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Series GSE85747 Query DataSets for GSE85747
Status Public on Sep 30, 2016
Title Initiation of mtDNA transcription is followed by pausing, and diverge across human cell types and during evolution
Sample organisms Caenorhabditis elegans; Drosophila melanogaster; Macaca mulatta; Pan troglodytes; Homo sapiens; Mus musculus; Rattus norvegicus
Experiment type Expression profiling by high throughput sequencing
Third-party reanalysis
Summary We analyzed nascent mtDNA-encoded RNA transcripts from GRO-seq and PRO-seq experiments in a collection of diverse human cell lines and Metazoan organisms. Accurate detection of human mtDNA transcription initiation sites (TIS) in the heavy and light strands revealed a novel transcription pausing site near the light strand TIS, upstream to the transcription-replication transition region, in conserved sequence block III (CSBIII). The transcription pausing index varied quantitatively among the cell lines. In addition to the human cell lines experiments, our experiments and analysis of non-human organisms enabled detection of previously unknown mtDNA TIS, pausing, and transcription termination sites with unprecedented accuracy. Whereas mammals (chimpanzee, rhesus macaque, rat, and mouse) showed a human-like mtDNA transcription pattern, the invertebrate pattern (Drosophila and C. elegans) profoundly diverged.
Overall design We re-mapped GRO-seq and PRO-seq data. We calibrated the parameters required to identify sites of transcription initiation, termination and pausing, which was first assigned to human samples but was applied to diverse non-human organisms.
Fastq files of the tested samples were trimmed by Trim-galore to reach a minimum read length of 30 nucleotides.
The trimmed fastq files of the samples were mapped against the entire revised mitochondrial genome using BWA-aln (-q=5, -l=20, -k=2, -t=1). BWA was used to convert SAI into SAM format, which in turn was converted into a BAM file and sorted using Samtools.
Samtools was used to generate VCF files of each sample (mpileup (-uf) command). Then, sample-specific mtDNA sequence was re-constructed for each of the analyzed samples using bcftools call (-c) (Samtools) in combination with vcf2fq from the package.
The Fastq files were uniquely re-mapped against the reconstructed sample-specific mtDNA using BWA-aln (-q=5, -l=32, -k=2, -t=1), and BAM files were generated again. Removal of low MAPQ reads was performed using the Samtools ‘view’ command (-F=1804, -q=30)
Coverage per base was calculated for a given sequence interval (separately for each strand) using Bedtolls (version 2.25), by the ‘genomecov’ command ('-d' and '-strand' options).
Since the mtDNA is a circular molecule and some reads may have been erroneously excluded we re-analyzed the Fastq files. To this end we remapped the reads to the sample-specific mtDNA sequence that was rearranged such, that the last 500 nucleotides of the standard mtDNA sequence were cut and pasted at the beginning of the sequence. Mapping was performed and read coverage at the former circle junction of the rearranged sequence was calculated and added to the previous mapping results
Genome build:
Homo sapiens: NC_012920.1; Revised Cambridge Reference Sequence ("rCRS")
Pan troglodytes: NC_001643.1
Macaca mulatta: NC_005943.1
Rattus norvegicus: NC_001665.2
Mus musculus: NC_005089.1
Drosophila melanogaster: NC_024511.2
Caenorhabditis elegans: NC_001328.1
Contributor(s) Mishmar D, Blumberg A
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Submission date Aug 17, 2016
Last update date Sep 30, 2016
Contact name Dan Mishmar
Organization name Ben Gurion University of the Negev
Department Life sciences
Street address Ben Gurion University of the Negev
