CUT&RUN sequencing of wild-type and ZNF217-knockdown KOPN-8 cells

Here, we elucidate the oncogenic role of ZNF217 in B-ALL and reveal that ZNF217 interacts with the CoREST complex to mediate histone modifications such as H3K4me1, H3K4me2, and H3K27ac. This interaction partially contributes to ZNF217’s oncogenic function in B-ALL. To identify genes directly bound by ZNF217, we conducted high-throughput CUT&RUN sequencing using an anti-ZNF217 antibody in both wild-type and ZNF217-knockdown KOPN-8 cells. Additionally, to distinguish between CoREST-dependent and CoREST-independent targets, we performed CUT&RUN sequencing in the same cell models using an antibody against LSD1, a core component of the CoREST complex that interacts with ZNF217, as well as antibodies against H3K4me1, H3K4me2, and H3K27ac. KOPN-8 cells transduced with either a ZNF217-targeting shRNA or a scrambled shRNA control were used in CUT&RUN assays. In the ZNF217 and LSD1 groups, live cells were used, while in the H3K4me1, H3K4me2, and H3K27ac groups, cells were lightly cross-linked with 0.1% formaldehyde for 1 minute. The antibodies employed included anti-ZNF217 (A303-265A, Thermo Fisher Scientific), anti-LSD1 (2139S, Cell Signaling Technology), anti-H3K4me1 (5326T, Active Motif), anti-H3K4me2 (9725T, Active Motif), and anti-H3K27ac (39034, Active Motif). For each sample, 10 ng of CUT&RUN-enriched DNA was used for library construction.