Dataset Information

Identification of ChIP-seq and RIME grade antibodies for Estrogen Receptor alpha.

ABSTRACT: Estrogen Receptor alpha (ERα) plays a major role in most breast cancers, and it is the target of endocrine therapies used in the clinic as standard of care for women with breast cancer expressing this receptor. The two methods ChIP-seq (chromatin immunoprecipitation coupled with deep sequencing) and RIME (Rapid Immunoprecipitation of Endogenous Proteins) have greatly improved our understanding of ERα function during breast cancer progression and in response to anti-estrogens. A critical component of both ChIP-seq and RIME protocols is the antibody that is used against the bait protein. To date, most of the ChIP-seq and RIME experiments for the study of ERα have been performed using the sc-543 antibody from Santa Cruz Biotechnology. However, this antibody has been discontinued, thereby severely impacting the study of ERα in normal physiology as well as diseases such as breast cancer and ovarian cancer. Here, we compare the sc-543 antibody with other commercially available antibodies, and we show that 06-935 (EMD Millipore) and ab3575 (Abcam) antibodies can successfully replace the sc-543 antibody for ChIP-seq and RIME experiments.

SUBMITTER: Glont SE

PROVIDER: S-EPMC6457525 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Identification of ChIP-seq and RIME grade antibodies for Estrogen Receptor alpha.

Glont Silvia-E SE Papachristou Evangelia K EK Sawle Ashley A Holmes Kelly A KA Carroll Jason S JS Siersbaek Rasmus R

PloS one 20190410 4

Estrogen Receptor alpha (ERα) plays a major role in most breast cancers, and it is the target of endocrine therapies used in the clinic as standard of care for women with breast cancer expressing this receptor. The two methods ChIP-seq (chromatin immunoprecipitation coupled with deep sequencing) and RIME (Rapid Immunoprecipitation of Endogenous Proteins) have greatly improved our understanding of ERα function during breast cancer progression and in response to anti-estrogens. A critical componen ...[more]

PMID: 30970003

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Project description:BackgroundThe largest group of patients with breast cancer are estrogen receptor-positive (ER+) type. The estrogen receptor acts as a transcription factor and triggers cell proliferation and differentiation. Hence, investigating ER-DNA interaction genomic regions can help identify genes directly regulated by ER and understand the mechanism of ER action in cancer progression.MethodsIn the present study, we employed a workflow to do a meta-analysis of ChIP-seq data of ER+ cell lines stimulated with 10 nM and 100 nM of E2. All publicly available data sets were re-analyzed with the same platform. Then, the known and unknown batch effects were removed. Finally, the meta-analysis was performed to obtain meta-differentially bound sites in estrogen-treated MCF7 cell lines compared to vehicles (as control). Also, the meta-analysis results were compared with the results of T47D cell lines for more precision. Enrichment analyses were also employed to find the functional importance of common meta-differentially bound sites and associated genes among both cell lines.ResultsRemarkably, POU5F1B, ZNF662, ZNF442, KIN, ZNF410, and SGSM2 transcription factors were recognized in the meta-analysis but not in individual studies. Enrichment of the meta-differentially bound sites resulted in the candidacy of pathways not previously reported in breast cancer. PCGF2, HNF1B, and ZBED6 transcription factors were also predicted through the enrichment analysis of associated genes. In addition, comparing the meta-analysis results of both ChIP-seq and RNA-seq data showed that many transcription factors affected by ER were up-regulated.ConclusionThe meta-analysis of ChIP-seq data of estrogen-treated MCF7 cell line leads to the identification of new binding sites of ER that have not been previously reported. Also, enrichment of the meta-differentially bound sites and their associated genes revealed new terms and pathways involved in the development of breast cancer which should be examined in future in vitro and in vivo studies.

Dataset Information

Identification of ChIP-seq and RIME grade antibodies for Estrogen Receptor alpha.

Publications

Identification of ChIP-seq and RIME grade antibodies for Estrogen Receptor alpha.

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