Note to users. If you're seeing this message, it means that your browser cannot find this page's style/presentation instructions -- or possibly that you are using a browser that does not support current Web standards. Find out more about why this message is appearing, and what you can do to make your experience of our site the best it can be.


Logo for

J. Biol. Chem. 283 (52): 36636-36645

© 2008 by The American Society for Biochemistry and Molecular Biology, Inc.

Role of Phosphatidic Acid in the Coupling of the ERK Cascade*


Catherine A. Kraft1, José Luis Garrido1, Eric Fluharty, Luis Leiva-Vega, , and Guillermo Romero2

Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261

Abstract: The production of phosphatidic acid plays a crucial role in the activation of the ERK cascade. This role was linked to the binding of phosphatidate to a specific polybasic site within the kinase domain of Raf-1. Here we show that phosphatidate promotes ERK phosphorylation in intact cells but does not activate Raf in vitro. The kinase suppressor of Ras (KSR) contains a sequence homologous to the phosphatidate binding site of Raf-1. Direct binding of phosphatidate to synthetic peptides derived from the sequences of the binding domains of Raf-1 and KSR was demonstrated by spectroscopic techniques. The specificity of these interactions was confirmed using synthetic lipids and mutated peptides in which the core of the phosphatidic acid binding domain was disrupted. Insulin and exogenous dioleoyl phosphatidate induced a rapid translocation of a mouse KSR1-EGFP construct to the plasma membrane of HIRcB cells. Mutation of two arginines located in the core of the putative phosphatidate binding site abolished dioleoyl phosphatidate- and insulin-induced translocation of KSR1. Overexpression of the mutant KSR1 in HIRcB cells inhibited insulin-dependent MEK and ERK phosphorylation. The addition of dioleoyl phosphatidate or insulin increased the co-localization of KSR1 and H-Ras and promoted the formation of plasma membrane patches enriched in both proteins and phosphatidic acid. These results, in conjunction with our previous work, suggest the formation of phosphatidate-enriched membrane microdomains that contain all components of the ERK cascade. We propose that these domains act as molecular scaffolds in the coupling of signaling events.

Received for publication June 17, 2008. Revision received October 22, 2008.

* This work was supported, in whole or in part, by National Institutes of Health Grants R01-DK-54782 and T32-GM-54813. This work was also supported by an internal grant from the Office of the Senior Vice Chancellor for the Health Sciences of the University of Pittsburgh. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.


The on-line version of this article (available at contains supplemental Figs. S1 and S2 and Movie S1.

1 Both of these authors contributed equally to this work.

2 To whom correspondence should be addressed. Tel.: 412-648-9408; E-mail: ggr{at}

Temporal Production of the Signaling Lipid Phosphatidic Acid by Phospholipase D2 Determines the Output of Extracellular Signal-Regulated Kinase Signaling in Cancer Cells.
F. Zhang, Z. Wang, M. Lu, Y. Yonekubo, X. Liang, Y. Zhang, P. Wu, Y. Zhou, S. Grinstein, J. F. Hancock, et al. (2014)
Mol. Cell. Biol. 34, 84-95
   Abstract »    Full Text »    PDF »
A New Signaling Pathway (JAK-Fes-phospholipase D) That Is Enhanced in Highly Proliferative Breast Cancer Cells.
Q. Ye, S. Kantonen, K. M. Henkels, and J. Gomez-Cambronero (2013)
J. Biol. Chem. 288, 9881-9891
   Abstract »    Full Text »    PDF »
Diabetes Diminishes Phosphatidic Acid in the Retina: A Putative Mediator for Reduced mTOR Signaling and Increased Neuronal Cell Death.
T. E. Fox, M. M. Young, M. M. Pedersen, X. Han, T. W. Gardner, and M. Kester (2012)
Invest. Ophthalmol. Vis. Sci. 53, 7257-7267
   Abstract »    Full Text »    PDF »
Lipin-1 Phosphatidic Phosphatase Activity Modulates Phosphatidate Levels to Promote Peroxisome Proliferator-activated Receptor {gamma} (PPAR{gamma}) Gene Expression during Adipogenesis.
P. Zhang, K. Takeuchi, L. S. Csaki, and K. Reue (2012)
J. Biol. Chem. 287, 3485-3494
   Abstract »    Full Text »    PDF »
Raf Family Kinases: Old Dogs Have Learned New Tricks.
D. Matallanas, M. Birtwistle, D. Romano, A. Zebisch, J. Rauch, A. von Kriegsheim, and W. Kolch (2011)
Genes & Cancer 2, 232-260
   Abstract »    Full Text »    PDF »
Drosophila Raf's N Terminus Contains a Novel Conserved Region and Can Contribute to Torso RTK Signaling.
J. Ding, O. Tchaicheeyan, and L. Ambrosio (2010)
Genetics 184, 717-729
   Abstract »    Full Text »    PDF »
Quantitative Analysis of Protein-Lipid Interactions Using Tryptophan Fluorescence.
C. A. Kraft, J. L. Garrido, L. Leiva-Vega, and G. Romero (2009)
Science Signaling 2, pl4
   Abstract »    Full Text »    PDF »

To Advertise     Find Products

Science Signaling. ISSN 1937-9145 (online), 1945-0877 (print). Pre-2008: Science's STKE. ISSN 1525-8882