AIMS: Telomeric repeat-containing RNAs are long non-coding RNAs generated from the telomeres. TERRAs are essential for the establishment of heterochromatin marks at telomeres, which serve for the binding of members of the heterochromatin protein 1 (HP1) protein family of epigenetic modifiers involved with chromatin compaction and gene silencing. While HP1γ is enriched on gene bodies of actively transcribed human and mouse genes, it is unclear if its transcriptional role is important for HP1γ function in telomere cohesion and telomere maintenance. We aimed to study the effect of mouse HP1γ on the transcription of telomere factors and molecules that can affect telomere maintenance. MAIN METHODS: We investigated the telomere function of HP1γ by using HP1γ deficient mouse embryonic fibroblasts (MEFs). We used gene expression analysis of HP1γ deficient MEFs and validated the molecular and mechanistic consequences of HP1γ loss by telomere FISH, immunofluorescence, RT-qPCR and DNA-RNA immunoprecipitation (DRIP). KEY FINDINGS: Loss of HP1γ in primary MEFs led to a downregulation of various telomere and telomere-accessory transcripts, including the shelterin protein TRF1. Its downregulation is associated with increased telomere replication stress and DNA damage (γH2AX), effects more profound in females. We suggest that the source for the impaired telomere maintenance is a consequence of increased telomeric DNA-RNA hybrids and TERRAs arising at and from mouse chromosomes 18 and X. SIGNIFICANCE: Our results suggest an important transcriptional control by mouse HP1γ of various telomere factors including TRF1 protein and TERRAs that has profound consequences on telomere stability, with a potential sexually dimorphic nature.
Fibroblasts , Telomere , Animals , Humans , Mice , Chromatin , DNA , Fibroblasts/metabolism , RNA/genetics , RNA/metabolism , Telomere/genetics , Telomere/metabolism , Transcription Factors/genetics , Telomeric Repeat Binding Protein 1/metabolism
Epigenetic modifications and nucleosome positioning play an important role in modulating gene expression. However, how the patterns of epigenetic modifications and nucleosome positioning are established around promoters is not well understood. Here, we have addressed these questions in a series of genome-wide experiments coupled to a novel bioinformatic analysis approach. Our data reveal a clear correlation between CpG density, promoter activity and accumulation of active or repressive histone marks. CGI boundaries define the chromatin promoter regions that will be epigenetically modified. CpG-rich promoters are targeted by histone modifications and histone variants, while CpG-poor promoters are regulated by DNA methylation. CGIs boundaries, but not transcriptional activity, are essential determinants of H2A.Z positioning in vicinity of the promoters, suggesting that the presence of H2A.Z is not related to transcriptional control. Accordingly, H2A.Z depletion has no impact on gene expression of arrested mouse embryonic fibroblasts. Therefore, the underlying DNA sequence, the promoter CpG density and, to a lesser extent, transcriptional activity, are key factors implicated in promoter chromatin architecture.
CpG Islands , Epigenesis, Genetic , Epigenome , Histones/genetics , Promoter Regions, Genetic , Protein Processing, Post-Translational , Animals , Chromatin/metabolism , Chromatin/ultrastructure , Chromatin Assembly and Disassembly , Computational Biology/methods , DNA Methylation , Embryo, Mammalian , Fibroblasts/cytology , Fibroblasts/metabolism , Histones/chemistry , Histones/deficiency , Histones/metabolism , Mice , Mice, Knockout , Primary Cell Culture , Protein Isoforms/chemistry , Protein Isoforms/genetics , Protein Isoforms/metabolism
