Gelsolin binding to phosphatidylinositol 4,5-bisphosphate is modulated by calcium and pH

Keng Mean Lin, Elizabeth Wenegieme, Pei-Jung Lu, Ching Shih Chen, Helen L. Yin

Research output: Contribution to journalArticle

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Abstract

The actin cytoskeleton of nonmuscle cells undergoes extensive remodeling during agonist stimulation. Lamellipodial extension is initiated by uncapping of actin nuclei at the cortical cytoplasm to allow filament elongation. Many actin filament capping proteins are regulated by phosphatidylinositol 4,5- bisphosphate (PIP2), which is hydrolyzed by phospholipase C. It is hypothesized that PIPe dissociates capping proteins from filament ends to promote actin assembly. However, since actin polymerization often occurs at a time when PIP2 concentration is decreased rather than increased, capping protein interactions with PIP2 may not be regulated solely by the bulk PIP2 concentration. We present evidence that PIP2 binding to the gelsolin family of capping proteins is enhanced by Ca2+. Binding was examined by equilibrium and nonequilibrium gel filtration and by monitoring intrinsic tryptophan fluorescence. Gelsolin and CapG affinity for PIP2 were increased 8- and 4-fold, respectively, by μM Ca2+, and the Ca2+ requirement was reduced by lowering the pH from 7.5 to 7.0. Studies with the NH2- and COOH- terminal halves of gelsolin showed that PIP2 binding occurred primarily at the NH2-terminal half, and Ca2+ exposed its PIP2 binding sites through a change in the COOH-terminal half. Mild acidification promotes PIP2 binding by directly affecting the NH2-terminal sites. Our findings can ex plain increased PIP2-induced uncapping even as the PIP2 concentration drops during cell activation. The change in gelsolin family PIP2 binding affinity during cell activation can impact divergent PIP2-dependent processes by altering PIP2 availability. Cross-talk be tween these proteins provides a multilayered mechanism for positive and negative modulation of signal transduction from the plasma membrane to the cytoskeleton.

Original languageEnglish
Pages (from-to)20443-20450
Number of pages8
JournalJournal of Biological Chemistry
Volume272
Issue number33
DOIs
Publication statusPublished - 1997 Aug 15

Fingerprint

Gelsolin
Phosphatidylinositols
Actins
Calcium
Actin Capping Proteins
Proteins
Chemical activation
Signal transduction
Polysorbates
Acidification
Type C Phospholipases
Cell membranes
Cytoskeleton
Actin Cytoskeleton
Tryptophan
Polymerization
Gel Chromatography
Elongation
Signal Transduction
Cytoplasm

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Lin, Keng Mean ; Wenegieme, Elizabeth ; Lu, Pei-Jung ; Chen, Ching Shih ; Yin, Helen L. / Gelsolin binding to phosphatidylinositol 4,5-bisphosphate is modulated by calcium and pH. In: Journal of Biological Chemistry. 1997 ; Vol. 272, No. 33. pp. 20443-20450.
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abstract = "The actin cytoskeleton of nonmuscle cells undergoes extensive remodeling during agonist stimulation. Lamellipodial extension is initiated by uncapping of actin nuclei at the cortical cytoplasm to allow filament elongation. Many actin filament capping proteins are regulated by phosphatidylinositol 4,5- bisphosphate (PIP2), which is hydrolyzed by phospholipase C. It is hypothesized that PIPe dissociates capping proteins from filament ends to promote actin assembly. However, since actin polymerization often occurs at a time when PIP2 concentration is decreased rather than increased, capping protein interactions with PIP2 may not be regulated solely by the bulk PIP2 concentration. We present evidence that PIP2 binding to the gelsolin family of capping proteins is enhanced by Ca2+. Binding was examined by equilibrium and nonequilibrium gel filtration and by monitoring intrinsic tryptophan fluorescence. Gelsolin and CapG affinity for PIP2 were increased 8- and 4-fold, respectively, by μM Ca2+, and the Ca2+ requirement was reduced by lowering the pH from 7.5 to 7.0. Studies with the NH2- and COOH- terminal halves of gelsolin showed that PIP2 binding occurred primarily at the NH2-terminal half, and Ca2+ exposed its PIP2 binding sites through a change in the COOH-terminal half. Mild acidification promotes PIP2 binding by directly affecting the NH2-terminal sites. Our findings can ex plain increased PIP2-induced uncapping even as the PIP2 concentration drops during cell activation. The change in gelsolin family PIP2 binding affinity during cell activation can impact divergent PIP2-dependent processes by altering PIP2 availability. Cross-talk be tween these proteins provides a multilayered mechanism for positive and negative modulation of signal transduction from the plasma membrane to the cytoskeleton.",
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Gelsolin binding to phosphatidylinositol 4,5-bisphosphate is modulated by calcium and pH. / Lin, Keng Mean; Wenegieme, Elizabeth; Lu, Pei-Jung; Chen, Ching Shih; Yin, Helen L.

