Fig. 2
From: Enhanced breast cancer progression by mutant p53 is inhibited by the circular RNA circ-Ccnb1
circ-Ccnb1 interacts with H2AX. a The circ-Ccnb1 probe pulled-down proteins were subject to proteomic assay. In the Beas2B cells, H2AX and p53 were precipitated. In the HTB126 cells, H2AX and pBclaf1 were precipitated. The last two columns represent the number of pulled down peptides detected by the system. b Lysates prepared from vector- or circ-Ccnb1-transfected HTB126 and Beas2B cells were subject to probe pull-down followed by Western blotting. The circ-Ccnb1 probe precipitated p53 in Beas2B cells. c In lysates prepared from vector- or circ-Ccnb1-transfected HTB126 and MB231 cells, the probe could not precipitate p53, but could precipitate Bclaf1. Ectopic expression of circ-Ccnb1 increased Bclaf1 precipitation. d Lysate prepared from 293T cells was incubated with antibodies as shown, followed by real-time PCR for levels of linear Ccnb1 mRNA and circ-Ccnb1. Anti-H2AX, p53, γH2AX, and Bclaf1 antibodies pulled-down circ-Ccnb1, but not linear Ccnb1 mRNA. **p < 0.01. Error bars, SD (n = 4). e Models depicting circ-Ccnb1 interacting with H2AX, p53 and Bclaf1. f Prediction of probable RNA-binding residues of H2AX was carried out by submitting the H2AX sequence to Pprint, RNABindRPlus and RBscore servers. “+” indicates the predicted RNA-binding residues. g Graphical representation of three-dimensional structures of the docking models of circ-Ccnb1 with the binding fragment of H2AX by NPDock
