Unknown,Transcriptomics,Genomics,Proteomics

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Bru-seq nascent RNA sequencing of human and mouse cells: correlation to replication timing and genome instability

ABSTRACT: Bru-seq nascent RNA sequencing (PubMed ID 23973811) was performed on two primary human fibroblast cell lines, mouse embryonic stem cells, and GM12878 human lymphoblastoid cells. Read data, which include both exon and intron signals, were used to identify transcription unit spans genome-wide, where a transcription unit is roughly correspondent to the longest expressed isoform of a gene. However, because algorithms were not constrained by annotated genes, transcription units need not and often do not correspond precisely to gene boundaries and include extragenic transcription. Transcription units were then compared to separate data sets that comprised induced copy number variants, common fragile sites, and Repli-seq replication timing. The objective was to discover the relationships between transcription unit span and size, local genomic instability, and replication timing. This GEO sample series provides the span and intensity of transcription units called genome-wide in the various samples. Correlations to genome stability and replication timing are provided in the associated manuscript. In addition, one human fibroblast line and the mouse embryonic stem cells had paired samples treated and untreated with low dose aphidicolin. Gene RPKM signal intensities are provided for these samples, although comparing these was not the principal objective of the study. Bru-seq single-read nascent RNA sequencing on human 090 fibroblasts +/- aphidicolin treatment, human UMHF1 fibroblasts (3 replicates), human GM12878 lymphoblastoid cells, and mouse embryonic stem cells+/- aphidicolin treatment.

ORGANISM(S): Homo sapiens

SUBMITTER: Thomas Wilson

PROVIDER: E-GEOD-55862 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Several copy number altered regions (CNA) have been identified in the genome of cervical cancer, especially amplifications of 3q and 5p. However, the contribution of those alterations to cervical carcinogenesis is still a matter of debate, since genome-wide, there is a lack of correlation between CNAs and gene expression. In this study, we investigated whether the CNAs in cell lines (CaLo, CasKi, HeLa, SiHa), at a gene-by-gene level, are related to changes in gene expression. On average 19.2% of the whole genome of cell lines had CNA. However, only 2.4% consisted of minimal recurrent regions (MRR), common to all cell lines. Whereas 3q had just some sparse common gains (13%), 5p was entirely duplicated recurrently. Genome-wide, only 11% of genes located in CNAs changed gene expression. In contrast, the rate increased over 3 fold times in MRRs. Chr 5p was confirmed entirely amplified by FISH. In spite of this, at most 32.9% of the explored genes in 5p (n=202) were de-regulated. In 3q, the rate was just 11.8%. Even in 3q26, which had five MRRs and 38.7% of SNPs was gained recurrently, the rate rose slightly to 13.6% (10 out of 73). Interestingly, up to 16% of de-regulated genes in 5p and 80% in 3q26 were down-regulated, suggesting additional factors are involved in gene repression. The de-regulated genes in 3q and 5p were found in clusters, suggesting local chromatin factors may also influence gene expression. In regions amplified discontinuously, the rate of down-regulated genes rose steadily as the number of amplified SNPs increased (p<0.01, Spearman's correlation). This suggests partial gene amplification as a mechanism of silencing gene expression. Additional genes were identified up- or down-regulated in 5p and 3q, which could be involved in cervical carcinogenesis, especially implicated in apoptosis. Those include CLPTM1L, AHRR, PDCD6 and DAP in 5p and TNFSF10 and ECT2 in 3q. Overall, the gene expression and copy number profiles suggest other factors, like epigenetic or chromatin domains, may influence gene expression within the entirely amplified genome segments. Further studies are needed to elucidate how these mechanisms regulate gene expression. Affymetrix SNP arrays were performed according to the manufacturer's directions on DNA extracted from cell lines or peripheral blood samples. Copy number analysis of Affymetrix 100K SNP arrays was performed for 4 cervical cancer cell lines. DNA from 38 healty women were used as references for copy number inference.

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Bru-seq nascent RNA sequencing of human and mouse cells: correlation to replication timing and genome instability

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