c-Src phosphorylates epidermal growth factor receptor on tyrosine 845

Area of Science:

• Cell Biology
• Molecular Biology
• Biochemistry

Background:

• Previous work identified c-Src association and activation with the epidermal growth factor (EGF) receptor in A431 cells upon EGF treatment.
• The precise interaction and phosphorylation events within the c-Src-EGF receptor complex remained to be elucidated.

Purpose of the Study:

• To investigate the phosphorylation of the EGF receptor by c-Src within their complex.
• To identify the specific site(s) on the EGF receptor phosphorylated by c-Src.

Main Methods:

• Utilized a synthetic peptide (Y845 peptide) corresponding to residues 837-856 of the EGF receptor for phosphorylation studies with c-Src.
• Analyzed cyanogen bromide-digested fragments of phosphorylated c-Src-associated EGF receptor using phosphopeptide mapping.
• Confirmed phosphorylation site in vitro and in EGF-treated A431 cells.

Main Results:

• c-Src phosphorylated the synthetic Y845 peptide.
• A 7 kDa phosphopeptide was detected in vitro, which co-migrated with the phosphorylated Y845 peptide.
• The 7 kDa phosphopeptide was exclusively phosphorylated on tyrosine, indicating Y845 as the site.
• Phosphorylation of Y845 was confirmed in EGF-treated A431 cells.

Conclusions:

• c-Src directly phosphorylates the EGF receptor at tyrosine residue 845 (Y845).
• This phosphorylation event is dependent on epidermal growth factor (EGF) stimulation.
• The findings elucidate a specific mechanism of EGF receptor regulation by c-Src.