Sustained activation of protein kinase C induces delayed phosphorylation and regulates the fate of epidermal growth factor receptor.
It is well established that acute activation of members of the protein kinase C (PKC) family induced by activation of cellular receptors can transduce extracellular stimuli to intracellular signaling. However, the functions of sustained activation of PKC are not well studied. We have previously shown that sustained activation of classical PKC isoforms over 15-60 min induced the formation of the pericentrion, a subset of recycling endosomes that are sequestered perinuclearly in a PKC- and phospholipase D (PLD)-dependent manner. In this study, we investigated the role of this process in the phosphorylation of EGFR on threonine 654 (Thr-654) and in the regulation of intracellular trafficking and fate of epidermal growth factor receptor (EGFR). Sustained stimulation of the angiotensin II receptor induced translocation of the EGFR to the pericentrion, which in turn prevents full access of EGF to the EGFR. These effects required PKC and PLD activities, and direct stimulation of PKC with phorbol esters was sufficient to reproduce these effects. Furthermore, activation of PKC induced delayed phosphorylation of EGFR on Thr-654 that coincided with the formation of the pericentrion and which was dependent on PLD and endocytosis of EGFR. Thus, Thr-654 phosphorylation required the formation of the pericentrion. On the other hand, using a T654A mutant of EGFR, we find that the phosphorylation on Thr-654 was not required for translocation of EGFR to the pericentrion but was required for protection of EGFR from degradation in response to EGF. Taken together, these results demonstrate a novel role for the pericentrion in the regulation of EGFR phosphorylation, which in turn is important for the fates of EGFR.
Department of Medicine and The Stony Brook Cancer Center, Stony Brook University, Stony Brook, New York, United States of America ; Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina, United States of America.
Mengling
M
Liu
Jolanta
J
Idkowiak-Baldys
Patrick L
PL
Roddy
Aleksander
A
Baldys
John
J
Raymond
Christopher J
CJ
Clarke
Yusuf A
YA
Hannun
D002460:Cell Line; D066246:ErbB Receptors / Q000378:metabolism*; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D015151:Immunoblotting; D018613:Microscopy, Confocal; D010739:Phospholipase D / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D044140:Receptor, Angiotensin, Type 1 / Q000378:metabolism
D044140:Receptor, Angiotensin, Type 1; D066246:ErbB Receptors; D011493:Protein Kinase C; D010739:Phospholipase D