Long range spreading of dosage compensation in Drosophila captures transcribed autosomal genes inserted on X
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ABSTRACT:
ORGANISM(S): Drosophila melanogaster
PROVIDER: GSE17729 | GEO | 2009/08/20
SECONDARY ACCESSION(S): PRJNA118377
REPOSITORIES: GEO
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Project description:Dosage compensation in D. melanogaster males is achieved via targeting of the MSL complex to X chromosomal genes. This is proposed to involve initial sequence-specific recognition of the X at ~150-300 chromatin entry sites, and subsequent spreading to nearby active genes. Here we test a model in which the spreading step requires transcription and is sequence-independent. We ask whether, in the native context of the X chromosome, MSL complex will target genes of autosomal origin. We find that MSL complex does bind such genes, but only if transcriptionally active. Targeting is accompanied by acetylation of the histone H4K16 residue and two-fold transcriptional up-regulation. We conclude that the presence of a long-sought specific DNA sequence within X-linked genes is not obligatory for MSL complex binding. Instead, physical linkage and transcription play the pivotal roles in the identification of MSL targets irrespective of their origin and DNA sequence. Keywords: Epigenetics ChIP-seq measurements of MSL complex binding and input control. Chromatin was prepared from third instar male larvae of a genotype y w TrojanElephant; MSL3-TAP; msl3. TrojanElephant is a mini-white- and yellow-marked transposition of genomic region spanning 65 kb from cg13773 to snRNP70K. MSL3-TAP is a mini-white-marked TAP-tagged genomic msl3 transgene. IgG beads were used for pull-down.
2009-08-20 | E-GEOD-17729 | biostudies-arrayexpress
Project description:Domains of heterochromatin play important roles in the maintenance and regulation of eukaryotic genomes. However, the repressive nature of heterochromatin combined with its propensity to self-propagate necessitates the existence of robust mechanisms that limit heterochromatin spreading and thereby avoid silencing of expressed genes. A number of specific sequence elements have been found to serve as barriers to heterochromatin spreading; however, the mechanisms by which spreading is curtailed are generally not well understood. Here we uncover a role for PAF complex component Leo1 in regulating heterochromatin cis-spreading. A genetic screen revealed that loss of Leo1 results in spreading of heterochromatin across a centromeric (IRC) boundary element in fission yeast. Similar heterochromatin spreading was seen upon deletion of other components of the PAF complex, but not other factors involved in transcription-coupled chromatin modification, indicating a specific role for the PAF complex in heterochromatin regulation. Loss of Leo1 is associated with reduced levels of H4K16 acetylation at the boundary, while tethering of the H4K16 acetyltransferase Mst1 to boundary chromatin suppresses heterochromatin spreading in leo1? cells, suggesting that Leo1 antagonises heterochromatin spreading by facilitating H4K16 acetylation. Interestingly, Leo1 also regulates heterochromatin spreading independently of boundaries, and loss of Leo1 causes redistribution of heterochromatin, in particular resulting in substantial expansion of telomeric heterochromatin domains. The PAF complex is known to be an important regulator of transcription-related chromatin modifications; our findings reveal a previously undescribed role for this complex in global regulation of heterochromatin spreading in cis. 8 samples: input (whole cell extract) and IP from H3K9me2 ChIP in wild-type and leo1? cells, in duplicate
2015-05-22 | E-GEOD-61688 | biostudies-arrayexpress
Project description:Domains of heterochromatin play important roles in the maintenance and regulation of eukaryotic genomes. However, the repressive nature of heterochromatin combined with its propensity to self-propagate necessitates the existence of robust mechanisms that limit heterochromatin spreading and thereby avoid silencing of expressed genes. A number of specific sequence elements have been found to serve as barriers to heterochromatin spreading; however, the mechanisms by which spreading is curtailed are generally not well understood. Here we uncover a role for PAF complex component Leo1 in regulating heterochromatin cis-spreading. A genetic screen revealed that loss of Leo1 results in spreading of heterochromatin across a centromeric (IRC) boundary element in fission yeast. Similar heterochromatin spreading was seen upon deletion of other components of the PAF complex, but not other factors involved in transcription-coupled chromatin modification, indicating a specific role for the PAF complex in heterochromatin regulation. Loss of Leo1 is associated with reduced levels of H4K16 acetylation at the boundary, while tethering of the H4K16 acetyltransferase Mst1 to boundary chromatin suppresses heterochromatin spreading in leo1Δ cells, suggesting that Leo1 antagonises heterochromatin spreading by facilitating H4K16 acetylation. Interestingly, Leo1 also regulates heterochromatin spreading independently of boundaries, and loss of Leo1 causes redistribution of heterochromatin, in particular resulting in substantial expansion of telomeric heterochromatin domains. The PAF complex is known to be an important regulator of transcription-related chromatin modifications; our findings reveal a previously undescribed role for this complex in global regulation of heterochromatin spreading in cis.
