Distinct regulators of Shh transcription in the floor plate and notochord indicate separate origins for these tissues in the mouse node.
The establishment of the floor plate at the ventral midline of the CNS is dependent on an inductive signaling process mediated by the secreted protein Sonic hedgehog (Shh). To understand molecularly how floor plate induction proceeds we identified a Shh-responsive regulatory element that directs transgene reporter expression to the ventral midline of the CNS and notochord in a Shh-like manner and characterized critical cis-acting sequences regulating this element. Cross-species comparisons narrowed the activity of the Shh floor plate enhancer to an 88-bp sequence within intron 2 of Shh that included highly conserved binding sites matching the consensus for homeodomain, Tbx and Foxa transcription factors. Mutational analysis revealed that the homeodomain and Foxa binding sites are each required for activation of the Shh floor plate enhancer, whereas the Tbx site was required for repression in regions of the CNS where Shh is not normally expressed. We further show that Shh enhancer activity was detected in the mouse node from where the floor plate and notochord precursors derive. Shh reporter expression was restricted to the ventral (mesodermal) layer of the node in a pattern similar to endogenous Shh. X-gal-positive cells emerging from the node were only detected in the notochord lineage, suggesting that the floor plate and notochord arise from distinct precursors in the mouse node.
Jeong
Yongsu
Y
Department of Genetics, University of Pennsylvania School of Medicine, Clinical Research Building, Room 470, 415 Curie Blvd, Philadelphia, PA 19104, USA.
Epstein
Douglas J
DJ
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D019070:Cell Lineage; D002490:Central Nervous System / