Open Access Research

Genome-wide investigation of in vivo EGR-1 binding sites in monocytic differentiation

Atsutaka Kubosaki1, Yasuhiro Tomaru12, Michihira Tagami1, Erik Arner1, Hisashi Miura12, Takahiro Suzuki12, Masanori Suzuki12, Harukazu Suzuki1 and Yoshihide Hayashizaki12*

Author Affiliations

1 RIKEN Omics Science Center, RIKEN Yokohama Institute 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan

2 International Graduate School of Arts and Sciences, Yokohama City University, 1-7-29 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045, Japan

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Genome Biology 2009, 10:R41  doi:10.1186/gb-2009-10-4-r41

Published: 19 April 2009

Additional files

Additional data file 1:

(a) siRNA mediated knockdown of EGR-1 mRNA. EGR-1 mRNA were quantified using quantitative RT-PCR. EGR-1 mRNA levels were normalized to GAPDH mRNA and are presented relative to RNA levels in mock cells. RNA levels are representative of four independent experiments. (b) Effect of siRNA on EGR-1 in THP-1 differentiation. Phase contrast and fluorescence images were taken at the same time. Photographs show transfect efficiency indicated by Alexa Fluor 555 (upper) and typical morphological changes in EGR-1 or control siRNA transfected THP-1 cells at 48 hours after PMA stimulation (lower). The white arrows indicate differentiating THP-1 cells. (c) A sample of sonicated DNA. The sonication conditions were optimized to achieve enrichment of fragments between 150 and 600 bp in length.

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Additional data file 2:

Myeloid related genes within predicted EGR-1 targets.

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Additional data file 3:

PCR primers were designed for nine regions in selected clusters and six negative regions without enrichment in CpG islands. Data are relative fold enrichments, calculated by determining the apparent immunoprecipitation efficiency and normalized to the level observed at a control region (mean ± standard deviation, n = 2).

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Additional data file 4:

Schematic Venn diagram representing the overlaps between EGR-1 binding sites, H3K9ac domains of each biological replicate and CAGE tag clusters.

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Additional data file 5:

SP1 protein levels over a time course following PMA stimulation were observed by western blot analysis using a specific polyclonal antibody.

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Additional data file 6:

Venn diagram showing the overlaps between EGR-1 binding sites and SP1 binding sites of each biological replicate.

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Additional data file 7:

Quantile normalized NAB1 and NAB2 transcript levels were produced by Illumina Human Sentrix-6 bead chips v.2.

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Additional data file 8:

deepCAGE tag clusters indicate transcriptional start sites in THP-1 differentiation.

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Additional data file 9:

Real-time PCR primers for ChIP samples.

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