2015-05-22 | GSE61688 | GEO
Project description:The Drosophila MSL complex mediates dosage compensation by increasing transcription of the single X chromosome in males approximately two-fold. This is accomplished through recognition of the X chromosome and subsequent acetylation of histone H4K16 on X-linked genes. Initial binding to the X is thought to occur at a subset of sites. However, the consensus sequence motif of entry sites (M-bM-^@M-^\MSL recognition elementM-bM-^@M-^] or MRE) is only slightly enriched on the X (~2 fold), and only a fraction of them is utilized by the MSL complex. Here we ask whether chromatin context could distinguish between utilized and non-utilized copies of the motif, by comparing their relative enrichment for histone modifications and chromosomal proteins mapped in the NHGRI modENCODE project. Through a comparative analysis of the chromatin features in male S2 cells, which contain MSL complex, and female Kc cells, which lack the complex, we find that the presence of active chromatin modifications, together with an elevated local GC content in surrounding sequence, has strong predictive value for functional MSL entry sites, independent of MSL binding. We tested these sites for function in Kc cells by RNAi knockdown of Sxl, resulting in induction of MSL complex. We show that ectopic MSL expression in Kc cells leads to H4K16 acetylation around these sites, and a relative increase in X chromosome transcription. Collectively, our results support a model in which a pre-existing active chromatin environment, coincident with H3K36me3, contributes to MSL entry site selection. The consequences of MSL targeting of the male X chromosome include increase in nucleosome lability, enrichment for H4K16 acetylation and JIL-1 kinase, and depletion of linker histone H1 on active X-linked genes. Our finding serves as a model to understand how chromatin and local sequence features are involved in the selection of functional protein binding sites in the genome. The key Drosophila female sex determinant protein, SXL, represses dosage compensation by inhibiting MSL2 translation. Loss of SXL results in the expression, stabilization, and targetting of the MSL complex in female cells. Therefore, depletion of SXL by RNA interference (RNAi) in female Kc cells will lead to a MSL2-dependent increase in transcription from the female X chromosomes, consistent with the induction of dosage compensation. In this experiment, we generated ChIP-chip profiles of H4K16 acetylation (H4K16ac) in Kc cells of control (GFP) and Sxl RNAi. For ChIP, we used Upstate (now Millipore) anti-H4K16ac antibody, catalog # 07-329, lot #JBC1355376.
