The DAXX co-repressor is directly recruited to active regulatory elements genome-wide to regulate autophagy programs in a model of human prostate cancer

Abstract

While carcinoma of the prostate is the second most common cause of cancer death in the US, current methods and markers used to predict prostate cancer (PCa) outcome are inadequate. This study was aimed at understanding the genome-wide binding and regulatory role of the DAXX transcriptional repressor, recently implicated in PCa. ChIP-Seq analysis of genome-wide distribution of DAXX in PC3 cells revealed over 59,000 DAXX binding sites, found at regulatory enhancers and promoters. ChIP-Seq analysis of DNA methyltransferase 1 (DNMT1), which is a key epigenetic partner for DAXX repression, revealed that DNMT1 binding was restricted to a small number of DAXX sites. DNMT1 and DAXX bound close to transcriptional activator motifs. DNMT1 sites were found to be dependent on DAXX for recruitment by analyzing DNMT1 ChIP-Seq following DAXX knockdown (K/D), corroborating previous findings that DAXX recruits DNMT1 to repress its target genes. Massively parallel RNA sequencing (RNA-Seq) was used to compare the transcriptomes of WT and DAXX K/D PC3 cells. Genes induced by DAXX K/D included those involved in autophagy, and DAXX ChIP-Seq peaks were found close to the transcription start sites (TSS) of autophagy genes, implying they are more likely to be regulated by DAXX. In conclusion, DAXX binds active regulatory elements and co-localizes with DNMT1 in the prostate cancer genome. Given DAXX's putative regulatory role in autophagy, future studies may consider DAXX as a candidate marker and therapeutic target for prostate cancer.

Transcriptional Repressor DAXX Promotes Prostate Cancer Tumorigenicity via Suppression of Autophagy

“…10B) cannot proceed. Although ULK1 has multiple targets in cell signaling and regulates other cellular processes besides autophagy (55), additional support that autophagy is indeed a mechanism by which DAXX affects tumorigenesis in PCa comes through our use of the ATG5-deficient cell line, DU145 (this study) and ChIP-seq/RNA-seq analyses (13). DAXX ChIP-seq peaks were found close to the TSSs of autophagy genes, and genes increased by DAXX K/D included those involved in autophagy.…”

“…DAXX ChIP-seq peaks were found close to the TSSs of autophagy genes, and genes increased by DAXX K/D included those involved in autophagy. Furthermore, RNA-seq showed that DAXX was inversely correlated with the expression of positive regulators of autophagy (13).…”

“…Anti-DAXX (sc-7001) goat polyclonal antibody from Santa Cruz Biotechnology, Inc. was used. A two-step cross-linking procedure preceded ChIP as described (13). Deep sequencing (ChIP-seq) was performed using an Illumina HiSeq 2500 system.…”

“…Deep sequencing (ChIP-seq) was performed using an Illumina HiSeq 2500 system. Genomic data analysis and visualization were done as described, using Bowtie2 and HOMER (13).…”

Epigenetic Control of Autophagy in Cancer Cells: A Key Process for Cancer-Related Phenotypes

“…ChIPsequencing analyses showed in a prostate cancer cell model that the co-repressor DAXX (death domain associated protein) colocalized with DNMT1 near the TSS (transcriptional start site) of several ATG-related genes including ULK1 and DAPK3 (death-associated protein kinase 3) [5]. Since DAXX has already been shown to specifically recruit DNMTs on specific target promoters, these data strongly suggested that DAXX may actively mediate a methylation-dependent silencing of ATG genes [6].…”