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DNA‐Binding Specificity of Cardiac Transcription Factor Complex Formed by NKX2‐5 and TBX5
Author(s) -
RodríguezRíos Jessica,
RosadoRodríguez Emili,
RodríguezMartínez José
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.03485
Subject(s) - transcription factor , dna , gene , microbiology and biotechnology , biology , binding site , dna binding protein , dna binding site , transcription (linguistics) , in vitro , computational biology , genetics , chemistry , gene expression , promoter , linguistics , philosophy
Transcription factors (TFs) are sequence‐specific DNA‐binding proteins responsible for cellular differentiation and development. Eukaryotic TFs are known for binding DNA as multimeric complexes to regulate gene expression. Transcription factors NKX2‐5 and TBX5 are fundamental elements of the gene regulatory networks that control mammalian heart development. More importantly, NKX2‐5 and TBX5 can bind to DNA as a cooperative complex. Our aim was to identify the intrinsic DNA‐binding preferences and the motif spacing and orientation pattern of the NKX2‐5:TBX5 heterodimer complex. In this study, we expressed full‐length GST‐tagged NKX2‐5 and TBX5 using a wheat germ cell‐free system. We successfully determined the in vitro DNA‐binding specificity of the NKX2‐5:TBX5 complex using Systematic Evolution of Ligands by Exponential Enrichment (SELEX‐seq). Our preliminary results shows NKX2‐5:TBX5 binding in a head‐to‐tail orientation with 0bp and 3bps spacing between core binding sites: GGTGT for TBX5 and CACTT for NKX2‐5. The spacing preference is consistent with ChIP‐exo data from cardiomyocytes and cardiac precursors. Our results highlight the utility of using in vitro techniques to determined DNA‐binding specificity to understand genomic binding. The findings of our study will help understand the spatial and temporal gene regulation rules involved in normal heart development.