title: Genomic selection identifies vertebrate transcription factor Fezf2 binding sites and target genes.
abstract: Identification of transcription factor targets is critical to understanding gene regulatory networks. Here, we uncover transcription factor binding sites and target genes employing systematic evolution of ligands by exponential enrichment (SELEX). Instead of selecting randomly synthesized DNA oligonucleotides as in most SELEX studies, we utilized zebrafish genomic DNA to isolate fragments bound by Fezf2, an evolutionarily conserved gene critical for vertebrate forebrain development. This is, to our knowledge, the first time that SELEX is applied to a vertebrate genome. Computational analysis of bound genomic fragments predicted a core consensus binding site, which identified response elements that mediated Fezf2-dependent transcription both in vitro and in vivo. Fezf2-bound fragments were enriched for conserved sequences. Surprisingly, ∼20% of these fragments overlapped well annotated protein-coding exons. Through loss of function, gain of function, and chromatin immunoprecipitation, we further identified and validated eomesa/tbr2 and lhx2b as biologically relevant target genes of Fezf2. Mutations in eomesa/tbr2 cause microcephaly in humans, whereas lhx2b is a critical regulator of cell fate and axonal targeting in the developing forebrain. These results demonstrate the feasibility of employing genomic SELEX to identify vertebrate transcription factor binding sites and target genes and reveal Fezf2 as a transcription activator and a candidate for evaluation in human microcephaly.
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, California 94158, USA.
authors.forename: Lishan
authors.initials: L
authors.lastname: Chen
authors.forename: Jiashun
authors.initials: J
authors.lastname: Zheng
authors.forename: Nan
authors.initials: N
authors.lastname: Yang
authors.forename: Hao
authors.initials: H
authors.lastname: Li
authors.forename: Su
authors.initials: S
authors.lastname: Guo
mesh_terms: D000818:Animals; D004247:DNA / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008831:Microcephaly / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D016548:Prosencephalon / Q000196:embryology*; D020218:Response Elements / Q000502:physiology*; D052156:SELEX Aptamer Technique; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics; D029961:Zebrafish Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D009419:Nerve Tissue Proteins; D014157:Transcription Factors; D029961:Zebrafish Proteins; D004247:DNA