cellular Responses* , 1999, The Journal of Biological Chemistry.

[16]  C J Weijer,et al.  Morphogenetic cell movement in Dictyostelium. , 1999, Seminars in cell & developmental biology.

[17]  S. Dowler,et al.  DAPP1: a dual adaptor for phosphotyrosine and 3-phosphoinositides. , 1999, The Biochemical journal.

[18]  S. Dowler,et al.  Identification of pleckstrin-homology-domain-containing proteins with novel phosphoinositide-binding specificities. , 2000, The Biochemical journal.

[19]  J W Sedat,et al.  Polarization of chemoattractant receptor signaling during neutrophil chemotaxis. , 2000, Science.

[20]  D. Lucas,et al.  Structure, function, and biology of SHIP proteins. , 2000, Genes & development.

[22]  S. Dowler,et al.  Crystal structure of the phosphatidylinositol 3,4-bisphosphate-binding pleckstrin homology (PH) domain of tandem PH-domain-containing protein 1 (TAPP1): molecular basis of lipid specificity , 2001 .

[23]  C J Weijer,et al.  Propagating chemoattractant waves coordinate periodic cell movement in Dictyostelium slugs. , 2001, Development.

[24]  J. Cardelli,et al.  p110-related PI 3-kinases regulate phagosome-phagosome fusion and phagosomal pH through a PKB/Akt dependent pathway in Dictyostelium. , 2001, Journal of cell science.

[25]  G S Taylor,et al.  PTEN and myotubularin: novel phosphoinositide phosphatases. , 2001, Annual review of biochemistry.

[26]  S. Grinstein,et al.  Distinct roles of class I and class III phosphatidylinositol 3-kinases in phagosome formation and maturation , 2001, The Journal of cell biology.

[27]  C. Helgason,et al.  A Regulatory Role for Src Homology 2 Domain–Containing Inositol 5′-Phosphatase (Ship) in Phagocytosis Mediated by Fcγ Receptors and Complement Receptor 3 (αMβ2; Cd11b/Cd18) , 2001, The Journal of experimental medicine.

[28]  Paul Tempst,et al.  PtdIns(3)P regulates the neutrophil oxidase complex by binding to the PX domain of p40phox , 2001, Nature Cell Biology.

[29]  M. Yaffe,et al.  The PX domains of p47phox and p40phox bind to lipid products of PI(3)K , 2001, Nature Cell Biology.

[30]  J. Cardelli,et al.  Phagocytosis and Macropinocytosis in Dictyostelium: Phosphoinositide‐Based Processes, Biochemically Distinct , 2001, Traffic.

[31]  M. Waterfield,et al.  Synthesis and function of 3-phosphorylated inositol lipids. , 2001, Annual review of biochemistry.

[32]  S. Grinstein,et al.  Restricted Accumulation of Phosphatidylinositol 3-Kinase Products in a Plasmalemmal Subdomain during Fcγ Receptor-Mediated Phagocytosis , 2001, The Journal of cell biology.

[33]  P. Lipp,et al.  Phosphatidylinositol 3-phosphate is generated in phagosomal membranes , 2001, Current Biology.

[34]  K. Lee,et al.  Sequential activities of phosphoinositide 3-kinase, PKB/Aakt, and Rab7 during macropinosome formation in Dictyostelium. , 2001, Molecular biology of the cell.

[35]  P. Devreotes,et al.  Tumor Suppressor PTEN Mediates Sensing of Chemoattractant Gradients , 2002, Cell.

[36]  Richard A. Firtel,et al.  Spatial and Temporal Regulation of 3-Phosphoinositides by PI 3-Kinase and PTEN Mediates Chemotaxis , 2002, Cell.

[37]  D. Dormann,et al.  Visualizing PI3 Kinase-Mediated Cell-Cell Signaling during Dictyostelium Development , 2002, Current Biology.

[38]  D. Dormann,et al.  Simultaneous quantification of cell motility and protein-membrane-association using active contours. , 2002, Cell motility and the cytoskeleton.

[39]  Jerónimo Bravo,et al.  Binding of the PX domain of p47phox to phosphatidylinositol 3,4‐bisphosphate and phosphatidic acid is masked by an intramolecular interaction , 2002, The EMBO journal.

[40]  P. Hawkins,et al.  Roles of PI3Ks in leukocyte chemotaxis and phagocytosis. , 2002, Current opinion in cell biology.

[41]  P. Devreotes,et al.  Temporal and spatial regulation of chemotaxis. , 2002, Developmental cell.

[42]  Richard A. Firtel,et al.  Leading the way: directional sensing through phosphatidylinositol 3-kinase and other signaling pathways , 2003, Journal of Cell Science.

[43]  P. V. van Haastert,et al.  A Diverse Family of Inositol 5-Phosphatases Playing a Role in Growth and Development in Dictyostelium discoideum * 210 , 2003, The Journal of Biological Chemistry.

[44]  P. Devreotes,et al.  Two phases of actin polymerization display different dependencies on PI(3,4,5)P3 accumulation and have unique roles during chemotaxis. , 2003, Molecular biology of the cell.

[45]  M. Lemmon,et al.  Phosphoinositide Recognition Domains , 2003, Traffic.

[46]  R. Firtel,et al.  Receptor-mediated regulation of PI3Ks confines PI(3,4,5)P3 to the leading edge of chemotaxing cells. , 2003, Molecular biology of the cell.