Figure 5.

PTRF is phosphorylated at multiple sites. (A) 2D gel electrophoresis of PTRF. Nuclear extract proteins were separated by 2D electrophoresis, and PTRF was detected on western blots using anti-PTRF antibodies. (B) Tryptic phosphopeptide map of PTRF labeled in vivo. NIH 3T3 cells were metabolically labeled with ³²P-orthophosphate, PTRF was immunoprecipitated, resolved by 10% SDS–PAGE and detected by autoradiography (lane 1). In parallel, PTRF was immunoprecipitated from unlabeled cells and visualized on immunoblots with anti-PTRF antibodies (lane 2). (C) Radiolabeled PTRF was digested with trypsin and subjected to 2D peptide mapping (right panel).