Fig. 2: USP29 interacts with Snail1 and requires catalytic USP ___domain to stabilize Snail1.

a HEK293T cells were co-transfected with Flag-Snail1 vector along with Flag-HA-GFP, Flag-HA-USP28, or Flag-HA-USP29 constructs for 24 h. Cells were treated with cycloheximide (100 μg/ml) to stop protein synthesis for indicated times and lysed for Western blotting assays with indicated antibodies. b turnover curves show the relative amounts of Snail1 at each time point from (a) (n = 3). c schematic illustration of the ___domain structure in USP29 and corresponding truncations with indicated amino acid sequences. d Flag-Snail1 and various GFP-USP29 constructs were co-transfected into HEK293T cells for 24 h, and then cell lysates were analyzed with Western blotting using indicated antibodies. Below column chart shows the relative quantification of Snail1 levels (n = 3). e HEK293T cells were co-transfected with indicated combinations of Flag-Snail1 and diverse USP29 constructs for 24 h. GFP or GFP-tagged USP29 variants were immunoprecipitated with anti-GFP antibodies. Samples were processed for Western blotting assays to probe for co-immunoprecipitated Snail1. f Flag-tagged Snail1 was immunoprecipitated from HEK293T cell lysates following co-transfections of Flag-Snail1 with pEGFP, pEGFP-USP28, or pEGFP-USP29-USP constructs. Immunoprecipitates were analyzed by Western blotting using indicated antibodies. All error bars represent the standard error of the mean (SEM), with *p value < 0.05 and n.s. representing not significant.