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Phosphorylation of Spinophilin Modulates Its Interaction with Actin Filaments

Hsieh-Wilson, Linda C. and Benfenati, Fabio and Snyder, Gretchen L. and Allen, Patrick B. and Nairn, Angus C. and Greengard, Paul (2003) Phosphorylation of Spinophilin Modulates Its Interaction with Actin Filaments. Journal of Biological Chemistry, 278 (2). pp. 1186-1194. ISSN 0021-9258. http://resolver.caltech.edu/CaltechAUTHORS:HSIjbc03

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Abstract

Spinophilin is a protein phosphatase 1 (PP1)- and actin-binding protein that modulates excitatory synaptic transmission and dendritic spine morphology. We report that spinophilin is phosphorylated in vitro by protein kinase A (PKA). Phosphorylation of spinophilin was stimulated by treatment of neostriatal neurons with a dopamine D1 receptor agonist or with forskolin, consistent with spinophilin being a substrate for PKA in intact cells. Using tryptic phosphopeptide mapping, site-directed mutagenesis, and microsequencing analysis, we identified two major sites of phosphorylation, Ser-94 and Ser-177, that are located within the actin-binding domain of spinophilin. Phosphorylation of spinophilin by PKA modulated the association between spinophilin and the actin cytoskeleton. Following subcellular fractionation, unphosphorylated spinophilin was enriched in the postsynaptic density, whereas a pool of phosphorylated spinophilin was found in the cytosol. F-actin co-sedimentation and overlay analysis revealed that phosphorylation of spinophilin reduced the stoichiometry of the spinophilin-actin interaction. In contrast, the ability of spinophilin to bind to PP1 remained unchanged. Taken together, our studies suggest that phosphorylation of spinophilin by PKA modulates the anchoring of the spinophilin-PP1 complex within dendritic spines, thereby likely contributing to the efficacy and plasticity of synaptic transmission.


Item Type:Article
Additional Information:Copyright © 2003 by The American Society for Biochemistry and Molecular Biology, Inc. Received for publication, June 10, 2002, and in revised form, October 31, 2002. Originally published In Press as doi:10.1074/jbc.M205754200 on November 1, 2002 We thank Martin Lan, Mercedes Paredes, Dr. Stacie Grossman, Jean Whitesell (Cocalico Biologicals, Inc.), and Dr. Joseph Fernandez (Rockefeller University Protein/DNA Technology Center) for valuable assistance. This work was supported by United States Public Health Services Grants MH40899 and DA10044 and by Fellowship DRG-1451 of the Cancer Research Fund of the Damon Runyon-Walter Winchell Foundation (to L. C. H.-W.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Record Number:CaltechAUTHORS:HSIjbc03
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:HSIjbc03
Alternative URL:http://dx.doi.org/10.1074/jbc.M205754200
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:6903
Collection:CaltechAUTHORS
Deposited By: Archive Administrator
Deposited On:01 Jan 2007
Last Modified:26 Dec 2012 09:26

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