title: Identification of target genes for the Xenopus Hes-related protein XHR1, a prepattern factor specifying the midbrain-hindbrain boundary.
abstract: The midbrain-hindbrain boundary (MHB) acts as a local organizer in the development of the CNS in vertebrates. Previously, we identified an MHB-specific bHLH-WRPW transcriptional repressor gene, Xenopus Hes-related 1 (XHR1), which is initially expressed in the presumptive MHB (pre-MHB) region at the early gastrula stage. To better understand the gene cascades involved in MHB formation, we investigated the genes downstream from XHR1 by differential screening using a Xenopus cDNA macroarray and a dexamethasone (DEX)-inducible, dominant-negative transcriptional activator construct of XHR1 (XHR1-VP16-GR). Among the newly identified candidate target genes of XHR1 were Enhancer of split-related genes (ESR1, ESR3/7, and ESR9) and Xenopus laevis cleavage 2 (XLCL2). XHR1-VP16-GR induced the expression of the ESR genes and XLCL2 as well as Xdelta1, Xngnr1, and XHR1 itself in the presence of DEX even after pretreatment with the protein synthesis inhibitor, cycloheximide. This suggests that these genes are direct targets of XHR1. XHR1-knockdown experiments with antisense morpholino oligos and ectopic expression of wild-type XHR1 revealed that XHR1 is necessary and sufficient to repress ESR genes in the pre-MHB region. Misexpression of the ESR genes in the pre-MHB region repressed the MHB marker gene, Pax2, suggesting that the repression of the ESR genes by XHR1 is at least partly required for the early development of the pre-MHB. Our data also show that XHR1 is not activated by Notch signaling, differing from ESR genes. Taken together, we propose a model in which XHR1 defines the pre-MHB region as a prepattern gene by repressing those possible direct target genes.
authors.affiliation: Department of Biological Sciences, Graduate School of Science, University of Tokyo, Japan.
authors.forename: Hitomi
authors.initials: H
authors.lastname: Takada
authors.forename: Daisuke
authors.initials: D
authors.lastname: Hattori
authors.forename: Atsushi
authors.initials: A
authors.lastname: Kitayama
authors.forename: Naoto
authors.initials: N
authors.lastname: Ueno
authors.forename: Masanori
authors.initials: M
authors.lastname: Taira
mesh_terms: D000818:Animals; D001483:Base Sequence; D019521:Body Patterning; D017931:DNA Primers; D004625:Embryo, Nonmammalian / Q000502:physiology*; D005775:Gastrula / Q000502:physiology; D018507:Gene Expression Regulation, Developmental; D008636:Mesencephalon / Q000033:anatomy & histology / Q000196:embryology*; D009024:Morphogenesis; D009419:Nerve Tissue Proteins / Q000502:physiology*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D015183:Restriction Mapping; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012249:Rhombencephalon / Q000033:anatomy & histology / Q000196:embryology*; D014981:Xenopus / Q000196:embryology*; D029867:Xenopus Proteins / Q000235:genetics / Q000502:physiology*
chemical_list: D017931:DNA Primers; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; D029867:Xenopus Proteins; C449809:hes7.1 protein, Xenopus