Global Egr1-miRNAs Binding Analysis in PMA-Induced K562 Cells Using ChIP-Seq

Wei Wang,Xueying Xie,Chao Tang,Jing Tu,Dequang Zhou,Zuhong Lu,Xiaolong Shi,Qinyu Ge

doi:10.1155/2010/867517

Abstract

Although much is known about microRNAs' regulation in gene expression and their contributions in cell fate, to date, globally lineage-(cell-) specific identification of the binding events between a transcription factor and its targeting microRNA genes is still waiting for elucidation. In this paper, we performed a ChIP-Seq experiment to find the targeting microRNA genes of a transcription factor, Egr1, in human erythroleukemia cell line K562. We found Egr1 binding sites near the promoters of 124 distinct microRNA genes, accounting for about 42% of the miRNAs which have high-confidence predicted promoters (294). We also found EGR1 bind to another 63 pre-miRNAs. We chose 12 of the 187 microRNAs with Egr1 binding sites to perform ChIP-PCR assays and the positive binding signal from ChIP-PCR confirmed the ChIP-Seq results. Our experiments provide the first global binding profile between Egr1 and its targeting microRNA genes in PMA-treated K562 cells, which may facilitate the understanding of pathways controlling microRNA biology in this specific cell line.

Highlights

MicroRNAs are a family of ∼22-nucleotide small noncoding RNAs in eukaryotes and mainly involve in regulation at posttranscriptional level by translational repression or degradation their target mRNAs [1, 2]
A recent published miRNAs expression profile in the similar cell condition facilitated to analyze the expression changes of those miRNAs which are bound by Egr1 in Phorbol-12-myristate 13-acetate (PMA)-induced MK differentiation of K562 cells [11]
We found that hsa-mir-142 has 45 consensus sites dispersed among its promoters and correspondingly, 10 Egr1 binding regions identified by Chromatin Immunoprecipitation (ChIP)-Seq were enriched with reads count above nine. miR-142 is one of five miRNAs which are highly specific for hematopoietic cells according expression comparison [35]

Summary

Introduction

MicroRNAs (miRNAs) are a family of ∼22-nucleotide small noncoding RNAs in eukaryotes and mainly involve in regulation at posttranscriptional level by translational repression or degradation their target mRNAs [1, 2]. MiRNAs have been reported to regulate hematopoietic lineage differentiation, angiogenesis, cell adhesion and so on [1, 5]. K562 is a cell line deriving from chronic myeloid leukemia, which is a common progenitor of megakaryocytic and erythroid lineages of the hematopoietic stem cell differentiation. MiRNAs have been found to play a key role in K562 differentiation. The changes of miRNAs expression level gave clues for their functions in hematopoietic lineage differentiation but a more detailed regulation pathway is anticipated to be understood: by which targets miRNAs realize their functions in hematopoietic differentiation and miRNAs are subjected to which factors controlling their transcription? Garzon et al confirmed that miR-130 targets the transcription factor MAFB and participates in MK differentiation by up-regulating its expression level in CD34+ hematopoietic progenitors [14]. Navarro et al found that independently of p53, miR-34a directly regulates expression

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Conclusion