FIGURE 5.

TC-PTP recognizes GADD34 phosphorylated on tyrosine 262. Panel A, lysates of HEK293T cells expressing WT GADD34-GFP or mutant GADD34(Y262F)-GFP with or without the catalytically inactive FLAG-TC-PTP(C/S) containing a serine substitution in place of the catalytic cysteine were incubated with FLAG-M2 beads (IP) with or without 10 mm Na₃VO₄. The bound proteins were eluted with sample buffer and with lysates (Input) subjected to SDS-PAGE and immunoblotting (IB) with anti-GADD34 and anti-FLAG. Panel B, lysates of HEK293T cells expressing WT GADD34-GFP or the mutant GADD34(Y262F)-GFP either alone or with GST-PTP1B(C/S) were incubated with GST-agarose beads in the presence or absence of 1 mm Na₃VO₄. The bound proteins were eluted in sample buffer and with lysates (input) subjected to SDS-PAGE prior to immunoblotting with anti-GFP and anti-PTP1B antibodies.