Fig. 4a–c.

Identification of Src-dependent phosphorylation sites among mitochondrial proteins. Mitochondria were treated as described in the legend of Fig. 1. Proteins were then solubilized with n-dodecyl-maltoside (2.5 mg/mg protein), and 100 to 400 μg per lane was subjected to BN-PAGE (5–13%). a The BN-PAGE was directly transferred to PVDF membranes, and tyrosine-phosphorylated proteins were detected with an antibody to phosphotyrosine (PY20). Membranes were then stripped and reprobed with antibodies against one subunit of each OxPhos complex. b Individual lanes corresponding to each treatment (CTL, ATP, ATP + PP2, ATP + PTP1Bi) were cut, soaked with 1% sodium dodecyl sulfate (w/v) and 1% (v/v) β-mercaptoethanol, and subjected to a second-dimension SDS-PAGE (12.5%) for immunodetection of tyrosine phosphorylation. For each lane, one second-dimension gel was kept for Coomassie blue staining. c Spots (arrows) detected both in the Coomassie blue-stained gel and in the immunoblot corresponding to the ATP treatment were analyzed by LC-MS/MS. Data are representative of at least three experiments