2012-03-19 | E-GEOD-34857 | biostudies-arrayexpress
Project description:The Drosophila MSL complex mediates dosage compensation by increasing transcription of the single X chromosome in males approximately two-fold. This is accomplished through recognition of the X chromosome and subsequent acetylation of histone H4K16 on X-linked genes. Initial binding to the X is thought to occur at a subset of sites. However, the consensus sequence motif of entry sites (“MSL recognition element” or MRE) is only slightly enriched on the X (~2 fold), and only a fraction of them is utilized by the MSL complex. Here we ask whether chromatin context could distinguish between utilized and non-utilized copies of the motif, by comparing their relative enrichment for histone modifications and chromosomal proteins mapped in the NHGRI modENCODE project. Through a comparative analysis of the chromatin features in male S2 cells, which contain MSL complex, and female Kc cells, which lack the complex, we find that the presence of active chromatin modifications, together with an elevated local GC content in surrounding sequence, has strong predictive value for functional MSL entry sites, independent of MSL binding. We tested these sites for function in Kc cells by RNAi knockdown of Sxl, resulting in induction of MSL complex. We show that ectopic MSL expression in Kc cells leads to H4K16 acetylation around these sites, and a relative increase in X chromosome transcription. Collectively, our results support a model in which a pre-existing active chromatin environment, coincident with H3K36me3, contributes to MSL entry site selection. The consequences of MSL targeting of the male X chromosome include increase in nucleosome lability, enrichment for H4K16 acetylation and JIL-1 kinase, and depletion of linker histone H1 on active X-linked genes. Our finding serves as a model to understand how chromatin and local sequence features are involved in the selection of functional protein binding sites in the genome.
2012-03-19 | GSE34858 | GEO
Project description:The Drosophila MSL complex mediates dosage compensation by increasing transcription of the single X chromosome in males approximately two-fold. This is accomplished through recognition of the X chromosome and subsequent acetylation of histone H4K16 on X-linked genes. Initial binding to the X is thought to occur at a subset of sites. However, the consensus sequence motif of entry sites (“MSL recognition element” or MRE) is only slightly enriched on the X (~2 fold), and only a fraction of them is utilized by the MSL complex. Here we ask whether chromatin context could distinguish between utilized and non-utilized copies of the motif, by comparing their relative enrichment for histone modifications and chromosomal proteins mapped in the NHGRI modENCODE project. Through a comparative analysis of the chromatin features in male S2 cells, which contain MSL complex, and female Kc cells, which lack the complex, we find that the presence of active chromatin modifications, together with an elevated local GC content in surrounding sequence, has strong predictive value for functional MSL entry sites, independent of MSL binding. We tested these sites for function in Kc cells by RNAi knockdown of Sxl, resulting in induction of MSL complex. We show that ectopic MSL expression in Kc cells leads to H4K16 acetylation around these sites, and a relative increase in X chromosome transcription. Collectively, our results support a model in which a pre-existing active chromatin environment, coincident with H3K36me3, contributes to MSL entry site selection. The consequences of MSL targeting of the male X chromosome include increase in nucleosome lability, enrichment for H4K16 acetylation and JIL-1 kinase, and depletion of linker histone H1 on active X-linked genes. Our finding serves as a model to understand how chromatin and local sequence features are involved in the selection of functional protein binding sites in the genome.
2012-03-19 | GSE34857 | GEO
Project description:The Drosophila MSL complex mediates dosage compensation by increasing transcription of the single X chromosome in males approximately two-fold. This is accomplished through recognition of the X chromosome and subsequent acetylation of histone H4K16 on X-linked genes. Initial binding to the X is thought to occur at a subset of sites. However, the consensus sequence motif of entry sites (“MSL recognition element” or MRE) is only slightly enriched on the X (~2 fold), and only a fraction of them is utilized by the MSL complex. Here we ask whether chromatin context could distinguish between utilized and non-utilized copies of the motif, by comparing their relative enrichment for histone modifications and chromosomal proteins mapped in the NHGRI modENCODE project. Through a comparative analysis of the chromatin features in male S2 cells, which contain MSL complex, and female Kc cells, which lack the complex, we find that the presence of active chromatin modifications, together with an elevated local GC content in surrounding sequence, has strong predictive value for functional MSL entry sites, independent of MSL binding. We tested these sites for function in Kc cells by RNAi knockdown of Sxl, resulting in induction of MSL complex. We show that ectopic MSL expression in Kc cells leads to H4K16 acetylation around these sites, and a relative increase in X chromosome transcription. Collectively, our results support a model in which a pre-existing active chromatin environment, coincident with H3K36me3, contributes to MSL entry site selection. The consequences of MSL targeting of the male X chromosome include increase in nucleosome lability, enrichment for H4K16 acetylation and JIL-1 kinase, and depletion of linker histone H1 on active X-linked genes. Our finding serves as a model to understand how chromatin and local sequence features are involved in the selection of functional protein binding sites in the genome. The key Drosophila female sex determinant protein, SXL, represses dosage compensation by inhibiting MSL2 translation. Loss of SXL results in the expression, stabilization, and targetting of the MSL complex in female cells. Therefore, depletion of SXL by RNA interference (RNAi) in female Kc cells will lead to a MSL2-dependent increase in transcription from the female X chromosomes, consistent with the induction of dosage compensation. In this experiment, we generated gene expression profiles of Kc cells of control (GFP), Sxl RNAi and Sxl-Msl2 RNAi experiments.
