Role of ubiquitination in IGF-1 receptor signaling and degradation.
BACKGROUND
The insulin-like growth factor 1 receptor (IGF-1R) plays numerous crucial roles in cancer biology. The majority of knowledge on IGF-1R signaling is concerned with its role in the activation of the canonical phosphatidyl inositol-3 kinase (PI3K)/Akt and MAPK/ERK pathways. However, the role of IGF-1R ubiquitination in modulating IGF-1R function is an area of current research. In light of this we sought to determine the relationship between IGF-1R phosphorylation, ubiquitination, and modulation of growth signals.


METHODS
Wild type and mutant constructs of IGF-1R were transfected into IGF-1R null fibroblasts. IGF-1R autophosphorylation and ubiquitination were determined by immunoprecipitation and western blotting. IGF-1R degradation and stability was determined by cyclohexamide-chase assay in combination with lysosome and proteasome inhibitors.


RESULTS
IGF-1R autophosphorylation was found to be an absolute requirement for receptor ubiquitination. Deletion of C-terminal domain had minimal effect on IGF-1 induced receptor autophosphorylation, however, ubiquitination and ERK activation were completely abolished. Cells expressing kinase impaired IGF-1R, exhibited both receptor ubiquitination and ERK phosphorylation, however failed to activate Akt. While IGF-1R mutants with impaired PI3K/Akt signaling were degraded mainly by the proteasomes, the C-terminal truncated one was exclusively degraded through the lysosomal pathway.


CONCLUSIONS
Our data suggest important roles of ubiquitination in mediating IGF-1R signaling and degradation. Ubiquitination of IGF-1R requires receptor tyrosine kinase activity, but is not involved in Akt activation. In addition we show that the C-terminal domain of IGF-1R is a necessary requisite for ubiquitination and ERK phosphorylation as well as for proteasomal degradation of the receptor.
Sehat
Bita
B
Department of Oncology and Pathology, Cancer Centre Karolinska (CCK), Karolinska Institutet and Karolinska University Hospital-Solna, Stockholm, Sweden.
Andersson
Sandra
S
Vasilcanu
Radu
R
Girnita
Leonard
L
Larsson
Olle
O
D000818:Animals; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006868:Hydrolysis; D047468:Immunoprecipitation; D051379:Mice; D018345:Mice, Knockout; D010766:Phosphorylation; D017526:Receptor, IGF Type 1 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D054875:Ubiquitination
D017526:Receptor, IGF Type 1; D048049:Extracellular Signal-Regulated MAP Kinases