Figure 3. Panx1 and Panx3 are post-translationally modified by S-nitrosylation. Western blots for Panx1 (A), Panx2 (B), and Panx3 (C) following an assay for detection of S-nitrosylated proteins. HEK-293T cells ectopically expressing murine Panx1, Panx2, or Panx3 were treated with 100 µM GSNO to facilitate S-nitrosylation and samples were processed by the Biotin Switch assay. For controls, Panx expressing cells were stimulated with 100 µM reduced GSH, the backbone molecule in GSNO that lacks the capacity to release NO, or with the reducing agent DTT (1mM) following GSNO treatment to reduce S-nitrosylated protein thiols. As a negative control, ascorbate was omitted from the reducing step to prevent reduction of S-nitrosylated cysteines and subsequent biotinylation. GAPDH was used as input controls. Protein sizes are noted in kDa.