Metastasis suppressor tetraspanin CD82/KAI1 regulates ubiquitylation of epidermal growth factor receptor

Ligand induced ubiquitylation of Epidermal Growth Factor Receptor (EGFR) is an important regulatory mechanism which controls endocytic trafficking of the receptor and its signalling potential. Here we report that tetraspanin CD82/KAI1 specifically suppresses ubiquitylation of EGFR after stimulation with Heparin-Binding Epidermal Growth Factor (HBEGF) or amphiregulin (AR) and alters the rate of recruitment of the activated receptor to EEA1-positive endosomes. The suppressive effect of CD82 is dependent on the heparin-binding domain of the ligand. Deletion of the C terminal cytoplasmic domain of CD82 (CD82ΔC mutant) inhibits endocytic trafficking of the tetraspanin and compromises its activity towards HBEGF activated EGFR. Reduced ubiquitylation of EGFR is accompanied by PKC dependent increase in serine phosphorylation of cCbl in cells expressing elevated levels of CD82. Furthermore, phosphorylation of Threonine 654 (PKC phosphorylation site) in juxtamembrane domain of the receptor is considerably increased in CD82 expressing cells. These results describe previously unsuspected links between tetraspanin proteins and ubiquitylation of their molecular partners (e.g. EGFR). Our data identify CD82 as a new regulator of cCbl, which discriminatively controls the activity of this E3 ubiquitin ligase towards heparin-binding ligand EGFR pairs. Taken together these observations provide an important new insight into the modulatory role of CD82 in endocytic trafficking of EGF receptor.