Developmentally Regulated Recruitment of Transcription Factors and Chromatin Modification Activities to Chicken Lysozyme cis-Regulatory Elements In Vivo

Abstract

Expression of the chicken lysozyme gene is upregulated during macrophage differentiation and reaches its highest level in bacterial lipopolysaccharide (LPS)-stimulated macrophages. This is accompanied by complex alterations in chromatin structure. We have previously shown that chromatin fine-structure alterations precede the onset of gene expression in macrophage precursor cells and mark the lysozyme chromatin domain for expression later in development. To further examine this phenomenon and to investigate the basis for the differentiation-dependent alterations of lysozyme chromatin, we studied the recruitment of transcription factors to the lysozyme locus in vivo at different stages of myeloid differentiation. Factor recruitment occurred in several steps. First, early-acting transcription factors such as NF1 and Fli-1 bound to a subset of enhancer elements and recruited CREB-binding protein. LPS stimulation led to an additional recruitment of C/EBPβ and a significant change in enhancer and promoter structure. Transcription factor recruitment was accompanied by specific changes in histone modification within the lysozyme chromatin domain. Interestingly, we present evidence for a transient interaction of transcription factors with lysozyme chromatin in lysozyme-nonexpressing macrophage precursors, which was accompanied by a partial demethylation of CpG sites. This indicates that a partially accessible chromatin structure of lineage-specific genes is a hallmark of hematopoietic progenitor cells.

ACKNOWLEDGMENTS

We thank Peter Cockerill and George Follows for comments on the manuscript. P.L thanks Thierry Grange, Valerie Orlando, and the team from the EMBO course on DNA-Protein Interaction in vivo for their expert training in chromatin immunoprecipitation technology. We also thank Achim Leutz and Elisabeth Kowentz-Leutz for anti-C/EBPβ (NF-M) antibodies and Naoko Tanese for anti-NF1 antibodies.

Research in C. Bonifer's laboratory is supported by grants from the Wellcome Trust, the BBSRC, the Leukemia Research Fund, and the City of Hope Medical Centre. Nicola Wilson is a recipient of a Leukemia Research Fund summer studentship grant.