Zebrafish promoterome based on Haberle et al. and Nepal et al.
Tracks display features of zebrafish promoter utilization before, during and after the mid-blastula transition, MBT, representing maternal stages and zygotic stages. RNA-seq was performed in four stages. Time course of CAGE is shown across twelve stages. H3K4me3 histone mark is presented for four stages in teal. H2A.Z ChIP-seq during MBT is shown in green. ChIP subtracted signal shows nucleosome positioning, as described in Haberle et al..
All stages are named according to ZFIN recommendations, note references indicating prim-6 have been renamed here to prim-5 (embryos collected were an indistinguishable mixture of prim-5 and prim-6).
Display Conventions and Configuration
- CAGE transcription start sites (CTSSs) - normalized (tag per million: tpm) tag counts at tag's 5' end only, for a particular stage and strand
- CAGE tag clusters (TCs) - clusters of normalized CAGE tags: 0.1-0.9 quantile range (as described in Nepal et al.) shown as a block, dominant CTSS position shown as a thick bar
- RNA-seq coverage - read depth at every genomic position
- ChIP-seq raw coverage - read depth based on reads extended to 200bp
- ChIP-seq subtracted coverage - depth of unextended reads on reverse strand is subtracted from depth of unextended reads on forward strand (see Haberle et al.)
CAGE and RNA-seq
Embryos were collected from wild type AB zebrafish in 10 minutes intervals to allow accurate staging. Total RNA was extracted for each stage, using TriZol Reagent (Invitrogen). CAGE libraries were prepared using total RNA and sequenced on Illumina GA II. Sequenced reads were mapped to the reference zebrafish genome (Zv9/danRer7 assembly) using Bowtie with default parameters allowing up to two mismatches and keeping only uniquely mapped reads. Mismatching G at the first (5') position of CAGE reads were removed and tag counts were normalized using power law normalization. See Nepal et. al. for more details.
ChIP experiments were carried out using the ChIP-IT Express Enzymatic kit (Active Motif) in line with the manufacturer's instructions. Chromatin was prepared using 5000 embryos (512-cell), 3000 embryos (oblong), 1500 embryos (30% epiboly) and 200 embryos (Prim-5/6). Anti-H3K4me3 (Abcam ab8580) or anti-H2A.Z (Abcam ab4174) were used for immunoprecipitation. Sequenced reads were mapped to the reference zebrafish genome (Zv9/danRer7 assembly) using Bowtie with default parameters allowing up to two mismatches and keeping only uniquely mapped reads. Significantly enriched regions (peaks) were detected using model-based analysis for ChIP-seq (MACS) with default parameters.
Haberle V, Li N, Hadzhiev Y et al. Two independent transcription initiation codes overlap on vertebrate core promoters. Nature 2014;507(7492):381-5. PMID: 24531765
Nepal C, Hadzhiev Y, Previti C et al. Dynamic regulation of the transcription initiation landscape at single nucleotide resolution during vertebrate embryogenesis. Genome Res. 2013;23(11):1938-50. PMID: 24002785; PMCID: PMC3814893
For the individual contributions in the data generation and analyses see references above.