Figure 3.

p130 compensates for the absence of Rb in mediating the transcriptional repression activity of AR

(A and B) ChIP-seq analysis of p130 was performed in uninduced C4-2-tet-shRB cells (no doxycycline). The Venn diagram for p130 and Rb (A) or p130 and E2F1 (B) overlapping binding peaks was shown. (C) ChIP-seq analysis of p130 was performed in C4-2-tet-shRB cells treated with vehicle or doxycycline (3 d). The Venn diagram for p130 binding peaks in both conditions was shown. (D) BETA for the association of p130 binding peaks (doxycycline treated) with the expression of Rb-repressed genes identified from RNA-seq of C4-2-tet-shRB (doxycycline versus vehicle) is shown. (E) Immunoprecipitation (IP) of E2F1 in C4-2-tet-shRB cells (doxycycline for 0, 3, or 30 d), followed by immunoblotting for p130 or E2F1, is shown. (F and G) IP of p130 (F) or E2F1 (G) in C4-2-tet-shRB cells cultured with or without doxycycline and treated with vehicle or palbociclib (1μM, 24 h), followed by immunoblotting for total p130, p-p130 (S672 phosphorylated), or E2F1 is shown. (H) BETA for the association of p130 binding peaks (doxycycline treated) with the expression of AR-repressed genes identified from RNA-seq of C4-2-tet-shRB (doxycycline treated) is shown. (I–K) C4-2-tet-shRB cells stably infected by lentiviral shRNAs against NTC or RBL2 were treated with vehicle or doxycycline (6 d), followed by immunoblotting for Rb, p130, and E2F1 (I), quantitative real-time PCR for E2F-regulated genes (J), and the flow cytometry cell cycle analysis (K). Data in bar graphs represent the mean ± SD. n.s., not significant. ∗p < 0.05.