Similar requirement for clathrin in EGF- and HGF- stimulated Akt phosphorylation

ABSTRACT

Receptor tyrosine kinases, such as the epidermal growth factor (EGF) receptor (EGFR) and Met lead to activation of intracellular signals including Akt, a critical regulator of cell survival, metabolism and proliferation. Upon binding their respective ligands, each of these receptors is recruited into clathrin coated pits (CCPs) eventually leading to endocytosis. We have recently shown that phosphorylation of Gab1 and Akt following EGFR activation requires clathrin, but does not require receptor endocytosis. We examined whether clathrin regulates Akt signaling downstream of Met, as it does for EGFR signaling. Stimulation with the Met ligand Hepatocyte Growth Factor (HGF) leads to enrichment of phosphorylated Gab1 (pGab1) within CCPs in ARPE-19 cells. Perturbation of clathrin using the inhibitor pitstop2 decreases HGF-stimulated Akt phosphorylation. These results indicate that clathrin may regulate Met signaling leading to Akt phosphorylation similarly as it does for EGFR signaling.

KEYWORDS:

• clathrin
• endocytosis
• membrane traffic
• phosphorylation
• receptor tyrosine kinase
• signaling adaptor
• signaling scaffold
• signaling microdomain
• TIRF microscopy

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.

Funding

Funding for this research was provided by an Operating Grant from the Canadian Institutes of Health Research to C.N.A. (grant no. 125854) and by Ryerson University.