Fig. 6: Functional importance of ANLN super-enhancer in TNBC clonogenicity.

a Higher expression of ANLN is associated with poor prognosis of breast cancer patients in the METABRIC cohort. The statistical significance was calculated by a log-rank test (one-sided). Increased expression of ANLN in TNBC samples (n = 327), compared to other subtypes (Her2 n = 237, LumA n = 706, LumB n = 484, and Normal-like n = 199; two-sided Wilcoxon signed-rank tests). The boxes represent the 25th percentile, median, and 75th percentile, whiskers were extended to the furthest value that is no more than 1.5 times the inter-quartile range. b Immunoblotting detection of ANLN expression in a panel of breast cancer cell lines, repeated independently twice with similar results. c–e H3K27ac (c), Brd4 (d), and P300 (e) ChIP-qPCR of indicated cell lines using primer amplifying e1 of ANLN SE. n of independent experiments is indicated by scatter dots. f mRNA expression levels of ANLN, data from Cancer RNA-seq Nexus. TNBC n = 42 samples, Normal tissue n = 21 samples adjacent to TNBC. g Schematic of ANLN SE (SSE256) and detection of e1 deletion of ANLN SE by PCR in Hs578T, MDA-MB-231 and BT549 cells, repeated independently twice with similar results. h Immunoblotting detection of ANLN upon deletion of SSE256 e1 region, repeated independently twice with similar results. i Clonogenic assay of Hs578t and BT549 cells with or without deletion of SSE256 e1. n = 3 independent experiments. Data are represented as mean ± SEM in (c–e) and (i). P-values were calculated by one-sided Student’s t test in (c–e). P-values were calculated by two-sided Student’s t test in (i). Source data are provided as a Source Data file.