title: Cell type and context-specific function of PLAG1 for IGF2 P3 promoter             activity.
abstract: The fetal transcription factor PLAG1 is found to be overexpressed in cancers,             and has been suggested to bind the insulin like growth factor 2 (IGF2) P3 promoter,             and to activate the IGF2 gene. The expression of IGF2 has partly been linked to             loss of CTCF-dependent chromatin insulator function at the H19 imprinting control             region (ICR). We investigated the role of PLAG1 for IGF2 regulation in Hep3B and             JEG-3 cell lines. Chromatin immunoprecipitation revealed cell type-specific binding             of PLAG1 to the IGF2 P3 promoter, which was substantially insensitive to recombinant             PLAG1 overexpression in the endogenous context. We hypothesized that the H19 chromatin             insulator may be involved in the cell type-specific PLAG1 response. By using a             GFP reporter gene/insulator assay plasmid construct with and without the H19 ICR             and/or an SV40 enhancer, we confirm that the effect of the insulator is specifically             associated with the activity of the IGF2 P3 promoter in the GFP reporter system,             and furthermore, that the reporter insulator is functional in JEG-3 but not in             Hep3B cells. FACS analysis was used to assess the function of PLAG1 in low endogenously             expressing, but Zn-inducible stable PLAG1 expressing JEG-3 cell clones. Considerable             increase in IGF2 expression upon PLAG1 induction with a partial insulator overriding             activity was found using the reporter constructs. This is in contrast to the effect             of the endogenous IGF2 gene which was insensitive to PLAG1 expression in JEG-3,             while modestly induced the already highly expressed IGF2 gene in Hep3B cells.             We suggest that the PLAG1 binding to the IGF2 P3 promoter and IGF2 expression             is cell type-specific, and that the PLAG1 transcription factor acts as a transcriptional             facilitator that partially overrides the insulation by the H19 ICR.
authors.affiliation: Department of Clinical Neuroscience, Karolinska Institutet, Karolinska University Hospital, SE-171 76 Stockholm, Sweden.
authors.forename: Monira
authors.initials: M
authors.lastname: Akhtar
authors.forename: Claes
authors.initials: C
authors.lastname: Holmgren
authors.forename: Anita
authors.initials: A
authors.lastname: Göndör
authors.forename: Mattias
authors.initials: M
authors.lastname: Vesterlund
authors.forename: Chandrasekhar
authors.initials: C
authors.lastname: Kanduri
authors.forename: Catharina
authors.initials: C
authors.lastname: Larsson
authors.forename: Tomas J
authors.initials: TJ
authors.lastname: Ekström
mesh_terms: D002460:Cell Line; D047369:Chromatin Immunoprecipitation; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D038101:Insulator Elements; D007335:Insulin-Like Growth Factor II / Q000235:genetics*; D009928:Organ Specificity / Q000235:genetics; D011401:Promoter Regions, Genetic; D062085:RNA, Long Noncoding / Q000235:genetics / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; C094335:H19 long non-coding RNA; C104382:PLAG1 protein, human; D062085:RNA, Long Noncoding; D007335:Insulin-Like Growth Factor II