Inositol phospholipids regulate the guanine-nucleotide-exchange factor Tiam1 by facilitating its binding to the plasma membrane and regulating GDP/GTP exchange on Rac1

Ian Neil Fleming, Ian H Batty, Alan R Prescott, Alex Gray, Gursant Singh Kular, Hazel Stewart, C Peter Downes

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

Binding of the Rac1-specific guanine-nucleotide-exchange factor, Tiam1, to the plasma membrane requires the N-terminal pleckstrin homology domain. In the present study, we show that membrane-association is mediated by binding of PtdIns(4,5)P(2) to the pleckstrin homology domain. Moreover, in 1321N1 astrocytoma cells, translocation of Tiam1 to the cytosol, following receptor-mediated stimulation of PtdIns(4,5)P(2) breakdown, correlates with decreased Rac1-GTP levels, indicating that membrane-association is required for GDP/GTP exchange on Rac1. In addition, we show that platelet-derived growth factor activates Rac1 in vivo by increasing PtdIns(3,4,5)P(3) concentrations, rather than the closely related lipid, PtdIns(3,4)P(2). Finally, the data demonstrate that PtdIns(4,5)P(2) and PtdIns(3,4,5)P(3) bind to the same pleckstrin homology domain in Tiam1 and that soluble inositol phosphates appear to compete with lipids for this binding. Together, these novel observations provide strong evidence that distinct phosphoinositides regulate different functions of this enzyme, indicating that local concentrations of signalling lipids and the levels of cytosolic inositol phosphates will play crucial roles in determining its activity in vivo.
Original languageEnglish
Pages (from-to)857-865
Number of pages9
JournalBiochemical Journal
Volume382
Issue number3
DOIs
Publication statusPublished - 15 Sep 2004

Fingerprint

Guanine Nucleotide Exchange Factors
Cell membranes
Phosphatidylinositols
Guanosine Triphosphate
Cell Membrane
Inositol Phosphates
Lipids
Membranes
Platelet-Derived Growth Factor
Astrocytoma
Cytosol
Enzymes
phosphatidylinositol 5-phosphate
Pleckstrin Homology Domains

Keywords

  • androstadienes
  • tumor cell line
  • cell membrane
  • cytosol
  • guanine nucleotide exchange factors
  • guanosine diphosphate
  • guanosine triphosphate
  • humans
  • peptide fragments
  • phosphatidylinositol 3-kinases
  • phosphatidylinositol 4,5-diphosphate
  • phosphatidylinositol phosphates
  • platelet-derived growth factor
  • protein binding
  • protein structure, tertiary
  • protein transport
  • proteins
  • receptors, cytoplasmic and nuclear
  • recombinant fusion proteins
  • thrombin
  • rac1 GTP-binding protein
  • pleckstrin homology domains
  • structural basis
  • Tiam1
  • translocation
  • activation
  • cells
  • inositol phospholipids

Cite this

Inositol phospholipids regulate the guanine-nucleotide-exchange factor Tiam1 by facilitating its binding to the plasma membrane and regulating GDP/GTP exchange on Rac1. / Fleming, Ian Neil; Batty, Ian H; Prescott, Alan R; Gray, Alex; Kular, Gursant Singh; Stewart, Hazel; Downes, C Peter.

In: Biochemical Journal, Vol. 382, No. 3, 15.09.2004, p. 857-865.

