Bookmarking by “non-pioneer” transcription factors during liver development establishes competence for future gene activation

Transcription factor binding to enhancer and promoter regions is critical for gene activation. To understand how cell-specific gene expression patterns are generated, we studied the developmental timing of association of two prominent hepatic transcription factors with gene regulatory regions. We found that most individual binding events displayed extraordinarily high temporal variations during liver development. Early and persistent binding is necessary but not sufficient for gene activation. Stable gene expression patterns are mainly generated by the combinatorial activity of multiple transcription factors, which mark regulatory regions long before activation and promote progressive broadening of active chromatin domains. Both, temporally stable and dynamic binding events contribute to the developmental maturation of active promoter configurations. The results reveal a developmental “bookmarking” function of transcription factors, and illuminate remarkable parallels between the principles employed for gene activation during development, evolution and upon mitotic exit. Overall design: Chromatin immunoprecipitation assays were performed in formaldehyde-crosslinked mouse liver extracts using antibodies recognizing C/ABPa or HNF4a or RNA-Polymerase-II. The extracts were prepared from mouse embryonic day 15.5, 18.5, newborn, postnatal day 14 and postnatal day 60. Each ChIP-seq assay were performed with 2 biological replicates. mRNAs for RNA-seq analyses were prepared from E18.5 livers of wild type (WT) or CEBPalox/lox-Alfp-Cre or HNF4lox/lox-Alfp-Cre mice.