Twinfilin is a ubiquitous actin monomer–binding protein that regulates actin filament turnover in yeast and mammalian cells. To elucidate the mechanism by which twinfilin contributes to actin filament dynamics, we carried out an analysis of yeast twinfilin, and we show here that twinfilin is an abundant protein that localizes to cortical actin patches in wild-type yeast cells. Native gel assays demonstrate that twinfilin binds ADP-actin monomers with higher affinity than ATP-actin monomers. A mutant twinfilin that does not interact with actin monomers in vitro no longer localizes to cortical actin patches when expressed in yeast, suggesting that the ability to interact with actin monomers may be essential for the localization of twinfilin. The localization of twinfilin to the cortical actin cytoskeleton is also disrupted in yeast strains where either the CAP1 or CAP2 gene, encoding for the α and β subunits of capping protein, is deleted. Purified twinfilin and capping protein form a complex on native gels. Twinfilin also interacts with phosphatidylinositol 4,5-bisphosphate (PI[4,5]P2), and its actin monomer–sequestering activity is inhibited by PI(4,5)P2. Based on these results, we propose a model for the biological role of twinfilin as a protein that localizes actin monomers to the sites of rapid filament assembly in cells.
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15 October 2001
Article|
October 15 2001
Interactions with PIP2, ADP-actin monomers, and capping protein regulate the activity and localization of yeast twinfilin
Sandra Palmgren,
Sandra Palmgren
1Program in Cellular Biotechnology, Institute of Biotechnology, FIN-00014 University of Helsinki, Helsinki, Finland
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Pauli J. Ojala,
Pauli J. Ojala
1Program in Cellular Biotechnology, Institute of Biotechnology, FIN-00014 University of Helsinki, Helsinki, Finland
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Martin A. Wear,
Martin A. Wear
2Department of Cell Biology and Physiology, Washington University, St. Louis, MO 63110
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John A. Cooper,
John A. Cooper
2Department of Cell Biology and Physiology, Washington University, St. Louis, MO 63110
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Pekka Lappalainen
Pekka Lappalainen
1Program in Cellular Biotechnology, Institute of Biotechnology, FIN-00014 University of Helsinki, Helsinki, Finland
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Sandra Palmgren
1Program in Cellular Biotechnology, Institute of Biotechnology, FIN-00014 University of Helsinki, Helsinki, Finland
Pauli J. Ojala
1Program in Cellular Biotechnology, Institute of Biotechnology, FIN-00014 University of Helsinki, Helsinki, Finland
Martin A. Wear
2Department of Cell Biology and Physiology, Washington University, St. Louis, MO 63110
John A. Cooper
2Department of Cell Biology and Physiology, Washington University, St. Louis, MO 63110
Pekka Lappalainen
1Program in Cellular Biotechnology, Institute of Biotechnology, FIN-00014 University of Helsinki, Helsinki, Finland
Address correspondence to Pekka Lappalainen, Institute of Biotechnology, P.O. Box 56 (Viikinkaari 9), FIN-00014 University of Helsinki, Helsinki, Finland. Tel.: 358-9-1915-9499. Fax: 358-9-1915-9366. E-mail: [email protected]
*
Abbreviations used in this paper: ADF, actin-depolymerizing factor; GFP, green fluorescent protein; PI(4,5)P2, phosphatidylinositol 4,5-bisphosphate.
Received:
June 29 2001
Revision Received:
August 30 2001
Accepted:
September 05 2001
Online ISSN: 1540-8140
Print ISSN: 0021-9525
The Rockefeller University Press
2001
J Cell Biol (2001) 155 (2): 251–260.
Article history
Received:
June 29 2001
Revision Received:
August 30 2001
Accepted:
September 05 2001
Citation
Sandra Palmgren, Pauli J. Ojala, Martin A. Wear, John A. Cooper, Pekka Lappalainen; Interactions with PIP2, ADP-actin monomers, and capping protein regulate the activity and localization of yeast twinfilin . J Cell Biol 15 October 2001; 155 (2): 251–260. doi: https://doi.org/10.1083/jcb.200106157
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