Figure 2

Role of A2BP1 in cocaine responses. (A) Co-immunoprecipitation shows A2BP1 western blotting of H3K4me3 or IgG pulldown of whole NAc lysate. Two repeated cocaine-treated samples and two saline control samples are shown. (B) The two Venn diagrams show overlap between: A2BP1-motif containing genes versus H3K4me3 differential site-containing genes (upper); A2BP1 motif- and H3K4me3 differential site-containing genes versus cocaine-regulated differential and spliced genes (lower). The table shows the top five enriched functional terms of the 478 overlapped genes from the lower Venn diagram. (C) Western blot analysis of A2BP1 in nuclear lysates of NAc after repeated cocaine. Error bars are mean ± standard error of the mean (SEM) derived from eight cocaine treated and eight saline treated control mice, respectively. (D) Conditioned place preference (CPP) scores of AAV-Cre-GFP-injected A2bp1^loxP/loxP mice (cre) with AAV-GFP-injected A2bp1^loxP/loxP mice as control (con). Error bars are mean ± SEM derived from seven cre and eight control samples. (E) Four predicted A2BP1 target candidate genes were chosen for Nanostring validation. All show the same direction of transcription change after chronic cocaine treatment as observed in RNA-seq. Error bars are mean ± SEM derived from 14 cocaine and 14 saline treated samples. (F) Cocaine-induced transcription change as observed in (E) are lost (Rps6kb2, Zfp26, Dvl1) or in one case reversed (Ece2) when A2bp1 was conditionally knocked down by using AAV-Cre viral injection in NAc. Error bars are mean ± SEM derived from five cocaine and six saline treated samples. *P < 0.05, **P < 0.01.