Project description:The human silencing hub (HUSH) complex binds to transcripts of LINE-1 retrotransposons (L1s) and other genomic repeats, recruiting MORC2 and other effectors to remodel chromatin. However, how HUSH and MORC2 operate alongside DNA methylation, a central epigenetic regulator of repeat transcription, remains poorly understood. Here we interrogate this relationship in human neural progenitor cells (hNPCs), a somatic model of brain development that tolerates removal of DNA methyltransferase DNMT1. Upon loss of MORC2 or HUSH subunit TASOR in hNPCs, L1s remain silenced by robust promoter methylation. However, genome demethylation and activation of evolutionarily-young L1s attracts MORC2 binding. Simultaneous depletion of DNMT1 and MORC2 causes massive accumulation of L1 transcripts. We identify the same mechanistic hierarchy at pericentromeric α-satellites and clustered protocadherin genes, repetitive elements important for chromosome structure and neurodevelopment respectively. Our data delineate the independent epigenetic control of repeats in somatic cells, with implications for understanding the vital functions of HUSH-MORC2 in hypomethylated contexts throughout human development.
Project description:The human silencing hub (HUSH) complex binds to transcripts of LINE-1 retrotransposons (L1s) and other genomic repeats, recruiting MORC2 and other effectors to remodel chromatin. However, how HUSH and MORC2 operate alongside DNA methylation, a central epigenetic regulator of repeat transcription, remains poorly understood. Here we interrogate this relationship in human neural progenitor cells (hNPCs), a somatic model of brain development that tolerates removal of DNA methyltransferase DNMT1. Upon loss of MORC2 or HUSH subunit TASOR in hNPCs, L1s remain silenced by robust promoter methylation. However, genome demethylation and activation of evolutionarily-young L1s attracts MORC2 binding. Simultaneous depletion of DNMT1 and MORC2 causes massive accumulation of L1 transcripts. We identify the same mechanistic hierarchy at pericentromeric α-satellites and clustered protocadherin genes, repetitive elements important for chromosome structure and neurodevelopment respectively. Our data delineate the independent epigenetic control of repeats in somatic cells, with implications for understanding the vital functions of HUSH-MORC2 in hypomethylated contexts throughout human development.
Project description:The human silencing hub (HUSH) complex binds to transcripts of LINE-1 retrotransposons (L1s) and other genomic repeats, recruiting MORC2 and other effectors to remodel chromatin. However, how HUSH and MORC2 operate alongside DNA methylation, a central epigenetic regulator of repeat transcription, remains poorly understood. Here we interrogate this relationship in human neural progenitor cells (hNPCs), a somatic model of brain development that tolerates removal of DNA methyltransferase DNMT1. Upon loss of MORC2 or HUSH subunit TASOR in hNPCs, L1s remain silenced by robust promoter methylation. However, genome demethylation and activation of evolutionarily-young L1s attracts MORC2 binding. Simultaneous depletion of DNMT1 and MORC2 causes massive accumulation of L1 transcripts. We identify the same mechanistic hierarchy at pericentromeric α-satellites and clustered protocadherin genes, repetitive elements important for chromosome structure and neurodevelopment respectively. Our data delineate the independent epigenetic control of repeats in somatic cells, with implications for understanding the vital functions of HUSH-MORC2 in hypomethylated contexts throughout human development.
Project description:The human silencing hub (HUSH) complex binds to transcripts of LINE-1 retrotransposons (L1s) and other genomic repeats, recruiting MORC2 and other effectors to remodel chromatin. However, how HUSH and MORC2 operate alongside DNA methylation, a central epigenetic regulator of repeat transcription, remains poorly understood. Here we interrogate this relationship in human neural progenitor cells (hNPCs), a somatic model of brain development that tolerates removal of DNA methyltransferase DNMT1. Upon loss of MORC2 or HUSH subunit TASOR in hNPCs, L1s remain silenced by robust promoter methylation. However, genome demethylation and activation of evolutionarily-young L1s attracts MORC2 binding. Simultaneous depletion of DNMT1 and MORC2 causes massive accumulation of L1 transcripts. We identify the same mechanistic hierarchy at pericentromeric α-satellites and clustered protocadherin genes, repetitive elements important for chromosome structure and neurodevelopment respectively. Our data delineate the independent epigenetic control of repeats in somatic cells, with implications for understanding the vital functions of HUSH-MORC2 in hypomethylated contexts throughout human development.
Project description:The human silencing hub (HUSH) complex binds to transcripts of LINE-1 retrotransposons (L1s) and other genomic repeats, recruiting MORC2 and other effectors to remodel chromatin. However, how HUSH and MORC2 operate alongside DNA methylation, a central epigenetic regulator of repeat transcription, remains poorly understood. Here we interrogate this relationship in human neural progenitor cells (hNPCs), a somatic model of brain development that tolerates removal of DNA methyltransferase DNMT1. Upon loss of MORC2 or HUSH subunit TASOR in hNPCs, L1s remain silenced by robust promoter methylation. However, genome demethylation and activation of evolutionarily-young L1s attracts MORC2 binding. Simultaneous depletion of DNMT1 and MORC2 causes massive accumulation of L1 transcripts. We identify the same mechanistic hierarchy at pericentromeric α-satellites and clustered protocadherin genes, repetitive elements important for chromosome structure and neurodevelopment respectively. Our data delineate the independent epigenetic control of repeats in somatic cells, with implications for understanding the vital functions of HUSH-MORC2 in hypomethylated contexts throughout human development.
