A Structural Model for the Membrane-bound Form of the Juxtamembrane Domain of the Epidermal Growth Factor Receptor

Kiattawee Choowongkomon,Cathleen R Carlin,Frank D Sönnichsen

doi:10.1074/jbc.m502698200

Kiattawee Choowongkomon, Cathleen R Carlin + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m502698200

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Abstract

The epidermal growth factor receptor (EGFR) is a member of the receptor tyrosine kinase family involved in the regulation of cellular proliferation and differentiation. Its juxtamembrane domain (JX), the region located between the transmembrane and kinase domains, plays important roles in receptor trafficking. Two sorting signals, a PXXP motif and a 658LL659 motif, are responsible for basolateral sorting in polarized epithelial cells, and a 679LL680 motif targets the ligand-activated receptor for lysosomal degradation. To understand the regulation of these signals, we characterized the structural properties of recombinant JX domain in aqueous solution and in dodecylphosphocholine (DPC) detergent. JX is inherently unstructured in aqueous solution, albeit a nascent helix encompasses the lysosomal sorting signal. In DPC micelles, structures derived from NMR data showed three amphipathic, helical segments. A large, internally inconsistent group of long range nuclear Overhauser effects suggest a close proximity of the helices, and the presence of significant conformational averaging. Models were determined for the average JX conformation using restraints representing the translational restriction due to micelle-surface adsorption, and the helix orientations were determined from residual dipolar couplings. Two equivalent average structural models were obtained that differ only in the relative orientation between first and second helices. In these models, the 658LL659 and 679LL680 motifs are located in the first and second helices and face the micelle surface, whereas the PXXP motif is located in a flexible helix-connecting region. The data suggest that the activity of these signals may be regulated by their membrane association and restricted accessibility in the intact receptor.

Highlights

The epidermal growth factor receptor (EGFR) is a member of the receptor tyrosine kinase family involved in the regulation of cellular proliferation and differentiation
EGFR645– 697 in DPC Micelles—Because the EGF receptor juxtamembrane domain (JX) region is in close proximity to biomembranes, we investigated how the lipid microenvironment affected the structure of this domain
In EGFR, tyrosine kinase activity has been reported to be regulated by the JX domain despite the absence of an inhibitory tyrosine

Summary

Introduction

The epidermal growth factor receptor (EGFR) is a member of the receptor tyrosine kinase family involved in the regulation of cellular proliferation and differentiation. The epidermal growth factor receptor (EGFR) is the prototypic member of the ErbB family of tyrosine kinase receptor molecules It plays an important role in cell differentiation, proliferation, and epithelial organogenesis [1]. Several intrinsic sorting signals and protein binding sites have been mapped to the JX region, which regulates receptor trafficking and inactivation These include basolateral sorting signals [10], a lysosomal sorting motif [11], a nuclear localization signal [12], as well. The JX region includes post-translational modification sites; Thr654 is a known substrate for PKC [16], and Thr669 and Ser671 are substrates for MAPK [17, 18]

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