Regulation of the Rac1-specific exchange factor Tiam1 involves both phosphoinositide 3-kinase-dependent and -independent components

I N Fleming, A Gray, C P Downes

Research output: Contribution to journalArticle

98 Citations (Scopus)

Abstract

The small GTPase Rac1 is involved in regulating membrane ruffling, gene transcription, cell-cycle progression and cell transformation, and some of these events are blocked by inhibitors of phosphoinositide 3-kinase (PI 3-kinase). Moreover, Rac1 can be activated by several guanine nucleotide exchange factors, which facilitate the release of GDP. We therefore investigated the ability of PI 3-kinase lipid products to regulate Tiam1, a Rac1-specific exchange factor. Tiam1 bound to polyphosphorylated inositol lipids in the rank order PtdIns(3,4,5)P(3)>PtdIns(3,4)P(2) >>PtdIns(4,5)P(2), and this binding could be attributed to the N-terminal pleckstrin-homology (N-PH) domain. Both PtdIns(3,4,5)P(3) and PtdIns(3,4)P(2) enhanced Tiam1 guanine nucleotide exchange activity in vitro, but PtdIns(4,5)P(2) had no effect. Co-expression of a constitutively active PI 3-kinase with Tiam1 increased the amount of GTP-bound Rac1 in vivo, a response which required the N-PH domain of Tiam1. Ectopic expression of Tiam1 caused membrane ruffling in Swiss 3T3 cells that was characterized by wortmannin-sensitive and -insensitive components, which required the N-PH domain and the C-terminal PH domain of Tiam1 respectively. These results reveal novel facets of Tiam1-dependent regulation of Rac1 function.
Original languageEnglish
Pages (from-to)173-82
Number of pages10
JournalBiochemical Journal
Volume351
Issue numberPt 1
Publication statusPublished - 2000

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1-Phosphatidylinositol 4-Kinase
Phosphatidylinositols
Phosphotransferases
Swiss 3T3 Cells
Lipids
Guanine Nucleotide Exchange Factors
cdc Genes
Guanine Nucleotides
Membranes
Monomeric GTP-Binding Proteins
Inositol
Guanosine Triphosphate
Pleckstrin Homology Domains
Transcription
phosphatidylinositol 5-phosphate
Genes
Cells

Keywords

  • Androstadienes
  • Animals
  • Blood Proteins
  • Cell Line
  • Cell Membrane
  • Enzyme Activation
  • Guanine Nucleotide Exchange Factors
  • Guanosine Diphosphate
  • Guanosine Triphosphate
  • Humans
  • Mice
  • Mutation
  • Phosphatidylinositol 3-Kinases
  • Phosphatidylinositols
  • Phosphoproteins
  • Protein Binding
  • Protein Structure, Tertiary
  • Proteins
  • Sequence Homology, Amino Acid
  • Substrate Specificity
  • rac1 GTP-Binding Protein

Cite this

Regulation of the Rac1-specific exchange factor Tiam1 involves both phosphoinositide 3-kinase-dependent and -independent components. / Fleming, I N; Gray, A; Downes, C P.

In: Biochemical Journal, Vol. 351, No. Pt 1, 2000, p. 173-82.

Research output: Contribution to journalArticle

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T1 - Regulation of the Rac1-specific exchange factor Tiam1 involves both phosphoinositide 3-kinase-dependent and -independent components

AU - Fleming, I N

AU - Gray, A

AU - Downes, C P

PY - 2000

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N2 - The small GTPase Rac1 is involved in regulating membrane ruffling, gene transcription, cell-cycle progression and cell transformation, and some of these events are blocked by inhibitors of phosphoinositide 3-kinase (PI 3-kinase). Moreover, Rac1 can be activated by several guanine nucleotide exchange factors, which facilitate the release of GDP. We therefore investigated the ability of PI 3-kinase lipid products to regulate Tiam1, a Rac1-specific exchange factor. Tiam1 bound to polyphosphorylated inositol lipids in the rank order PtdIns(3,4,5)P(3)>PtdIns(3,4)P(2) >>PtdIns(4,5)P(2), and this binding could be attributed to the N-terminal pleckstrin-homology (N-PH) domain. Both PtdIns(3,4,5)P(3) and PtdIns(3,4)P(2) enhanced Tiam1 guanine nucleotide exchange activity in vitro, but PtdIns(4,5)P(2) had no effect. Co-expression of a constitutively active PI 3-kinase with Tiam1 increased the amount of GTP-bound Rac1 in vivo, a response which required the N-PH domain of Tiam1. Ectopic expression of Tiam1 caused membrane ruffling in Swiss 3T3 cells that was characterized by wortmannin-sensitive and -insensitive components, which required the N-PH domain and the C-terminal PH domain of Tiam1 respectively. These results reveal novel facets of Tiam1-dependent regulation of Rac1 function.

AB - The small GTPase Rac1 is involved in regulating membrane ruffling, gene transcription, cell-cycle progression and cell transformation, and some of these events are blocked by inhibitors of phosphoinositide 3-kinase (PI 3-kinase). Moreover, Rac1 can be activated by several guanine nucleotide exchange factors, which facilitate the release of GDP. We therefore investigated the ability of PI 3-kinase lipid products to regulate Tiam1, a Rac1-specific exchange factor. Tiam1 bound to polyphosphorylated inositol lipids in the rank order PtdIns(3,4,5)P(3)>PtdIns(3,4)P(2) >>PtdIns(4,5)P(2), and this binding could be attributed to the N-terminal pleckstrin-homology (N-PH) domain. Both PtdIns(3,4,5)P(3) and PtdIns(3,4)P(2) enhanced Tiam1 guanine nucleotide exchange activity in vitro, but PtdIns(4,5)P(2) had no effect. Co-expression of a constitutively active PI 3-kinase with Tiam1 increased the amount of GTP-bound Rac1 in vivo, a response which required the N-PH domain of Tiam1. Ectopic expression of Tiam1 caused membrane ruffling in Swiss 3T3 cells that was characterized by wortmannin-sensitive and -insensitive components, which required the N-PH domain and the C-terminal PH domain of Tiam1 respectively. These results reveal novel facets of Tiam1-dependent regulation of Rac1 function.

KW - Androstadienes

KW - Animals

KW - Blood Proteins

KW - Cell Line

KW - Cell Membrane

KW - Enzyme Activation

KW - Guanine Nucleotide Exchange Factors

KW - Guanosine Diphosphate

KW - Guanosine Triphosphate

KW - Humans

KW - Mice

KW - Mutation

KW - Phosphatidylinositol 3-Kinases

KW - Phosphatidylinositols

KW - Phosphoproteins

KW - Protein Binding

KW - Protein Structure, Tertiary

KW - Proteins

KW - Sequence Homology, Amino Acid

KW - Substrate Specificity

KW - rac1 GTP-Binding Protein

M3 - Article

VL - 351

SP - 173

EP - 182

JO - Biochemical Journal

JF - Biochemical Journal

SN - 0264-6021

IS - Pt 1

ER -