title: Mapping and functional characterisation of a CTCF-dependent insulator element at the 3' border of the murine Scl transcriptional domain.
abstract: The Scl gene encodes a transcription factor essential for haematopoietic development. Scl transcription is regulated by a panel of cis-elements spread over 55 kb with the most distal 3' element being located downstream of the neighbouring gene Map17, which is co-regulated with Scl in haematopoietic cells. The Scl/Map17 domain is flanked upstream by the ubiquitously expressed Sil gene and downstream by a cluster of Cyp genes active in liver, but the mechanisms responsible for delineating the domain boundaries remain unclear. Here we report identification of a DNaseI hypersensitive site at the 3' end of the Scl/Map17 domain and 45 kb downstream of the Scl transcription start site. This element is located at the boundary of active and inactive chromatin, does not function as a classical tissue-specific enhancer, binds CTCF and is both necessary and sufficient for insulator function in haematopoietic cells in vitro. Moreover, in a transgenic reporter assay, tissue-specific expression of the Scl promoter in brain was increased by incorporation of 350 bp flanking fragments from the +45 element. Our data suggests that the +45 region functions as a boundary element that separates the Scl/Map17 and Cyp transcriptional domains, and raise the possibility that this element may be useful for improving tissue-specific expression of transgenic constructs.
authors.affiliation: Department of Haematology, Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom. gf246@cam.ac.uk
authors.forename: George A
authors.initials: GA
authors.lastname: Follows
authors.forename: Rita
authors.initials: R
authors.lastname: Ferreira
authors.forename: Mary E
authors.initials: ME
authors.lastname: Janes
authors.forename: Dominik
authors.initials: D
authors.lastname: Spensberger
authors.forename: Francesco
authors.initials: F
authors.lastname: Cambuli
authors.forename: Amy F
authors.initials: AF
authors.lastname: Chaney
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Kinston
authors.forename: Josette R
authors.initials: JR
authors.lastname: Landry
authors.forename: Anthony R
authors.initials: AR
authors.lastname: Green
authors.forename: Berthold
authors.initials: B
authors.lastname: Göttgens
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics* / Q000502:physiology*; D001665:Binding Sites; D000076246:CCCTC-Binding Factor; D047369:Chromatin Immunoprecipitation; D002874:Chromosome Mapping / Q000379:methods; D003850:Deoxyribonuclease I / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D017930:Genes, Reporter; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D008099:Liver / Q000378:metabolism; D051379:Mice; D005810:Multigene Family; D020411:Oligonucleotide Array Sequence Analysis; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000502:physiology*; D012097:Repressor Proteins / Q000235:genetics*; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1; D014158:Transcription, Genetic; D019076:Transgenes
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D000076246:CCCTC-Binding Factor; D003850:Deoxyribonuclease I; D011518:Proto-Oncogene Proteins; D012097:Repressor Proteins; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1; C000619697:CTCF protein, human; C000619698:Ctcf protein, mouse; C489366:Tal1 protein, mouse