Relationship between estrogen receptor alpha location and gene induction reveals the importance of downstream sites and cofactors

BACKGROUND: To understand cancer-related modifications to transcriptional programs requires detailed knowledge about the activation of signal-transduction pathways and gene expression programs. To investigate the mechanisms of target gene regulation by human estrogen receptor alpha (hERalpha), we combine extensive location and expression datasets with genomic sequence analysis. In particular, we study the influence of patterns of DNA occupancy by hERalpha on expression phenotypes. RESULTS: We find that strong ChIP-chip sites co-localize with strong hERalpha consensus sites and detect nucleotide bias near hERalpha sites. The localization of ChIP-chip sites relative to annotated genes shows that weak sites are enriched near transcription start sites, while stronger sites show no positional bias. Assessing the relationship between binding configurations and expression phenotypes, we find binding sites downstream of the transcription start site (TSS) to be equally good or better predictors of hERalpha-mediated expression as upstream sites. The study of FOX and SP1 cofactor sites near hERalpha ChIP sites shows that induced genes frequently have FOX or SP1 sites. Finally we integrate these multiple datasets to define a high confidence set of primary hERalpha target genes. CONCLUSION: Our results support the model of long-range interactions of hERalpha with the promoter-bound cofactor SP1 residing at the promoter of hERalpha target genes. FOX motifs co-occur with hERalpha motifs along responsive genes. Importantly we show that the spatial arrangement of sites near the start sites and within the full transcript is important in determining response to estrogen signaling.

Epigenetic remodelling of enhancers in response to estrogen deprivation and re-stimulation

Ludovica Vanzan

Estrogen hormones are implicated in a majority of breast cancers and estrogen receptor alpha (ER), the main nuclear factor mediating estrogen signaling, orchestrates a complex molecular circuitry that is not yet fully elucidated. Here, we investigated genome-wide DNA methylation, histone acetylation and transcription after estradiol (E2) deprivation and re-stimulation to better characterize the ability of ER to coordinate gene regulation. We found that E2 deprivation mostly resulted in DNA hypermethylation and histone deacetylation in enhancers. Transcriptome analysis revealed that E2 deprivation leads to a global down-regulation in gene expression, and more specifically of TET2 demethylase that may be involved in the DNA hypermethylation following short-term E2 deprivation. Further enrichment analysis of transcription factor (TF) binding and motif occurrence highlights the importance of ER connection mainly with two partner TF families, AP-1 and FOX. These interactions take place in the proximity of E2 deprivation-mediated differentially methylated and histone acetylated enhancers. Finally, while most deprivation-dependent epigenetic changes were reversed following E2 re-stimulation, DNA hypermethylation and H3K27 deacetylation at certain enhancers were partially retained. Overall, these results show that inactivation of ER mediates rapid and mostly reversible epigenetic changes at enhancers, and bring new insight into early events, which may ultimately lead to endocrine resistance.

OXFORD UNIV PRESS2021

Mammalian TOR as a central modulator of the transcriptional response of keratinocyte stem cells to small temperature variations

Fahd Azzabi Zouraq

Being in the layer at the interface of the body with the outside world, keratinocyte stem cells constantly adapt and respond to environmental cues such as temperature and oxygen. However, until now, experiments involving temperature have only examined cold and heat shocks. Here, our work illustrates that small variations of temperature (~1°C) are enough to affect colony size of keratinocyte stem cells and alter expression of some genes involved in various signalling pathways. The promoter regions of the temperature sensitive genes share a sequence located in some Alu short interspersed nuclear elements (Alu SINEs). This sequence termed TORRID contains several IFHL-like motifs (IFHL motif is related to ribogenesis in yeast) and interestingly a minimal TORRID sequence might be enough for a gene to acquire temperature sensitivity. Our findings demonstrated that the expression of temperature sensitive genes is regulated through the mammalian target of rapamycin (mTOR) pathway. The transcription factors ILF1 and RAP1, and histone deacetylase II (HDAC2) –human orthologs of yeast Fhl1p, Rap1 and Rpd3 respectively– associate with TORRID sequences. Small variations of temperature were enough to alter this association. Moreover, mTOR can translocate to the nucleus and associate with the TORRID sequences in a rapamycin and temperature dependent manner. Furthermore, Raptor and Rictor, members of the mTOR complexes 1 and 2 are also involved in this binding. Hence, our results reveal not only that TOR transcriptional activity is evolutionary conserved from yeast to human, but also that a small variation in temperature represents a new input to the mTOR pathway and fine-tune regulation of transcription through TORRID sequences. These results shed new light on the role of Alus in gene regulation.

EPFL2008

Identification of molecular apocrine breast tumours by microarray analysis

Cathrin Brisken, Maryse Fiche, Darlene Goldstein

Previous microarray studies on breast cancer identified multiple tumour classes, of which the most prominent, named luminal and basal, differ in expression of the oestrogen receptor alpha gene (ER). We report here the identification of a group of breast tumours with increased androgen signalling and a 'molecular apocrine' gene expression profile. Tumour samples from 49 patients with large operable or locally advanced breast cancers were tested on Affymetrix U133A gene expression microarrays. Principal components analysis and hierarchical clustering split the tumours into three groups: basal, luminal and a group we call molecular apocrine. All of the molecular apocrine tumours have strong apocrine features on histological examination (P=0.0002). The molecular apocrine group is androgen receptor (AR) positive and contains all of the ER-negative tumours outside the basal group. Kolmogorov-Smirnov testing indicates that oestrogen signalling is most active in the luminal group, and androgen signalling is most active in the molecular apocrine group. ERBB2 amplification is commoner in the molecular apocrine than the other groups. Genes that best split the three groups were identified by Wilcoxon test. Correlation of the average expression profile of these genes in our data with the expression profile of individual tumours in four published breast cancer studies suggest that molecular apocrine tumours represent 8-14% of tumours in these studies. Our data show that it is possible with microarray data to divide mammary tumour cells into three groups based on steroid receptor activity: luminal (ER+ AR+), basal (ER- AR-) and molecular apocrine (ER- AR+).

2005