Fig. 6.

CBS protects STAT3 from oxidation induced loss of transcriptional activity. (A). Quantification of STAT3 phosphorylation at Tyr 705 by Western blot analyses. In our quantification we determined the ratios between STAT3/GAPDH and p-STAT3/GAPDH and then determined the ratio between (p-STAT3/GAPDH)/(STAT3/GAPDH) and finally calculate the % of control. The data are presented as mean ± SEM of 7 independent experiments, ***P < 0.001, ****P < 0.0001 compared to control. (B). Western blot analyses of STAT3 oligomerization in the presence and absence of H₂O₂ (100 μM for 2 min) after separation by SDS-PAGE gel electrophoresis in control and shCBS T3M4 as well as in BxPC3 cells. Image is representative of 3 independent experiments. (C). Visualization of STAT3 disulfide-linked dimers after separation by two dimensional sequential nonreducing-reducing SDS-PAGE and immunoblotting of the diagonal gel with a STAT3 specific antibody. Arrows indicate intermolecular disulfide linked STAT3 oligomers. Image is representative of 3 independent experiments. (D). Quantification of oxidized STAT3 oligomers. The data are presented as mean ± SEM of 4 independent experiments, *P < 0.05 compared to control.