Crystal structure of the p14/MP1 scaffolding complex: How a twin couple attaches mitogen-activated protein kinase signaling to late endosomes

doi:10.1073/pnas.0403435101

Crystal structure of the p14/MP1 scaffolding complex: How a twin couple attaches mitogen-activated protein kinase signaling to late endosomes

^†Institute for Molecular Pathology, Dr. Bohr-Gasse 7, A-1030 Vienna, Austria; ^§Department for Histology and Molecular Cell Biology, Medical University of Innsbruck, Müllerstrasse 59, A-6020 Innsbruck, Austria; ^¶Max-Planck-Arbeitsgruppen für Strukturelle Molekularbiologie, Deutsches Elektronen-Synchrotron, Notkestrasse 85, D-22603 Hamburg, Germany; and ^∥Institute for Medical Radiation and Cell Research, University of Würzburg, Würzburg 97078, Germany

Communicated by Robert Huber, Max Planck Institute for Biochemistry, Martinsried, Germany, May 24, 2004 (received for review March 24, 2004)

Abstract

Signaling pathways in eukaryotic cells are often controlled by the formation of specific signaling complexes, which are coordinated by scaffold and adaptor proteins. Elucidating their molecular architecture is essential to understand the spatial and temporal regulation of cellular signaling. p14 and MP1 form a tight (K _d = 12.8 nM) endosomal adaptor/scaffold complex, which regulates mitogen-activated protein kinase (MAPK) signaling. Here, we present the 1.9-Å crystal structure of a biologically functional p14/MP1 complex. The overall topology of the individual MP1 and p14 proteins is almost identical, having a central five-stranded β-sheet sandwiched between a two-helix and a one-helix layer. Formation of the p14/MP1 heterodimer proceeds by β-sheet augmentation and yields a unique, almost symmetrical, complex with several potential protein-binding sites on its surface. Mutational analysis allowed identification of the p14 endosomal adaptor motif, which seems to orient the complex relative to the endosomal membrane. Two highly conserved and hydrophobic protein-binding sites are located on the opposite “cytoplasmic” face of the p14/MP1 heterodimer and might therefore function as docking sites for the target proteins extracellular regulated kinase (ERK) 1 and MAPK/ERK kinase 1. Furthermore, detailed sequence analyses revealed that MP1/p14, together with profilins, define a protein superfamily of small subcellular adaptor proteins, named ProflAP. Taken together, the presented work provides insight into the spatial regulation of MAPK signaling, illustrating how p14 and MP1 collaborate as an endosomal adaptor/scaffold complex.

Footnotes

↵ †† To whom correspondence may be addressed. E-mail: lukas.a.huber{at}uibk.ac.at or clausen{at}imp.univie.ac.at.
↵ ‡ R.K. and D.T. contributed equally to this work.
Abbreviations: MAPK, mitogen-activated protein kinase; EGF, epidermal growth factor; KSR, kinase suppressor of Ras; ERK, extracellular regulated kinase; MEK, MAPK/ERK kinase; MP1, MEK1 partner; MP1-myc, myc-tagged MP1.
Data deposition: The atomic coordinates and structure factors for the two p14/MP1 complex structures have been deposited in the Protein Data Bank, www.pdb.org (PDB ID codes 1VET and 1VEU, respectively).