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Title: Protein kinase C regulates the phosphorylation and oligomerization of ERM binding phosphoprotein 50

Abstract

Ezrin-Radixin-Moesin (ERM) binding phosphoprotein 50 (EBP50, a.k.a. NHERF-1) is a scaffold protein essential for the localization and coordinated activity of apical transporters, enzymes and receptors in epithelial cells. EBP50 acts via multiple protein binding interactions, including oligomerization through interactions of its PSD95-Dlg-ZO1 (PDZ) domains. EBP50 can be phosphorylated on multiple sites and phosphorylation of specific sites modulates the extent of oligomerization. The aim of the present study was to test the capacity of protein kinase C (PKC) to phosphorylate EBP50 and to regulate its oligomerization. In vitro experiments showed that the catalytic subunit of PKC directly phosphorylates EBP50. In HEK-293 cells transfected with rat EBP50 cDNA, a treatment with 12 myristate 13-acetate (PMA) induced a translocation of PKC{alpha} and {beta} isoforms to the membrane and increased {sup 32}P incorporation into EBP50. In co-transfection/co-precipitation studies, PMA treatment stimulated EBP50 oligomerization. Mass spectrometry analysis of full-length EBP50 and phosphorylation analyses of specific domains, and of mutated or truncated forms of EBP50, indicated that PKC-induced phosphorylation of EBP50 occurred on the Ser{sup 337}/Ser{sup 338} residue within the carboxyl-tail domain of the protein. Truncation of Ser{sup 337}/Ser{sup 338} also diminished PKC-induced oligomerization of EBP50. These results suggest the PKC signaling pathway can impact EBP50-dependentmore » cellular functions by regulating EBP50 oligomerization.« less

Authors:
 [1];  [2];  [2];  [2];  [1];  [3];  [3];  [1];  [4]
  1. Inserm U680, UPMC and Service de Biochimie et d'Hormonologie AP-HP, Hopital Tenon, Faculte de Medecine Saint-Antoine, Paris, F-75012 (France)
  2. Department of Medicine, University of Colorado Health Sciences Center, Box B158, 4200 E. 9th Avenue, Denver, CO 80262 (United States)
  3. Department of Cellular and Structural Biology, University of Colorado Health Sciences Center, Denver, CO 80262 (United States)
  4. Department of Medicine, University of Colorado Health Sciences Center, Box B158, 4200 E. 9th Avenue, Denver, CO 80262 (United States) and Department of Cellular and Structural Biology, University of Colorado Health Sciences Center, Denver, CO 80262 (United States). E-mail: brian.doctor@uchsc.edu
Publication Date:
OSTI Identifier:
20717608
Resource Type:
Journal Article
Resource Relation:
Journal Name: Experimental Cell Research; Journal Volume: 306; Journal Issue: 1; Other Information: DOI: 10.1016/j.yexcr.2005.02.011; PII: S0014-4827(05)00079-0; Copyright (c) 2005 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; ACETATES; COPRECIPITATION; ENZYMES; IN VITRO; MASS SPECTROSCOPY; PHOSPHOPROTEINS; PHOSPHORYLATION; RATS; RECEPTORS; TRANSLOCATION

Citation Formats

Fouassier, Laura, Nichols, Matthew T., Gidey, Elizabeth, McWilliams, Ryan R., Robin, Helene, Finnigan, Claire, Howell, Kathryn E., Housset, Chantal, and Doctor, R. Brian. Protein kinase C regulates the phosphorylation and oligomerization of ERM binding phosphoprotein 50. United States: N. p., 2005. Web. doi:10.1016/j.yexcr.2005.02.011.
Fouassier, Laura, Nichols, Matthew T., Gidey, Elizabeth, McWilliams, Ryan R., Robin, Helene, Finnigan, Claire, Howell, Kathryn E., Housset, Chantal, & Doctor, R. Brian. Protein kinase C regulates the phosphorylation and oligomerization of ERM binding phosphoprotein 50. United States. doi:10.1016/j.yexcr.2005.02.011.
Fouassier, Laura, Nichols, Matthew T., Gidey, Elizabeth, McWilliams, Ryan R., Robin, Helene, Finnigan, Claire, Howell, Kathryn E., Housset, Chantal, and Doctor, R. Brian. Sun . "Protein kinase C regulates the phosphorylation and oligomerization of ERM binding phosphoprotein 50". United States. doi:10.1016/j.yexcr.2005.02.011.
@article{osti_20717608,
title = {Protein kinase C regulates the phosphorylation and oligomerization of ERM binding phosphoprotein 50},
author = {Fouassier, Laura and Nichols, Matthew T. and Gidey, Elizabeth and McWilliams, Ryan R. and Robin, Helene and Finnigan, Claire and Howell, Kathryn E. and Housset, Chantal and Doctor, R. Brian},
abstractNote = {Ezrin-Radixin-Moesin (ERM) binding phosphoprotein 50 (EBP50, a.k.a. NHERF-1) is a scaffold protein essential for the localization and coordinated activity of apical transporters, enzymes and receptors in epithelial cells. EBP50 acts via multiple protein binding interactions, including oligomerization through interactions of its PSD95-Dlg-ZO1 (PDZ) domains. EBP50 can be phosphorylated on multiple sites and phosphorylation of specific sites modulates the extent of oligomerization. The aim of the present study was to test the capacity of protein kinase C (PKC) to phosphorylate EBP50 and to regulate its oligomerization. In vitro experiments showed that the catalytic subunit of PKC directly phosphorylates EBP50. In HEK-293 cells transfected with rat EBP50 cDNA, a treatment with 12 myristate 13-acetate (PMA) induced a translocation of PKC{alpha} and {beta} isoforms to the membrane and increased {sup 32}P incorporation into EBP50. In co-transfection/co-precipitation studies, PMA treatment stimulated EBP50 oligomerization. Mass spectrometry analysis of full-length EBP50 and phosphorylation analyses of specific domains, and of mutated or truncated forms of EBP50, indicated that PKC-induced phosphorylation of EBP50 occurred on the Ser{sup 337}/Ser{sup 338} residue within the carboxyl-tail domain of the protein. Truncation of Ser{sup 337}/Ser{sup 338} also diminished PKC-induced oligomerization of EBP50. These results suggest the PKC signaling pathway can impact EBP50-dependent cellular functions by regulating EBP50 oligomerization.},
doi = {10.1016/j.yexcr.2005.02.011},
journal = {Experimental Cell Research},
number = 1,
volume = 306,
place = {United States},
year = {Sun May 15 00:00:00 EDT 2005},
month = {Sun May 15 00:00:00 EDT 2005}
}