title: High-throughput assessment of context-dependent effects of chromatin proteins.
abstract: Chromatin proteins control gene activity in a concerted manner. We developed a high-throughput assay to study the effects of the local chromatin environment on the regulatory activity of a protein of interest. The assay combines a previously reported multiplexing strategy based on barcoded randomly integrated reporters with Gal4-mediated tethering. We applied the assay toDrosophilaheterochromatin protein 1a (HP1a), which is mostly known as a repressive protein but has also been linked to transcriptional activation.
  
  
  
  Recruitment to over 1000 genomic locations revealed that HP1a is a potent repressor able to silence even highly expressing reporter genes. However, the local chromatin context can modulate HP1a function. In pericentromeric regions, HP1a-induced repression was enhanced by twofold. In regions marked by a H3K36me3-rich chromatin signature, HP1a-dependent silencing was significantly decreased. We found no evidence for an activating function of HP1a in our experimental system. Furthermore, we did not observe stable transmission of repression over mitotic divisions after loss of targeted HP1a.
  
  
  
  The multiplexed tethered reporter assay should be applicable to a large number of chromatin proteins and will be a useful tool to dissect combinatorial regulatory interactions in chromatin.
authors: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors: Laura
authors: L
authors: Brueckner
authors: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors: Joris
authors: J
authors: van Arensbergen
authors: Division of Molecular Genetics, Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors: Waseem
authors: W
authors: Akhtar
authors: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors: Ludo
authors: L
authors: Pagie
authors: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, The Netherlands ; Department of Cell Biology, Erasmus University Medical Center, Rotterdam, The Netherlands.
authors: Bas
authors: 0000-0003-2649-1939
authors: B
authors: van Steensel
mesh_terms: D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics* / Q000378:metabolism; D029721:Drosophila Proteins / Q000235:genetics* / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism; D020868:Gene Silencing; D006657:Histones / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D061805:Transcription Elongation, Genetic; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D029721:Drosophila Proteins; C557617:GAL4 protein, Drosophila; D006657:Histones; D014157:Transcription Factors
keywords: Chromatin states; Drosophila melanogaster; Epigenetic memory; HP1a; Transcriptional repression