Mutation of Di-leucine residues in the juxtamembrane region alters EGF receptor expression.
Di-leucine motifs have been implicated in the internalization or degradation of many membrane proteins. The epidermal growth factor receptor (EGFR) contains two di-leucine residues at 658 (TLRRLLQER) and 679 (NQALLRIL). To determine the role of these di-leucine motifs in regulating EGF receptor expression, activity, or ligand-induced degradation, the di-leucine residues at positions 658 or 679 were mutated to di-alanine residues, and the mutant receptors were stably expressed in CHO cells. The results indicate that mutation of either di-leucine motif generates and promotes cell surface expression of carboxy-truncated EGF receptors (M(r) 120, 140 kDa) that do not undergo EGF-induced autophosphorylation or degradation. In contrast, full-length EGF receptors (170 kDa) containing di-alanine substitutions resemble wild type receptors in that they respond to EGF by autophosphorylation, their tyrosine kinase activity is inhibited by protein kinase C, and they are degraded. The level of autophosphorylation of the 170 kDa mutant receptors and EGF-induced tyrosine phosphorylation of other cellular proteins is lower than that of the wild type receptor, consistent with formation of kinase-inactive heterodimers between the truncated and full-length mutant receptors. These results demonstrate that removal of either of the di-leucines leads to generation of inactivating carboxy-truncated receptors, suggesting that the two di-leucine motifs within the juxtamembrane region of the EGFR are important for ensuring normal receptor expression.
Morrison
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Ben May Institute for Cancer Research, Department of Molecular Genetics and Cell Biology, University of Chicago, Illinois 60637, USA.
Chung
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Rosner
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D000409:Alanine; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D066246:ErbB Receptors / Q000096:biosynthesis* / Q000235:genetics; D007930:Leucine; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D013329:Structure-Activity Relationship
D066246:ErbB Receptors; D007930:Leucine; D000409:Alanine