City Beer Sheva
State/province Israel
ZIP/Postal code 84105
Country Israel
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BioProject PRJNA339241

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE85747_AC16_0_rep1_heavy.bedGraph.gz 57.3 Kb (ftp)(http) BEDGRAPH
GSE85747_AC16_0_rep1_light.bedGraph.gz 64.8 Kb (ftp)(http) BEDGRAPH
GSE85747_AC16_0_rep2_heavy.bedGraph.gz 57.0 Kb (ftp)(http) BEDGRAPH
GSE85747_AC16_0_rep2_light.bedGraph.gz 63.6 Kb (ftp)(http) BEDGRAPH
GSE85747_CD4_Mouse_heavy.bedGraph.gz 86.6 Kb (ftp)(http) BEDGRAPH
GSE85747_CD4_Mouse_light.bedGraph.gz 84.1 Kb (ftp)(http) BEDGRAPH
GSE85747_CD4_Rat_heavy.bedGraph.gz 83.2 Kb (ftp)(http) BEDGRAPH
GSE85747_CD4_Rat_light.bedGraph.gz 80.2 Kb (ftp)(http) BEDGRAPH
GSE85747_CD4_pro_heavy.bedGraph.gz 61.6 Kb (ftp)(http) BEDGRAPH
GSE85747_CD4_pro_light.bedGraph.gz 70.1 Kb (ftp)(http) BEDGRAPH
GSE85747_Celegans_N2_L3_GRO_heavy.bedGraph.gz 77.2 Kb (ftp)(http) BEDGRAPH
GSE85747_Celegans_N2_L3_GRO_light.bedGraph.gz 48.5 Kb (ftp)(http) BEDGRAPH
GSE85747_Chimp5-PI_heavy.bedGraph.gz 61.9 Kb (ftp)(http) BEDGRAPH
GSE85747_Chimp5-PI_light.bedGraph.gz 67.8 Kb (ftp)(http) BEDGRAPH
GSE85747_Chimp5_U_heavy.bedGraph.gz 61.6 Kb (ftp)(http) BEDGRAPH
GSE85747_Chimp5_U_light.bedGraph.gz 68.4 Kb (ftp)(http) BEDGRAPH
GSE85747_GM12004_GRO_heavy.bedGraph.gz 85.5 Kb (ftp)(http) BEDGRAPH
GSE85747_GM12004_GRO_light.bedGraph.gz 90.8 Kb (ftp)(http) BEDGRAPH
GSE85747_GM12004_PRO_heavy.bedGraph.gz 61.3 Kb (ftp)(http) BEDGRAPH
GSE85747_GM12004_PRO_light.bedGraph.gz 76.5 Kb (ftp)(http) BEDGRAPH
GSE85747_GM12750_GRO_heavy.bedGraph.gz 84.4 Kb (ftp)(http) BEDGRAPH
GSE85747_GM12750_GRO_light.bedGraph.gz 87.8 Kb (ftp)(http) BEDGRAPH
GSE85747_GM12878_gro_heavy.bedGraph.gz 88.4 Kb (ftp)(http) BEDGRAPH
GSE85747_GM12878_gro_light.bedGraph.gz 91.0 Kb (ftp)(http) BEDGRAPH
GSE85747_Hela_rep1_heavy.bedGraph.gz 62.9 Kb (ftp)(http) BEDGRAPH
GSE85747_Hela_rep1_light.bedGraph.gz 73.0 Kb (ftp)(http) BEDGRAPH
GSE85747_Hela_rep2_heavy.bedGraph.gz 90.2 Kb (ftp)(http) BEDGRAPH
GSE85747_Hela_rep2_light.bedGraph.gz 87.9 Kb (ftp)(http) BEDGRAPH
GSE85747_IMR90_rep1_heavy.bedGraph.gz 65.6 Kb (ftp)(http) BEDGRAPH
GSE85747_IMR90_rep1_light.bedGraph.gz 67.2 Kb (ftp)(http) BEDGRAPH
GSE85747_MCF7_0_rep1_heavy.bedGraph.gz 64.8 Kb (ftp)(http) BEDGRAPH
GSE85747_MCF7_0_rep1_light.bedGraph.gz 65.4 Kb (ftp)(http) BEDGRAPH
GSE85747_MCF7_0_rep2_heavy.bedGraph.gz 58.2 Kb (ftp)(http) BEDGRAPH
GSE85747_MCF7_0_rep2_light.bedGraph.gz 69.2 Kb (ftp)(http) BEDGRAPH
GSE85747_Rhesus3_PI_heavy.bedGraph.gz 67.2 Kb (ftp)(http) BEDGRAPH
GSE85747_Rhesus3_PI_light.bedGraph.gz 73.0 Kb (ftp)(http) BEDGRAPH
GSE85747_U20S_C_rep1_heavy.bedGraph.gz 90.6 Kb (ftp)(http) BEDGRAPH
GSE85747_U20S_C_rep1_light.bedGraph.gz 91.0 Kb (ftp)(http) BEDGRAPH
GSE85747_U20S_C_rep2_heavy.bedGraph.gz 86.1 Kb (ftp)(http) BEDGRAPH
GSE85747_U20S_C_rep2_light.bedGraph.gz 86.0 Kb (ftp)(http) BEDGRAPH
GSE85747_drosophila_PRO_01A_heavy.bedGraph.gz 87.3 Kb (ftp)(http) BEDGRAPH
GSE85747_drosophila_PRO_01A_light.bedGraph.gz 78.2 Kb (ftp)(http) BEDGRAPH
GSE85747_drosophila_PRO_01N_heavy.bedGraph.gz 81.9 Kb (ftp)(http) BEDGRAPH
GSE85747_drosophila_PRO_01N_light.bedGraph.gz 77.1 Kb (ftp)(http) BEDGRAPH
GSE85747_jurkat_pro_heavy.bedGraph.gz 74.2 Kb (ftp)(http) BEDGRAPH
GSE85747_jurkat_pro_light.bedGraph.gz 81.6 Kb (ftp)(http) BEDGRAPH
GSE85747_k562_gro_heavy.bedGraph.gz 85.7 Kb (ftp)(http) BEDGRAPH
GSE85747_k562_gro_light.bedGraph.gz 86.4 Kb (ftp)(http) BEDGRAPH
GSE85747_k562_pro_heavy.bedGraph.gz 101.9 Kb (ftp)(http) BEDGRAPH
GSE85747_k562_pro_light.bedGraph.gz 103.5 Kb (ftp)(http) BEDGRAPH
GSE85747_readme.xls.gz 8.2 Kb (ftp)(http) XLS
Processed data are available on Series record

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