Fig. 6: Identification of direct target genes of ANLN.

A Flowchart of the process used to identify the genes directly targeted by ANLN. B Heatmap showing the RNA-Seq expression and CUT&Tag signals of 84 direct target genes. C ChIP-qPCR experiments measuring ANLN and Pol II binding enrichment on promoter regions of target genes. n = 3 for each group. The error bars indicate the mean ± SDs. D Metascape database was used for enrichment analysis of 84 target genes. Based on the selected terms, accumulative hypergeometric P-values and enrichment factors were calculated and used for filtering. Remaining significant terms were hierarchically clustered into a tree based on Kappa-statistical similarities among their gene memberships. The 0.3 kappa score was applied as the threshold to cast the tree into term clusters. E Representative tracks of target genes showing the occupancy of ANLN and Pol II. F Relative mRNA levels of target genes after ANLN knockdown in KYSE150 cells were measured by RT-qPCR. n = 3 for each group. The error bars indicate the mean ± SDs. Two-tailed Student’s t tests were performed to determine the significance. G H3K27ac ChIP-Seq data from SEdb database were used to identify the super-enhancer regions of representative target genes, and the graph shows the information of each super-enhancer regions. H Representative tracks of super-enhancer regions occupancy of ANLN, Pol II and H3K72ac. I Screen potential transcription factors from the Cistrome database and ANLN interactome. J Occupancy of ANLN, Pol II and different transcription factors in the promoter and super-enhancer regions of representative target gene. K A hypothetical model: ANLN, Pol II and different transcription factors form an active transcriptional cluster at super-enhancer regions. CUT&Tag analysis representative of two independent experiments, and other data are representative of at least three independent experiments with similar results. Source data are provided as a Source Data file.