Fig 2: CBL0137 induces partial dissociation of CTCF from its binding sites. a Western blot analysis of CTCF, Rad21 (cohesin subunit), and SMC2 (condensin subunit) in nuclear extracts prepared from control and CBL0137-treated (3 µM, 6 h) HT1080 cells. Histone H3 was used as a loading control. b Western blot analysis of CTCF, Rad21, SMC2, SPT16 (FACT subunit), and histone H3 in different chromatin fractions from control and CBL0137-treated (3 µM, 6 h) HT1080 cells prepared by sequential extraction with 0.1 and 0.4 M NaCl. Pellet (Pt): insoluble chromatin fraction. c Representative 1.1 Mb-long genomic segment from chromosome 3 containing two distinct chromatin loop domains bordered by CTCF binding sites in control cells. Corresponding Hi-C maps, CTCF peaks determined using the PePr computational approach[43], and an enlarged view of the particular groups of CTCF peaks are shown for control and CBL0137-treated HT1080 cells. d Bar plots showing the total number of CTCF peaks mapped by PePr in intact HT1080 cells, preserved after CBL0137 treatment, and the newly formed CTCF peaks in CBL0137-treated cells. e Bar plots showing the number of CTCF differential binding sites identified in control and CBL0137-treated HT1080 cells. Analysis of differential binding sites was performed using PePr43, which showed the number of peaks mapped only in one of the two samples (either control or CBL0137-treated). Source data of Fig. 6a, b are provided in a Source Data file

Fig 3: CDK7 is necessary to maintain expression of genes that control mitotic fidelity including those comprising condensin.A, Venn diagram of genes that are differentially downregulated (left) or upregulated (right) in MDA-MB-231 and MDA-MB-468 cells treated with THZ1 (75 nM and 50 nM, respectively) compared with vehicle (DMSO). B, Gene Set Enrichment Analysis of genes in the cell cycle G2/M phase was evaluated in MDA-MB-231 (left) and MDA-MB-468 (right) cells. Normalized enrichment score (NES) and false discovery rate (FDR) are shown. C, table of significantly downregulated Gene Ontology pathways as assessed by evaluating the shared 1367 significantly downregulated genes in MDA-MB-231 and MDA-MB-468 cells. D, volcano plot of significant differentially expressed genes (DEGs) in MDA-MB-231 cells. Genes comprising the condensin complex are indicated by red dots with labels. E, representative Western blot of SMC2 protein in MDA-MB-231 cells treated with vehicle (DMSO), THZ1 (75 nM), or CT7001 (CT7, 750 nM). Values are the average of three replicates representing SMC2/β-actin. F, RT-PCR validation of condensin gene suppression in MDA-MB-231 cells treated as in E. G and H, same as E and F but using MDA-MB-468 cells treated with vehicle or THZ1 (50 nM). I and J, same as E and F but using HCC38 cells treated with vehicle or THZ1 (50 nM). K, RT-PCR validation of condensin gene suppression in HCC38 cells treated with CT7001 (CT7, 750 nM). L and M, MDA-MB-231 Cas9-expressing cells were transfected with nonspecific sgRNA (NTS) or sgRNA targeted to CDK7 (sgCDK7 #1, sgCDK7 #2) for 72 h. L, representative blot of SMC2 protein expression following CDK7 gene disruption. Numbers represent relative SMC2/β-actin ratios. β-Actin blot is reused from Figure 3A. This Western was probed for both SMC2 and CDK7 (to confirm CRISPR-mediated knockout of CDK7) with β-actin as the loading control for each. M. quantitation of SMC2 mRNA relative to GAPDH 72 h after transfection of MDA-MB-231 Cas9 cells with sgCDK7 versus nontargeting control. Bars are means ± SEM. At least three biological replicates were completed per experiment in duplicate. ∗ = p < 0.05.