Epigenetic silencing of tumor suppressor Par-4 promotes chemoresistance in recurrent breast cancer
Nathaniel W. Mabe, … , J. Will Thompson, James V. Alvarez
Nathaniel W. Mabe, … , J. Will Thompson, James V. Alvarez
Published August 27, 2018
Citation Information: J Clin Invest. 2018;128(10):4413-4428. https://doi.org/10.1172/JCI99481.
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Epigenetic silencing of tumor suppressor Par-4 promotes chemoresistance in recurrent breast cancer

Abstract

Tumor relapse is the leading cause of death in breast cancer, largely due to the fact that recurrent tumors are frequently resistant to chemotherapy. We previously reported that downregulation of the proapoptotic protein Par-4 promotes tumor recurrence in genetically engineered mouse models of breast cancer recurrence. In the present study, we examined the mechanism and functional significance of Par-4 downregulation in recurrent tumors. We found that epithelial-to-mesenchymal transition (EMT) promotes epigenetic silencing of Par-4 in recurrent tumors. Par-4 silencing proceeded through binding of the EMT transcription factor Twist to the Par-4 promoter, where Twist induced a unique bivalent chromatin domain. This bivalent configuration conferred plasticity at the Par-4 promoter, and Par-4 silencing could be reversed with pharmacologic inhibitors of Ezh2 and HDAC1/2. Using an epigenome editing approach to reexpress Par-4 by specifically reversing the histone modifications found in recurrent tumors, we found that Par-4 reexpression sensitized recurrent tumors to chemotherapy in vitro and in vivo. Upon reexpression, Par-4 bound to the protein phosphatase PP1, caused widespread changes in phosphorylation of cytoskeletal proteins, and cooperated with microtubule-targeting drugs to induce mitotic defects. These results identify Twist-induced epigenetic silencing of Par-4 as a targetable axis that promotes chemoresistance in recurrent breast cancer.

Authors

Nathaniel W. Mabe, Douglas B. Fox, Ryan Lupo, Amy E. Decker, Stephanie N. Phelps, J. Will Thompson, James V. Alvarez

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Figure 5

Par-4 is epigenetically repressed in recurrent tumors through bivalent histone modifications.

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Par-4 is epigenetically repressed in recurrent tumors through bivalent h...
(A) Methylation of CpG dinucleotides (circles) within the region surrounding the TSS of Par-4 (–1 to +248) or the TSS of the E-cadherin promoter (–217 to +148). Bisulfite-treated DNA from primary and recurrent tumor cells was transformed into bacteria and 10 replicate colonies were sequenced (rows). Open circles denote unmethylated CpG dinucleotides, closed circles denote methylated CpG dinucleotides. (B) ChIP-qPCR analysis showing enrichment of histone marks H3K4me3, H3K9ac, H3K27me3, H3K9me2, and RNApol2 within the Par-4 promoter (left) in primary and recurrent tumor cells. The E-cadherin promoter (right) is shown to demonstrate the pattern of histone modification at a stably silenced gene. (C) ChIP-seq analysis showing enrichment of histone marks H3K4me3, H3K9ac, H3K27me3, and RNApol2 at the Par-4 promoter in primary and recurrent tumor cells. (D) ChIP-qPCR analysis showing enrichment of histone marks H3K4me3, H3K9ac, H3K27me3, H3K9me2, and RNApol2 at the Par-4 promoter (left) in control or Twist-expressing primary cell line 1. Enrichments for the E-cadherin promoter (right) is included as a control. (E) Western blot analysis showing Par-4 expression in recurrent tumor cells treated for 48 hours with inhibitors of EZH2 (1 μM EPZ6438) or HDACs (1 μM SAHA), either alone or in combination. Primary tumor cell lysate is shown as a control for Par-4 expression. Error bars denote mean ± SEM.