In: Journal of Biological Chemistry, Vol. 272, No. 33, 15.08.1997, p. 20443-20450.

Research output: Contribution to journalArticle

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T1 - Gelsolin binding to phosphatidylinositol 4,5-bisphosphate is modulated by calcium and pH

AU - Lin, Keng Mean

AU - Wenegieme, Elizabeth

AU - Lu, Pei-Jung

AU - Chen, Ching Shih

AU - Yin, Helen L.

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N2 - The actin cytoskeleton of nonmuscle cells undergoes extensive remodeling during agonist stimulation. Lamellipodial extension is initiated by uncapping of actin nuclei at the cortical cytoplasm to allow filament elongation. Many actin filament capping proteins are regulated by phosphatidylinositol 4,5- bisphosphate (PIP2), which is hydrolyzed by phospholipase C. It is hypothesized that PIPe dissociates capping proteins from filament ends to promote actin assembly. However, since actin polymerization often occurs at a time when PIP2 concentration is decreased rather than increased, capping protein interactions with PIP2 may not be regulated solely by the bulk PIP2 concentration. We present evidence that PIP2 binding to the gelsolin family of capping proteins is enhanced by Ca2+. Binding was examined by equilibrium and nonequilibrium gel filtration and by monitoring intrinsic tryptophan fluorescence. Gelsolin and CapG affinity for PIP2 were increased 8- and 4-fold, respectively, by μM Ca2+, and the Ca2+ requirement was reduced by lowering the pH from 7.5 to 7.0. Studies with the NH2- and COOH- terminal halves of gelsolin showed that PIP2 binding occurred primarily at the NH2-terminal half, and Ca2+ exposed its PIP2 binding sites through a change in the COOH-terminal half. Mild acidification promotes PIP2 binding by directly affecting the NH2-terminal sites. Our findings can ex plain increased PIP2-induced uncapping even as the PIP2 concentration drops during cell activation. The change in gelsolin family PIP2 binding affinity during cell activation can impact divergent PIP2-dependent processes by altering PIP2 availability. Cross-talk be tween these proteins provides a multilayered mechanism for positive and negative modulation of signal transduction from the plasma membrane to the cytoskeleton.

AB - The actin cytoskeleton of nonmuscle cells undergoes extensive remodeling during agonist stimulation. Lamellipodial extension is initiated by uncapping of actin nuclei at the cortical cytoplasm to allow filament elongation. Many actin filament capping proteins are regulated by phosphatidylinositol 4,5- bisphosphate (PIP2), which is hydrolyzed by phospholipase C. It is hypothesized that PIPe dissociates capping proteins from filament ends to promote actin assembly. However, since actin polymerization often occurs at a time when PIP2 concentration is decreased rather than increased, capping protein interactions with PIP2 may not be regulated solely by the bulk PIP2 concentration. We present evidence that PIP2 binding to the gelsolin family of capping proteins is enhanced by Ca2+. Binding was examined by equilibrium and nonequilibrium gel filtration and by monitoring intrinsic tryptophan fluorescence. Gelsolin and CapG affinity for PIP2 were increased 8- and 4-fold, respectively, by μM Ca2+, and the Ca2+ requirement was reduced by lowering the pH from 7.5 to 7.0. Studies with the NH2- and COOH- terminal halves of gelsolin showed that PIP2 binding occurred primarily at the NH2-terminal half, and Ca2+ exposed its PIP2 binding sites through a change in the COOH-terminal half. Mild acidification promotes PIP2 binding by directly affecting the NH2-terminal sites. Our findings can ex plain increased PIP2-induced uncapping even as the PIP2 concentration drops during cell activation. The change in gelsolin family PIP2 binding affinity during cell activation can impact divergent PIP2-dependent processes by altering PIP2 availability. Cross-talk be tween these proteins provides a multilayered mechanism for positive and negative modulation of signal transduction from the plasma membrane to the cytoskeleton.

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