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Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture [HTGTS-Seq]

ABSTRACT: Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture [HTGTS-Seq]

PROVIDER: PRJNA596476 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR10734000_1.fastq.gz	Fastqsanger.gz
	SRR10734000_2.fastq.gz	Fastqsanger.gz
	SRR10734001_1.fastq.gz	Fastqsanger.gz
	SRR10734001_2.fastq.gz	Fastqsanger.gz
	SRR10734002_1.fastq.gz	Fastqsanger.gz

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Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture [HTGTS-Seq]

Project description:Our laboratory identified robust recurrent DNA double-strand break (DSB) cluster (“RDC”) genes in mouse neural stem/progenitor cells (NSPCs) by applying the high-throughput, genome-wide, translocation sequencing (HTGTS) method. Genomic alterations of most of the identified RDC-genes have been associated with psychiatric disorders such as autism and schizophrenia and several are altered in brain cancer. The most robust mouse RDCs are all in genes that tend to be very long, actively transcribed and were enhanced after mild inhibition of replication stress. The common transcription and replication characteristics we observe in RDC-genes suggest that frequent RDC DSBs might be generated by collision between transcription and replication processes. However, the underlying mechanism of RDC formation is still unknown. To elucidate mechanisms that generate and resolve DSBs in these cells, we established an in vitro system of induced neural progenitor cells derived from embryonic stem cells. HTGTS bait-DSBs introduced by CRISPR/Cas9 on three mouse chromosomes identified only 5 RDC-genes in embryonic stem cells and 27 RDC-genes in the neural progenitor cells differentiated from them. All RDC-genes identified in induced neural progenitor cells belonged to the group of genes identified in primary neural and progenitor cells. These results indicate that our in vitro differentiation system is both effective and robust in terms of recapitulating our previous findings and facilitating mechanistic studies.

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Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture [GRO-seq]

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Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture

Project description:Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture

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Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture [GRO-seq]

Project description:Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture [GRO-seq]

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Three classes of recurrent DNA break clusters in brain progenitors identified by 3D proximity-based break joining assay

Project description:We recently discovered 27 recurrent DNA double-strand break (DSB) clusters (RDCs) in mouse neural stem/progenitor cells (NSPCs). Most RDCs occurred across long, late-replicating RDC genes and were found only after mild inhibition of DNA replication. RDC genes share intriguing characteristics, including encoding surface proteins that organize brain architecture and neuronal junctions, and are genetically implicated in neuropsychiatric disorders and/or cancers. RDC identification relies on high-throughput genome-wide translocation sequencing (HTGTS), which maps recurrent DSBs based on their translocation to "bait" DSBs in specific chromosomal locations. Cellular heterogeneity in 3D genome organization allowed unequivocal identification of RDCs on 14 different chromosomes using HTGTS baits on three mouse chromosomes. Additional candidate RDCs were also implicated, however, suggesting that some RDCs were missed. To more completely identify RDCs, we exploited our finding that joining of two DSBs occurs more frequently if they lie on the same cis chromosome. Thus, we used CRISPR/Cas9 to introduce specific DSBs into each mouse chromosome in NSPCs that were used as bait for HTGTS libraries. This analysis confirmed all 27 previously identified RDCs and identified many new ones. NSPC RDCs fall into three groups based on length, organization, transcription level, and replication timing of genes within them. While mostly less robust, the largest group of newly defined RDCs share many intriguing characteristics with the original 27. Our findings also revealed RDCs in NSPCs in the absence of induced replication stress, and support the idea that the latter treatment augments an already active endogenous process.

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Long Neural Genes Harbor Recurrent DNA Break Clusters in Neural Stem/Progenitor Cells

Project description:Repair of DNA double-strand breaks (DSBs) by non-homologous end-joining is critical for neural development, and brain cells frequently contain somatic genomic variations that might involve DSB intermediates. We now use an unbiased, high-throughput approach to identify genomic regions harboring recurrent DSBs in primary neural stem/progenitor cells (NSPCs). We identify 27 recurrent DSB clusters (RDCs) and, remarkably, all occur within gene bodies. Most of these NSPC RDCs were detected only upon mild, aphidicolin-induced replication stress, providing a nucleotide-resolution view of replication-associated genomic fragile sites. The vast majority of RDCs occur in long, transcribed, and late-replicating genes. Moreover, almost 90% of identified RDC-containing genes are involved in synapse function and/or neural cell adhesion, with a substantial fraction also implicated in tumor suppression and/or mental disorders. Our characterization of NSPC RDCs reveals a basis of gene fragility and suggests potential impacts of DNA breaks on neurodevelopment and neural functions. We performed high-throughput, genome-wide, translocation sequencing (HTGTS) and GRO-seq in primary mouse neural stem/progenitor cells of the indicated genotypes.

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Induction of recurrent break cluster genes in neural progenitor cells differentiated from embryonic stem cells in culture.

Project description:Mild replication stress enhances appearance of dozens of robust recurrent genomic break clusters, termed RDCs, in cultured primary mouse neural stem and progenitor cells (NSPCs). Robust RDCs occur within genes ("RDC-genes") that are long and have roles in neural cell communications and/or have been implicated in neuropsychiatric diseases or cancer. We sought to develop an in vitro approach to determine whether specific RDC formation is associated with neural development. For this purpose, we adapted a system to induce neural progenitor cell (NPC) development from mouse embryonic stem cell (ESC) lines deficient for XRCC4 plus p53, a genotype that enhances DNA double-strand break (DSB) persistence to enhance detection. We tested for RDCs by our genome-wide DSB identification approach that captures DSBs via their ability to join to specific genomic Cas9/single-guide RNA-generated bait DSBs. In XRCC4/p53-deficient ESCs, we detected seven RDCs, all of which were in genes and two of which were robust. In contrast, in NPCs derived from these ESC lines we detected 29 RDCs, a large fraction of which were robust and associated with long, transcribed neural genes that were also robust RDC-genes in primary NSPCs. These studies suggest that many RDCs present in NSPCs are developmentally influenced to occur in this cell type and indicate that induced development of NPCs from ESCs provides an approach to rapidly elucidate mechanistic aspects of NPC RDC formation.

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Intra-Vk Cluster Recombination Shapes the Ig Kappa Locus Repertoire [HTGTS]

Project description:Intra-Vk Cluster Recombination Shapes the Ig Kappa Locus Repertoire [HTGTS]

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Three classes of recurrent DNA break clusters in brain progenitors identified by 3D proximity-based break joining assay

Project description:Three classes of recurrent DNA break clusters in brain progenitors identified by 3D proximity-based break joining assay

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Induction of Recurrent Break Cluster Genes in Neural Progenitor Cells Differentiated from Embryonic Stem Cells In Culture

Project description:This SuperSeries is composed of the SubSeries listed below.

2020-01-11 | GSE142315 | GEO