Mechanismen die generelle und zelltypspezifische DNA-Bindung des Masterregulators PU.1 kontrollieren
Biochemie
Zusammenfassung der Projektergebnisse
Aim of this collaborative project was to combine high-throughput-sequencing, computational, biochemical and functional analyses to address the question, how the binding of the myeloid and B cell-specific transcription factor PU.1 to genomic DNA is controlled. We used both in-vitro and in-vivo analyses to characterize the binding of PU.1 to chromatinized DNA. Our in-vitro studies show that PU.1 binds to chromatinized DNA harboring PU.1 binding sites around the nucleosome entry exit (up to 16 bp inside) but not around the dyad axis. PU.1 binding is also inhibited by DNA methylation or hydroxymethylation, suggesting that the epigenetic/chromatin landscapes of individual cell types influence the binding of PU.1. This view is also supported through our mapping of PU.1 binding sites in a comprehensive panel of cell types, suggesting that only up to 15 % of putative binding sites are bound in at least one cell type. To further study the initial binding of PU.1 in “untouched” cells, we developed an RNA-transfection-based model which allowed the functional dissection of PU.1 protein domains that are required for the stable binding of PU.1. Finally, we show that the domain that is required for de-novo bound and remodeled sites is interacting with the SWI/SNF complex.
Projektbezogene Publikationen (Auswahl)
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Characterizing higher order structures of chromatin in human cells.
(bioRxiv.org - the preprint server for Biology)
Uwe Schwartz, Attila Németh, Sarah Diermeier, Josef Exler, Stefan Hansch, Rodrigo Maldonado, Leonhard Heizinger, Rainer Merkl, Gernot Läengst
