Prostate cancer stem cells (CSC) are implicated in tumor initiation, cancer progression, metastasis, and the development of therapeutic-resistant disease. It is well known that the bulk of prostate cancer cells express androgen receptor (AR) and that androgens are required for prostate cancer growth, progression, and emergence of castration-resistant disease. In contrast, the small subpopulation of self-renewing CSCs exhibits an AR-negative (AR−) signature. The mechanisms underlying the absence of AR are unknown. Using CSC-like cell models isolated from clinical biopsy tissues, we identify the E3 ligase MDM2 as a key regulator of prostate CSC integrity. First, unlike what has been reported for the bulk of AR+ tumor cells where MDM2 regulates the temporal expression of AR during transcriptional activity, MDM2 in CSCs promoted the constant ubiquitination and degradation of AR, resulting in sustained loss of total AR protein. Second, MDM2 promoted CSC self-renewal, the expression of stem cell factors, and CSC proliferation. Loss of MDM2 reversed these processes and induced expression of full-length AR (and not AR variants), terminal differentiation into luminal cells, and cell death. Selectively blocking MDM2-mediated activity in combination with androgen/AR-targeted therapy may offer a novel strategy for eliminating AR− CSCs in addition to the bulk of AR+ prostate cancer cells, decreasing metastatic tumor burden and inhibiting the emergence of therapeutic resistance. Significance: These findings provide a novel mechanistic aspect of prostate cancer cell stemness that advances our understanding of the diverse transcriptional activity that bypasses AR in contributing to therapeutic resistance, tumor progression, and metastasis.

Hexavalent chromium compounds are well-established respiratory carcinogens used in industrial processes. While inhalation exposure constitutes an occupational risk affecting mostly chromium workers, environmental exposure from drinking water is a widespread gastrointestinal cancer risk, affecting millions of people throughout the world. Cr(VI) is genotoxic, forming protein-Cr-DNA adducts and silencing tumor suppressor genes, but its mechanism of action at the molecular level is poorly understood. Our prior work using FAIRE showed that Cr(VI) disrupted the binding of transcription factors CTCF and AP-1 to their cognate chromatin sites. Here, we used two complementary approaches to test the hypothesis that chromium perturbs chromatin organization and dynamics. DANPOS2 analyses of MNase-seq data identified several chromatin alterations induced by Cr(VI) affecting nucleosome architecture, including occupancy changes at specific genome locations; position shifts of 10 nucleotides or more; and changes in position amplitude or fuzziness. ATAC-seq analysis revealed that Cr(VI) disrupted the accessibility of chromatin regions enriched for CTCF and AP-1 binding motifs, with a significant co-occurrence of binding sites for both factors in the same region. Cr(VI)-enriched CTCF sites were confirmed by ChIP-seq and found to correlate with evolutionarily conserved sites occupied by CTCF in vivo, as determined by comparison with ENCODE-validated CTCF datasets from mouse liver. In addition, more than 30% of the Cr(VI)-enriched CTCF sites were located in promoters of genes differentially expressed from chromium treatment. Our results support the conclusion that Cr(VI) exposure promotes broad changes in chromatin accessibility and suggest that the subsequent effects on transcription regulation may result from disruption of CTCF binding and nucleosome spacing, implicating transcription regulatory mechanisms as primary Cr(VI) targets.

Complex mixtures of environmental agents often cause mixture-specific health effects that cannot be accounted for by a single mechanism. To study the biological effects of exposure to a mixture of chromium-VI and benzo[a]pyrene (B[a]P), often found together in the environment, we exposed mice for 60 days to 0, 55, 550, or 5500 ppb Cr(VI) in drinking water followed by 90 days of coexposure to B[a]P at 0, 1.25, 12.5, or 125 mg/kg/day and examined liver and gastrointestinal (GI) tract for exposure effects. In the liver, the mixture caused more significant histopathology than expected from the sum of effects of the individual components, while in the GI tract, Cr(VI) alone caused significant enterocyte hypertrophy and increases in cell proliferation and DNA damage that were also observed in mice coexposed to B[a]P. Expression of genes involved in drug metabolism, tumor suppression, oxidative stress, and inflammation was altered in mixed exposures relative to control and to singly exposed mice. Drug metabolism and oxidative stress genes were upregulated and tumor suppressor and inflammation genes downregulated in the proximal GI tract, whereas most markers were upregulated in the distal GI tract and downregulated in the liver. Oral exposure to Cr(VI) and B[a]P mixtures appears to have tissue-specific differential consequences in liver and GI tract that cannot be predicted from the effects of each individual toxicant. Tissue specificity may be particularly critical in cases of extended exposure to mixtures of these agents, as may happen in the occupational setting or in areas where drinking water contains elevated levels of Cr(VI).

Hexavalent chromium (Cr(VI)) is a highly mutagenic and carcinogenic chemical used in many industrial processes. Occupational exposure to chromium, occurring mostly by inhalation, constitutes a major lung cancer risk affecting chromium workers. Environmental exposure, on the other hand, mainly by ingestion of contaminated drinking water, is a widespread gastrointestinal cancer risk, affecting millions of people throughout the world. One of the major mechanisms through which Cr(VI) causes carcinogenic transformation is thought to be the disruption of transcriptional regulation. Indeed, Cr(VI)-directed DNA damage and crosslinking occurs preferentially at sites where active DNA replication and transcription processes take place. Accordingly, numerous studies have shown that Cr(VI) causes gene expression changes in a wide range of cell signaling pathways, resulting from Cr(VI)-induced direct macromolecular damage, alteration in transcription factor function, and disruption of epigenetic signatures. This brief review highlights past and current information on the impact of Cr(VI) on the various mechanisms of transcriptional regulation. Impact statement: This mini-review highlights current evidence on the mechanisms through which hexavalent chromium (Cr(VI)) disrupts transcriptional regulation, an emerging area of interest and one of the central processes by which chromium induces carcinogenesis. Several studies have shown that Cr(VI) causes widespread DNA damage and disrupts epigenetic signatures, suggesting that chromatin may be a direct Cr(VI) target. The findings discussed here suggest that Cr(VI) disrupts transcriptional regulation by causing genomic architecture changes.

Hexavalent chromium compounds are well-established respiratory carcinogens to which humans are commonly exposed in industrial and occupational settings. In addition, natural and anthropogenic sources of these compounds contribute to the exposure of global populations through multiple routes, including dermal, ingestion and inhalation that elevate the risk of cancer by largely unresolved mechanisms. Cr(VI) has genotoxic properties that include ternary adduct formation with DNA, increases in DNA damage, mostly by double-strand break formation, and altered transcriptional responses. Our previous work using ATAC-seq showed that CTCF motifs were enriched in Cr(VI)-dependent differentially accessible chromatin, suggesting that CTCF, a key transcription factor responsible for the regulation of the transcriptome, might be a chromium target. To test this hypothesis, we investigated the effect of Cr(VI) treatment on the binding of CTCF to its cognate sites and ensuing changes in transcription-related histone modifications. Differentially bound CTCF sites were enriched by Cr(VI) treatment within transcription-related regions, specifically transcription start sites and upstream genic regions. Functional annotation of the affected genes highlighted biological processes previously associated with Cr(VI) exposure. Notably, we found that differentially bound CTCF sites proximal to the promoters of this subset of genes were frequently associated with the active histone marks H3K27ac, H3K4me3, and H3K36me3, in agreement with the concept that Cr(VI) targets CTCF in euchromatic regions of the genome. Our results support the conclusion that Cr(VI) treatment promotes the differential binding of CTCF to its cognate sites in genes near transcription-active boundaries, targeting these genes for dysregulation.