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The insulator protein SU(HW) modulates nuclear lamina interactions of the Drosophila genome: expression

ABSTRACT: Specific interactions of the genome with the nuclear lamina (NL) are thought to assist chromosome folding inside the nucleus and to contribute to the regulation of gene expression. High-resolution mapping has recently identified hundreds of large, sharply defined lamina-associated domains (LADs) in the human genome, and suggested that the insulator protein CTCF may help to demarcate these domains. Here, we report the detailed structure of LADs in Drosophila cells, and investigate the putative roles of five insulator proteins in LAD organization. We found that of these five proteins, only SU(HW) binds preferentially at LAD borders and at specific positions inside LADs, while GAF, CTCF, BEAF-32 and DWG are mostly absent from these regions. By knockdown and overexpression studies we demonstrate that SU(HW) weakens LAD – NL interactions by a local antagonistic effect. Our results provide insights into the evolution of LAD organization and reveal a role for SU(HW) in the regulation of genome – NL interactions.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE20312 | GEO | 2011/01/24

SECONDARY ACCESSION(S): PRJNA129843

REPOSITORIES: GEO

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The insulator protein SU(HW) modulates nuclear lamina interactions of the Drosophila genome: DamID

Project description:Specific interactions of the genome with the nuclear lamina (NL) are thought to assist chromosome folding inside the nucleus and to contribute to the regulation of gene expression. High-resolution mapping has recently identified hundreds of large, sharply defined lamina-associated domains (LADs) in the human genome, and suggested that the insulator protein CTCF may help to demarcate these domains. Here, we report the detailed structure of LADs in Drosophila cells, and investigate the putative roles of five insulator proteins in LAD organization. We found that of these five proteins, only SU(HW) binds preferentially at LAD borders and at specific positions inside LADs, while GAF, CTCF, BEAF-32 and DWG are mostly absent from these regions. By knockdown and overexpression studies we demonstrate that SU(HW) weakens LAD – NL interactions by a local antagonistic effect. Our results provide insights into the evolution of LAD organization and reveal a role for SU(HW) in the regulation of genome – NL interactions. DamID experiments for Lamin, CTCF, SU(HW), GAF, DWG, and BEAF-32, and for Lamin after overexpression and after knockdown of SU(HW), were performed in Drosophila cell cultures. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing. Every experiment was done in duplicate in the reverse dye orientation. The supplementary file 'GSE20311_DamID_norm_mean.txt' contains the mean log2(Dam-fusion/Dam-only) values of two replicates.

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2011-01-24 | E-GEOD-20312 | biostudies-arrayexpress

The insulator protein SU(HW) modulates nuclear lamina interactions of the Drosophila genome: DamID

2011-01-24 | GSE20311 | GEO

Decoding the determinants of genome-nuclear lamina interactions by a random scrambling approach (RNA-Seq)

Project description:Mammalian genomes contain hundreds of lamina-associated domains (LADs), which are large, often megabase-sized heterochromatin domains that are anchored to the nuclear lamina (NL). Even though LADs play a key role in the spatial organization of the genome and potently repress gene activity, it is not yet understood how their interactions with the NL are encoded in their DNA. Here, we investigated the principles that govern LAD-NL interactions by taking a "LAD scrambling" approach. This approach relies on 1) local "hopping" of a Sleeping Beauty transposon to randomly insert loxP sites within and around a LAD of choice; and 2) Cre-mediated recombination between the loxP sites. This approach is highly efficient, enabling us to establish a large collection of clonal cell lines with deletions and inversions up to ~2Mb in size, spanning LAD and their flanking inter-LAD sequences. Mapping of NL interactions in these clones revealed that a single LAD contacts the NL through multiple regions. These regions act cooperatively and redundantly; however, some have more affinity for the NL and can boost NL contacts of neighbouring sequences. We also observed a crosstalk between two neighbouring LADs, when close enough to each other. Finally, changes in the heterochromatin mark H3K9me3 only partially mirrored changes in NL contacts. They also correlated more with gene expression changes than NL contacts did. These principles of LAD - NL interactions provide insight into the overall architecture of the genome and the impact on gene regulation.

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Stochastic genome - nuclear lamina contacts are linked to histone H3K9 dimethylation (methylation data)

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2013-01-31 | E-GEOD-40094 | biostudies-arrayexpress

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Project description:The nuclear lamina (NL) interacts with hundreds of large genomic regions termed lamina-associated domains (LADs). The dynamics of these interactions and the relation to epigenetic modifications are poorly understood. We visualized the fate of LADs in single cells using a novel 'molecular contact memory' approach. In each interphase nucleus, only ~30% of LADs are positioned at the periphery; these LADs are in intermittent molecular contact with the NL but remain constrained to the periphery. Upon mitosis, LAD positioning is not detectably inherited but instead is stochastically reshuffled. Contact of individual LADs with the NL correlates with their degree of H3K9 dimethylation in single cells, and inactivation of the H3K9 methyltransferase G9a reduces the NL contact frequencies. These results indicate that nuclear positioning and histone modification of LADs are both stochastic yet linked in single cells. Collectively, these results highlight principles of the dynamic spatial architecture of chromosomes. 12 RNA-seq experiments for 6 samples, each with a biological replicate: m6ATracer-VP16+/DamLaminB1+ m6ATracer-VP16+/DamLaminB1- m6ATracer-VP16-/DamLaminB1- m6ATracer-GFP+/DamLaminB1+ m6ATracer-GFP+/DamLaminB1- m6ATracer-GFP-/DamLaminB1-

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NURF301 contributes to gypsy chromatin insulator-mediated nuclear organization

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2022-10-14 | PXD032165 | Pride

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