A generic tool for transcription factor target gene discovery in Arabidopsis cell suspension cultures based on tandem chromatin affinity purification.
Genome-wide identification of transcription factor (TF) binding sites is pivotal to our understanding of gene expression regulation. Although much progress has been made in the determination of potential binding regions of proteins by chromatin immunoprecipitation, this method has some inherent limitations regarding DNA enrichment efficiency and antibody necessity. Here, we report an alternative strategy for assaying in vivo TF-DNA binding in Arabidopsis (Arabidopsis thaliana) cells by tandem chromatin affinity purification (TChAP). Evaluation of TChAP using the E2Fa TF and comparison with traditional chromatin immunoprecipitation and single chromatin affinity purification illustrates the suitability of TChAP and provides a resource for exploring the E2Fa transcriptional network. Integration with transcriptome, cis-regulatory element, functional enrichment, and coexpression network analyses demonstrates the quality of the E2Fa TChAP sequencing data and validates the identification of new direct E2Fa targets. TChAP enhances both TF target mapping throughput, by circumventing issues related to antibody availability, and output, by improving DNA enrichment efficiency.
Department of Plant Systems Biology, VIB, B-9052 Ghent, Belgium.
Aurine
A
Verkest
Thomas
T
Abeel
Ken S
KS
Heyndrickx
Jelle
J
Van Leene
Christa
C
Lanz
Eveline
E
Van De Slijke
Nancy
N
De Winne
Dominique
D
Eeckhout
Geert
G
Persiau
Frank
F
Van Breusegem
Dirk
D
Inzé
Klaas
K
Vandepoele
Geert
G
De Jaeger
D017360:Arabidopsis / Q000166:cytology* / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D019921:Biotinylation; D002478:Cells, Cultured; D002843:Chromatin / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation; D002846:Chromatography, Affinity / Q000379:methods*; D050684:E2F Transcription Factors / Q000378:metabolism*; D017343:Genes, Plant; D056726:Genetic Association Studies / Q000379:methods*; D006639:Histidine / Q000378:metabolism; D058977:Molecular Sequence Annotation; D059372:Nucleotide Motifs / Q000235:genetics; D009842:Oligopeptides / Q000378:metabolism; D030821:Plants, Genetically Modified; D011485:Protein Binding / Q000235:genetics; D017422:Sequence Analysis, DNA
D029681:Arabidopsis Proteins; D002843:Chromatin; D050684:E2F Transcription Factors; C496141:E2Fa protein, Arabidopsis; C471213:His-His-His-His-His-His; D009842:Oligopeptides; D006639:Histidine