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Originally published In Press as doi:10.1074/mcp.M600147-MCP200 on September 6, 2006.
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Molecular & Cellular Proteomics 5:2211-2227, 2006.
© 2006 by The American Society for Biochemistry and Molecular Biology, Inc.

Research

Transgenic Mouse Proteomics Identifies New 14-3-3-associated Proteins Involved in Cytoskeletal Rearrangements and Cell Signaling*,S

Pierre-Olivier Angrand{ddagger},§,¶, Inmaculada Segura||, Pamela Völkel{ddagger},§, Sonja Ghidelli{ddagger}, Rebecca Terry||, Miro Brajenovic{ddagger},**, Kristina Vintersten{ddagger}{ddagger},§§, Rüdiger Klein||, Giulio Superti-Furga{ddagger},¶¶, Gerard Drewes{ddagger}, Bernhard Kuster{ddagger}, Tewis Bouwmeester{ddagger} and Amparo Acker-Palmer||,||||

From the {ddagger} Cellzome AG and the {ddagger}{ddagger} European Molecular Biology Laboratory, Meyerhofstrasse 1, D-69117 Heidelberg, Germany and || Max Planck Institute of Neurobiology, Am Klopferspitz 18, D-82152 Martinsried, Germany

Identification of protein-protein interactions is crucial for unraveling cellular processes and biochemical mechanisms of signal transduction. Here we describe, for the first time, the application of the tandem affinity purification (TAP) and LC-MS method to the characterization of protein complexes from transgenic mice. The TAP strategy developed in transgenic mice allows the emplacement of complexes in their physiological environment in contact with proteins that might only be specifically expressed in certain tissues while simultaneously ensuring the right stoichiometry of the TAP protein versus their binding partners and represents a novelty in proteomics approaches used so far. Mouse lines expressing TAP-tagged 14-3-3{zeta} protein were generated, and protein interactions were determined. 14-3-3 proteins are general regulators of cell signaling and represent up to 1% of the total brain protein. This study allowed the identification of almost 40 novel 14-3-3{zeta}-binding proteins. Biochemical and functional characterization of some of these interactions revealed new mechanisms of action of 14-3-3{zeta} in several signaling pathways, such as glutamate receptor signaling via binding to homer homolog 3 (Homer 3) and in cytoskeletal rearrangements and spine morphogenesis by binding and regulating the activity of the signaling complex formed by G protein-coupled receptor kinase-interactor 1 (GIT1) and p21-activated kinase-interacting exchange factor ß (ßPIX).

To whom correspondence may be addressed. E-mail: pierre-olivier.angrand{at}ibl.fr

|||| To whom correspondence may be addressed. Tel.: 49-89-8578-3159; Fax: 49-89-8578-3152; E-mail: palmer{at}neuro.mpg.de


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