title: JUMONJI, a critical factor for cardiac development, functions as a transcriptional repressor.
abstract: JUMONJI (JMJ) is a nuclear factor that is critical for normal cardiovascular development, evidenced by the analysis of jmj homozygous mutant mice. However, the molecular function of JMJ remains to be elucidated. In the present study, we investigated whether JMJ is a transcriptional modulator. Reporter gene assays using the GAL4-DNA binding domain fused to JMJ and a reporter gene consisting of the GAL4 binding sites upstream of a luciferase reporter gene indicated that JMJ functions as a powerful transcriptional repressor. The DNA binding motif of JMJ was determined using CASTing experiments by incubating a random oligonucleotide library with the GST-JMJ fusion protein coupled to agarose beads. Among the selected binding oligonucleotides, the high affinity DNA binding sequences were identified by gel retardation assays. JMJ repressed expression of the reporter genes containing the high affinity JMJ binding sequences, indicating that JMJ is a DNA-binding transcriptional repressor. The domains for transcriptional repression, DNA binding, and nuclear localization signal were mapped by mutational analyses using reporter gene assays, gel retardation assays, and immunostaining experiments, respectively. The present data demonstrate for the first time that JMJ functions as a DNA-binding transcriptional repressor. Therefore, JMJ may play a critical role in transcription factor cascade to regulate expression of heart-specific genes and normal cardiac development.
authors.affiliation: Department of Anatomy and Cardiovascular Research Center, University of Wisconsin Medical School, 1300 University Avenue, Madison, WI 53706, USA.
authors.forename: Tae-Gyun
authors.initials: TG
authors.lastname: Kim
authors.forename: Jonathan C
authors.initials: JC
authors.lastname: Kraus
authors.forename: Junqin
authors.initials: J
authors.lastname: Chen
authors.forename: Youngsook
authors.initials: Y
authors.lastname: Lee
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D006321:Heart / Q000254:growth & development; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000494:pharmacology / Q000502:physiology*; D019913:Nuclear Localization Signals; D009841:Oligonucleotides / Q000378:metabolism; D063151:Polycomb Repressive Complex 2; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000494:pharmacology; D012097:Repressor Proteins / Q000494:pharmacology*; D014162:Transfection
chemical_list: C093650:Jarid2 protein, mouse; D009419:Nerve Tissue Proteins; D019913:Nuclear Localization Signals; D009841:Oligonucleotides; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D063151:Polycomb Repressive Complex 2