Global binding pattern of the Wilms' tumor gene 1 (WT1) +17AA -KTS isoform in leukemic cells

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Abstract

The aim of this study was to investigate the global DNA-binding pattern of Wilms' tumor gene 1 (WT1) in leukemic cells. Clinical and preclinical data indicate the zinc finger transcription factor WT1 as an oncogene, but the full target gene repertoire of WT1 in leukemic cells has not been previously characterized. The -KTS isoforms (excluding the three amino acid (KTS) insert between zinc finger three and four) are considered as the most efficient DNA-binders. Among these, the 17AA isoform (including 17 amino acids encoded by exon 5) is the most abundant one. To specifically analyze the DNA-binding of WT1(+17AA/-KTS) in leukemic cells, we generated a K562 clone that stably expressed BIO-tagged WT1(+17AA/-KTS), as well as the biotinylating enzyme Bir A. From the cells chromatin immunoprecipitation (ChIP) by streptavidin capture was performed followed by sequencing with a minimum of 50 million reads per sample. After alignment to the genome and peak calling, peaks were characterized and compared to available K562 tracks in the ENCODE database. We found that 45% of identified WT1(+17AA/-KTS) peaks are in the proximity of transcription start sites (promoter area, first exon or first intron) of target genes, whereas only 11% of randomized peaks were found here. Within the peaks we show strong enrichment for three different previously published WT1-binding motifs. Comparison to ENCODE tracks showed that WT1(+17AA/-KTS) peaks are in close proximity to binding sites of other transcription factors, to histone marks for actively transcribed genes, and to binding sites of chromatin modifiers. Considering peaks within promoters and gene bodies only (for safe assignment to a target gene), Gene Ontology (GO) analysis revealed enrichment of GO groups important for proliferation, cell death, embryonic development, and cell motility. In conclusion, WT1(+17AA/-KTS) binds close to transcription start sites in areas of active transcription. The target genes implicated in proliferation, cell death, cell signaling and motility adds to the growing evidence of WT1 as an effector gene in leukemia.

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Subject classification (UKÄ) – MANDATORY

  • Medical Biotechnology

Keywords

  • amino acid, endogenous compound, histone, streptavidin, transcription factor, WT1 protein, zinc finger protein, binding site, cell death, cell motility, cell proliferation, chemical binding, chromatin immunoprecipitation, controlled clinical trial, controlled study, data base, DNA binding, DNA transcription, embryo cell, embryo development, gene expression, gene ontology, human, human cell, intron, leukemia cell, oncogene, preclinical study, promoter region, randomized controlled trial, transcription initiation site, transcription regulation
Original languageEnglish
Article numberAbstract nr 2005
JournalCancer Research
Volume76
Issue number14 Suppl.
Publication statusPublished - 2016 Jun 2
Publication categoryResearch
Peer-reviewedYes
EventAmerican Association for Cancer Research (AACR) 107th Annual Meeting 2016 - New Orleans, United States
Duration: 2016 Apr 162016 Apr 20