Fig. 8.

Tsg101 interacts with p62 through the PRR domain. (A) Schematic diagram of plasmid structures for Tsg101 full-length and its truncated mutations. (B) Representative immunoblots showing the expression of co-transfected Tsg101 mutants and Flag-p62 in HEK-293T cells. (C) Co-immunoprecipitations (Co-IP) with anti-Flag antibody, and immunoblotting with anti-Myc antibody in HEK-293T cells co-transfected with indicated Tsg101 mutants and Flag-p62. Similar results were observed in three independent experiments. HC stands for non-specific IgG heavy-chain signal bands. (D) Co-IP with anti-Myc antibody, and immunoblotting with anti-Flag antibody in HEK-293T cells co-transfected with indicated Tsg101 mutants and Flag-p62 plasmid. Similar results were observed in three independent experiments. (E) Representative images and (F) quantification analysis of DHE staining in HEK-293T cells transfected with Tsg101-full length (FL) and its PRR-deletion plasmids. *, p < 0.05. Similar results were observed in three independent experiments. (G) Cell survival rate was determined by MTS incorporation in HEK-293T cells transfected with Tsg101-FL and its PRR-deletion plasmids, followed by treatment with 500 μM H₂O₂ for 12 h *, p < 0.05. Similar results were observed in three independent experiments. (H) A diagram showing the structures of p62-FL and its truncated mutations. (I) Representative immunoblots showing the expression of indicated Flag-p62 mutants and mCherry-Tsg101 in HEK-293T cells. (J) Co-IP with mCherry antibody and immunoblotting with Flag antibody in HEK-293T cells co-transfected with indicated Flag-p62 mutants and mCherry-Tsg101. Similar results were observed in three independent experiments. HC stands for non-specific IgG heavy-chain bands. (K) Co-IP with anti-Flag antibody, and immunoblotting with mCherry antibody in HEK-293T cells co-transfected with indicated Flag-p62 mutants and mCherry-Tsg101. Similar results were observed in three independent experiments. HC stands for non-specific IgG heavy-chain bands.