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An epigenetic signature for monoallelic olfactory receptor expression.

ABSTRACT: Constitutive heterochromatin is traditionally viewed as the static form of heterochromatin that silences pericentromeric and telomeric repeats in a cell cycle- and differentiation-independent manner. Here, we show that, in the mouse olfactory epithelium, olfactory receptor (OR) genes are marked in a highly dynamic fashion with the molecular hallmarks of constitutive heterochromatin, H3K9me3 and H4K20me3. The cell type and developmentally dependent deposition of these marks along the OR clusters are, most likely, reversed during the process of OR choice to allow for monogenic and monoallelic OR expression. In contrast to the current view of OR choice, our data suggest that OR silencing takes place before OR expression, indicating that it is not the product of an OR-elicited feedback signal. Our findings suggest that chromatin-mediated silencing lays a molecular foundation upon which singular and stochastic selection for gene expression can be applied.

SUBMITTER: Magklara A

PROVIDER: S-EPMC3094500 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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An epigenetic signature for monoallelic olfactory receptor expression.

Magklara Angeliki A Yen Angela A Colquitt Bradley M BM Clowney E Josephine EJ Allen William W Markenscoff-Papadimitriou Eirene E Evans Zoe A ZA Kheradpour Pouya P Mountoufaris George G Carey Catriona C Barnea Gilad G Kellis Manolis M Lomvardas Stavros S

Cell 20110428 4

Constitutive heterochromatin is traditionally viewed as the static form of heterochromatin that silences pericentromeric and telomeric repeats in a cell cycle- and differentiation-independent manner. Here, we show that, in the mouse olfactory epithelium, olfactory receptor (OR) genes are marked in a highly dynamic fashion with the molecular hallmarks of constitutive heterochromatin, H3K9me3 and H4K20me3. The cell type and developmentally dependent deposition of these marks along the OR clusters ...[more]

PMID: 21529909

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Project description:Distant enhancer elements are a major source of specificity in mammalian gene expression. Although enhancers that regulate broad developmental decisions and inducible gene expression have been studied extensively, little is known about regulatory elements that govern monogenic and monoallelic expression. Here, using high throughput epigenetic and genetic techniques we identified a plethora of distant enhancers that regulate monoallelic olfactory receptor (OR) gene expression. Potential OR enhancers have unique, cell type specific epigenetic marks that distinguish them from other neuronal enhancers and correlate with enhancer activity in vivo. Using sequence capture to enrich for these sequences we identified Dnase-protected footprints that reveal novel regulatory sequences and transcription factors required for OR gene activation. Our experiments provide insight to the regulation of OR expression, and describe novel principles and methodologies towards the understanding of transcriptional mechanisms that generate cellular diversity. In vivo examination of H3K79me3 enrichment and DNAse protected footprints on olfactory receptor enhancer sequences. We performed ChIP-seq on native chromatin isolated from the mouse olfactory epithelium using antibodies against H3K79me3. To sequence accessible regions of the genome we treated nuclei with limiting amounts of DNAse I to digest accessible chromatin and perform Dnase Hypersensitivity (DHS)-seq. In the olfactory epithelium, the H enhancer – the first described enhancer for olfactory receptors has a well-defined DNAse I hypersensitivity peak and is flanked by high levels of H3K79me3. We find other intergenic sequences nearby olfactory receptor genes that share the same chromatin signature, and test their function in vivo. TO uncover transcription factor footprints on olfactory receptor enhancers we performed sequence capture of the DHS-seq library to enrich for these sequences. We find multiple DNAse-protected sequences and perform motif analysis on transcription factor footprints to reveal factors involved in olfactory receptor gene regulation.

Project description:BackgroundMonoallelic expression (MAE) is a frequent genomic phenomenon in normal tissues, however its role in cancer is yet to be fully understood. MAE is defined as the expression of a gene that is restricted to one allele in the presence of a diploid heterozygous genome. Constitutive MAE occurs for imprinted genes, odorant receptors and random X inactivation. Several studies in normal tissues have showed MAE in approximately 5-20% of the cases. However, little information exists on the MAE rate in cancer. In this study we assessed the presence and rate of MAE in melanoma. The genetic basis of melanoma has been studied in depth over the past decades, leading to the identification of mutations/genetic alterations responsible for melanoma development.MethodsTo examine the role of MAE in melanoma we used 15 melanoma cell lines and compared their RNA-seq data with genotyping data obtained by the parental TIL (tumor infiltrating lymphocytes). Genotyping was performed using the Illumina HumanOmni1 beadchip. The RNA-seq library preparation and sequencing was performed using the Illumina TruSeq Stranded Total RNA Human Kit and subsequently sequenced using a HiSeq 2500 according to manufacturer's guidelines. By comparing genotyping data with RNA-seq data, we identified SNPs in which DNA genotypes were heterozygous and corresponding RNA genotypes were homozygous. All homozygous DNA genotypes were removed prior to the analysis. To confirm the validity to detect MAE, we examined heterozygous DNA genotypes from X chromosome of female samples as well as for imprinted and olfactory receptor genes and confirmed MAE.ResultsMAE was detected in all 15 cell lines although to a different rate. When looking at the B-allele frequencies we found a preferential pattern of complete monoallelic expression rather then differential monoallelic expression across the 15 melanoma cell lines. As some samples showed high differences in the homozygous and heterozygous call rate, we looked at the single chromosomes and showed that MAE may be explained by underlying large copy number imbalances in some instances. Interestingly these regions included genes known to play a role in melanoma initiation and progression. Nevertheless, some chromosome regions showed MAE without CN imbalances suggesting that additional mechanisms (including epigenetic silencing) may explain MAE in melanoma.ConclusionThe biological implications of MAE are yet to be realized. Nevertheless, our findings suggest that MAE is a common phenomenon in melanoma cell lines. Further analyses are currently being undertaken to evaluate whether MAE is gene/pathway specific and to understand whether MAE can be employed by cancers to achieve a more aggressive phenotype.

Dataset Information

An epigenetic signature for monoallelic olfactory receptor expression.

Publications

An epigenetic signature for monoallelic olfactory receptor expression.

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