2012-03-19 | E-GEOD-34858 | biostudies-arrayexpress
Project description:In Drosophila, the global increase in transcription from the X chromosome in males to compensate for its monosomy is mediated by the male-specific-lethal complex (MSL-C) consisting of five proteins and two non-coding RNAs, roX1 and roX2. After an initial sequence dependent recognition by the MSL-C of 150-300 high affinity sites, the spreading to the majority of the X-linked genes depends on local MSL-C concentration and active transcription. Here we ask whether any additional RNA species are associated to the MSL-C. No additional roX were found but a strong association was found between the msl2 mRNA and the MSL-C. Based on our results we propose a model in which a non-chromatin associated partial or complete MSL-C titrates newly transcribed msl2 mRNA and thus feed-back regulates the amount of available MSL-C. In total 12 samples; 4 Input files (4 different conditions) with the corresponding 8 RIP samples (2 different antibodies, same 4 conditions as Input)
2011-04-13 | E-GEOD-20249 | biostudies-arrayexpress
Project description:Dosage compensation serves as a model for understanding how chromatin-modification enzymes are targeted to initiate and maintain gene regulation. In Drosophila, the MSL complex associates with active genes specifically on the male X chromosome to acetylate histone H4 at lysine 16, and increase expression of most X-linked genes approximately two-fold. To date, no DNA sequence has been discovered to explain the specificity of MSL binding. We previously hypothesized that sequence-specific targeting occurs at initiation or “chromatin entry sites”, but binding to the majority of sites is sequence-independent. Here we characterize more than 150 potential entry sites by ChIP-chip and ChIP-seq and discover a common GA-rich MSL recognition element (MRE). The motif is only slightly enriched on the X chromosome (~2 fold), but this is doubled when considering its preferential location within or 3’ to active X-linked genes (>4 fold enrichment). When inserted on an autosome, a newly identified site can direct local MSL spreading to flanking active genes on the autosome. These results provide strong evidence for both sequence-dependent and -independent steps in MSL targeting of dosage compensation to the male X chromosome.
2008-08-22 | GSE11485 | GEO
Project description:In Drosophila, the global increase in transcription from the X chromosome in males to compensate for its monosomy is mediated by the male-specific-lethal complex (MSL-C) consisting of five proteins and two non-coding RNAs, roX1 and roX2. After an initial sequence dependent recognition by the MSL-C of 150-300 high affinity sites, the spreading to the majority of the X-linked genes depends on local MSL-C concentration and active transcription. Here we ask whether any additional RNA species are associated to the MSL-C. No additional roX were found but a strong association was found between the msl2 mRNA and the MSL-C. Based on our results we propose a model in which a non-chromatin associated partial or complete MSL-C titrates newly transcribed msl2 mRNA and thus feed-back regulates the amount of available MSL-C.
2011-04-13 | GSE20249 | GEO