ChIP-chip by array of human HeLa cells exposed to 1% oxygen (hypoxia) or 21% oxygen (normoxic control) to study the DNA binding profile of endogenous HIF1A on proximal promoters.
ABSTRACT: DNA binding profiling of endogenous HIF1A on proximal promoters in human HeLa cells exposed to 1% oxygen (hypoxia), using normoxic cells (21% oxygen) as reference.
Biological background: The Hypoxia Inducible Factor Family of transcription factors is proposed as the main orchestrator of the cellular response to hypoxia. HIFs are heterodimers of a HIF alpha and a HIF beta subunit. HIF alpha protein stability is regulated by oxygen-dependent proteasomal degradation, and hence HIFs are strongly stabilized in hypoxia.
Purpose of the study: a number of HIF1 ChIP-chip studies have been reported, employing various cell types, array platforms and HIF antibodies, and the overlap of HIF binding locations in these studies is relatively small. The aim of this study was to characterize HIF1 binding in an additional cell line (HeLa), and employing a different HIFalpha antibody.
Experimental design: We conducted a total of six hybridizations employing four biological replicates. For two biological replicates, we performed dye-swap technical replicate experiments.
Results summary: We identified 55 HIF binding locations in HeLa cells (FDR<2%). While this number is relatively low compared to previous studies, presumably due to limiting antibody sensitivity, the overlap with data from other cell lines is comparable to our HeLa data.
The transcriptional response driven by Hypoxia-inducible factor (HIF) is central to the adaptation to oxygen restriction. Despite recent characterization of genome-wide HIF DNA binding locations and hypoxia-regulated transcripts in different cell types, the molecular bases of HIF target selection remain unresolved. Herein, we combined multi-level experimental data and computational predictions to identify sequence motifs that may contribute to HIF target selectivity. We obtained a core set of bo ...[more]
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