Project description:Partitioning of active gene loci to the nuclear envelope is a key mechanism by which organisms can increase the speed of adaptation and metabolic robustness to fluctuating resources in the environment. In the budding yeast Saccharomyces cerevisiae, adaptation and transcriptional memory induced by nutrient depletion or other stresses, results from relocalization of active gene loci from nucleoplasm to the nuclear envelope, leading to increased transport of mRNAs to the cytosol and their translation. The mechanism by which this translocation occurs remains a mystery. We show here, that for the inositol depletion-responsive gene locus INO1 in yeast, translocation to the nuclear envelope is caused by a local phase transition of the mechanical stiffness of chromatin surrounding activated INO1 that favors phase separation of the active gene locus into low density regions of chromatin proximal to the nuclear envelope. Gene regulatory elements essential to translocation encode binding sites for histone acetyl transferases, which are necessary for chromatin decompaction. INO1 locus partitioning can be explained by a phenomenological model of chromatin decompaction, which reflects stiffening of chromatin and its partitioning into a dilute chromatin phase adjacent to the nuclear envelope, from the dense chromatin found in the nucleoplasmic phase. Recent evidence suggests that this demixing of chromatin could be due to the dissolution of multivalent chromatin interactions mediated by histone post-translational modifications.
Project description:Epigenetic gene silencing is of central importance to maintain genome integrity and is mediated by an elaborate interplay between DNA methylation, histone posttranslational modifications and chromatin remodeling complexes. DNA methylation and repressive histone marks usually correlate with transcriptionally silent heterochromatin, however there are exceptions to this interdependence. In Arabidopsis, mutation of MORPHEUS MOLECULE 1 (MOM1) causes transcriptional derepression of heterochromatin independently of changes in DNA methylation. More recently, two Arabidopsis homologs of mouse Microrchidia (MORC) have also been implicated in gene silencing and heterochromatin condensation without altering genome-wide DNA methylation patterns. In this study, we show that AtMORC6 physically interacts with AtMORC1 and with its close homologue AtMORC2 in two mutually exclusive protein complexes. RNA-seq analysis of high-order mutants indicates that AtMORC1 and AtMORC2 act redundantly to repress a common set of loci. We also examined the genetic interactions between AtMORC6 and MOM1 pathways. Although AtMORC6 and MOM1 control the silencing of a very similar set of genomic loci, we observed synergistic transcriptional regulation in the mom1/atmorc6 double mutant, suggesting that these epigenetic regulators act mainly by independent silencing mechanisms. RNA-seq libraries were prepared for two suites of mutants to allow direct comparisons between mutants within each set. The two sets consisted of the following samples: Set_1) A wildtype (Col) control, the morc1 mutant, the morc2 mutant, the morc1 morc2 double mutant, the morc6 mutant, and the morc1 morc2 morc6 triple mutant ; Set_2) A wildtpe (Col) control, the morc6 mutant, the mom1 mutant, and the mom1 morc6 double mutant. For each sample, two biological replicates were performed (denoted "bio_replicate_1" or "bio_replicate_2"). Whole genome bisulifte libraries were sequenced from material grown in parallel.
Project description:Variants in the poorly characterised oncoprotein, MORC2, a chromatin remodelling ATPase, lead to defects in epigenetic regulation and DNA damage response. MORC2 variants are associated with multiple cancers and neurological disorders1,2. The C-terminal domain (CTD) of MORC2 is often phosphorylated in DNA damage response and is known to induce cancer progression in in vivo and in vitro cancer models3-5. However, it remains unclear how MORC2 CTD and its phosphorylation impacts its chromatin remodelling activity. Here, we report a molecular characterisation of full-length, phosphorylated MORC2. We show that MORC2 preferentially binds open chromatin, has multiple DNA binding sites with varying affinities and acts as a DNA sliding clamp. We identified a phosphate interacting motif within the CTD that dictates ATP hydrolysis and cooperative DNA binding. The DNA binding impacts several structural domains within the N-terminal ATPase region. We provide the first visual proof that MORC2 induces chromatin remodelling through ATP hydrolysis-dependent DNA compaction, where the speed of compaction is affected by its phosphorylation state. Together, our results reveal that phosphorylation of MORC2 CTD modulates its chromatin remodelling and could be an attractive target for therapeutics.
Project description:We report an in vitro reconstitution of full-length MORC2, the most commonly mutated MORC member, linked to various cancers and neurological disorders. MORC2 possesses multiple DNA binding sites that undergo structural rearrangement upon DNA binding. MORC2 locks onto the DNA using its C-terminal domain (CTD) and acts as a sliding clamp. A conserved phosphate-interacting motif within the CTD was found to regulate ATP hydrolysis and cooperative DNA binding. Importantly, MORC2 mediates chromatin remodelling via ATP hydrolysis-dependent DNA compaction, regulated by the phosphorylation state of its CTD.
Project description:We report an in vitro reconstitution of full-length MORC2, the most commonly mutated MORC member, linked to various cancers and neurological disorders. MORC2 possesses multiple DNA binding sites that undergo structural rearrangement upon DNA binding. MORC2 locks onto the DNA using its C-terminal domain (CTD) and acts as a sliding clamp. A conserved phosphate-interacting motif within the CTD was found to regulate ATP hydrolysis and cooperative DNA binding. Importantly, MORC2 mediates chromatin remodelling via ATP hydrolysis-dependent DNA compaction, regulated by the phosphorylation state of its CTD.