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[Preprint]. 2023 Oct 4:rs.3.rs-3270539. [Version 1] doi: 10.21203/rs.3.rs-3270539/v1

Figure 5. REST is a novel epigenetic target of EZH2 and it reverses EZH2-induction of NE markers.

Figure 5

(A) C4-2 and 144-13 cells were treated with DMSO or 10uM GSK126 for 48hr. Whole cell lysate was analyzed by Western blotting. (B) Western blotting of indicated proteins in C4-2 and NE1.3 cells expressing scramble shRNA or shEZH2. (C) RT-qPCR showing that EZH2-mediated REST downregulation and SYP upregulation was reversed by shEZH2 (genetic rescue). (D) 144-13 NEPC cells were treated with DMSO or 10uM GSK126 for 48hr, followed by ChIP with control IgG or anti-H3K27me3 antibody, and qPCR of REST’s promoter region near its transcriptional starting site. Y-axis represents % of ChIPed DNA relative to input. (E) Scatter plots showing negative correlation of the expression of EZH2 and REST in the Beltran_NM2016 CRPC genomic dataset. The plot, Pearson correlation coefficient R and P value were directly downloaded from the cBioPortal genomics interface. (F)A genetic rescue experiment in LNCaP cells indicates that the induction of NE markers by EZH2 was reversed by additional overexpression of REST.