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. Author manuscript; available in PMC: 2021 Jun 18.
Published in final edited form as: Mol Cell. 2020 May 23;78(6):1114–1132.e10. doi: 10.1016/j.molcel.2020.04.034

Figure 4. EN1 functions similarly to BRD4-S in Upregulating Matrisome Gene Expression and Interacts Preferentially with BRD4-S in MDA-MB-231 Cells.

Figure 4.

(A) 70 TF motifs identified from H3K27ac-enriched and IDC-associated BRD4-S-upregulated type II cystatin gene cluster and CTSW/MUC5 loci by MISP (Cistrome).

(B) Heatmap of 73 TF expression profiles classified according to PAM50 subtypes. Heatmap scale represents mRNA mean. Asterisks (*) mark the three TFs not enriched in IDC used as controls.

(C) Sequence logos for EN1 (MA0027.2) and EN2 (MA0642.1).

(D) RT-qPCR analysis of TF RNA. Data are mean ± s.d. P, two-tailed t-test.

(E) Knockdown of EN1 reduces S-Up CST1/2/4, MUC5AC/B and CTSW/Z RNAs as seen with BRD4-S knockdown. Data are mean ± s.d. P, two-tailed t-test.

(F) BRD4-S upregulates, while BRD4-L downregulates, matrisome gene expression in 3D culture. Data are mean ± s.d. P, two-tailed t-test.

(G) IP-IB showing EN1 preferentially interacts with BRD4-S in MDA-MB-231.

(H) EN1 interacts with both BRD4 isoforms better than EN2 using purified FLAG-tagged (f:) proteins as indicated.

(I) EN1 binds both BRD4 isoforms in SUM159 and MDA-MB-468. IP was similarly performed as in (G).