Research output: Contribution to journalArticle

Fleming, Ian Neil ; Batty, Ian H ; Prescott, Alan R ; Gray, Alex ; Kular, Gursant Singh ; Stewart, Hazel ; Downes, C Peter. / Inositol phospholipids regulate the guanine-nucleotide-exchange factor Tiam1 by facilitating its binding to the plasma membrane and regulating GDP/GTP exchange on Rac1. In: Biochemical Journal. 2004 ; Vol. 382, No. 3. pp. 857-865.
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T1 - Inositol phospholipids regulate the guanine-nucleotide-exchange factor Tiam1 by facilitating its binding to the plasma membrane and regulating GDP/GTP exchange on Rac1

AU - Fleming, Ian Neil

AU - Batty, Ian H

AU - Prescott, Alan R

AU - Gray, Alex

AU - Kular, Gursant Singh

AU - Stewart, Hazel

AU - Downes, C Peter

PY - 2004/9/15

Y1 - 2004/9/15

N2 - Binding of the Rac1-specific guanine-nucleotide-exchange factor, Tiam1, to the plasma membrane requires the N-terminal pleckstrin homology domain. In the present study, we show that membrane-association is mediated by binding of PtdIns(4,5)P(2) to the pleckstrin homology domain. Moreover, in 1321N1 astrocytoma cells, translocation of Tiam1 to the cytosol, following receptor-mediated stimulation of PtdIns(4,5)P(2) breakdown, correlates with decreased Rac1-GTP levels, indicating that membrane-association is required for GDP/GTP exchange on Rac1. In addition, we show that platelet-derived growth factor activates Rac1 in vivo by increasing PtdIns(3,4,5)P(3) concentrations, rather than the closely related lipid, PtdIns(3,4)P(2). Finally, the data demonstrate that PtdIns(4,5)P(2) and PtdIns(3,4,5)P(3) bind to the same pleckstrin homology domain in Tiam1 and that soluble inositol phosphates appear to compete with lipids for this binding. Together, these novel observations provide strong evidence that distinct phosphoinositides regulate different functions of this enzyme, indicating that local concentrations of signalling lipids and the levels of cytosolic inositol phosphates will play crucial roles in determining its activity in vivo.

AB - Binding of the Rac1-specific guanine-nucleotide-exchange factor, Tiam1, to the plasma membrane requires the N-terminal pleckstrin homology domain. In the present study, we show that membrane-association is mediated by binding of PtdIns(4,5)P(2) to the pleckstrin homology domain. Moreover, in 1321N1 astrocytoma cells, translocation of Tiam1 to the cytosol, following receptor-mediated stimulation of PtdIns(4,5)P(2) breakdown, correlates with decreased Rac1-GTP levels, indicating that membrane-association is required for GDP/GTP exchange on Rac1. In addition, we show that platelet-derived growth factor activates Rac1 in vivo by increasing PtdIns(3,4,5)P(3) concentrations, rather than the closely related lipid, PtdIns(3,4)P(2). Finally, the data demonstrate that PtdIns(4,5)P(2) and PtdIns(3,4,5)P(3) bind to the same pleckstrin homology domain in Tiam1 and that soluble inositol phosphates appear to compete with lipids for this binding. Together, these novel observations provide strong evidence that distinct phosphoinositides regulate different functions of this enzyme, indicating that local concentrations of signalling lipids and the levels of cytosolic inositol phosphates will play crucial roles in determining its activity in vivo.

KW - androstadienes

KW - tumor cell line

KW - cell membrane

KW - cytosol

KW - guanine nucleotide exchange factors

KW - guanosine diphosphate

KW - guanosine triphosphate

KW - humans

KW - peptide fragments

KW - phosphatidylinositol 3-kinases

KW - phosphatidylinositol 4,5-diphosphate

KW - phosphatidylinositol phosphates

KW - platelet-derived growth factor

KW - protein binding

KW - protein structure, tertiary

KW - protein transport

KW - proteins

KW - receptors, cytoplasmic and nuclear

KW - recombinant fusion proteins

KW - thrombin

KW - rac1 GTP-binding protein

KW - pleckstrin homology domains

KW - structural basis

KW - Tiam1

KW - translocation

KW - activation

KW - cells

KW - inositol phospholipids

U2 - 10.1042/BJ20040916

DO - 10.1042/BJ20040916

M3 - Article

VL - 382

SP - 857

EP - 865

JO - Biochemical Journal

JF - Biochemical Journal

SN - 0264-6021

IS - 3

ER -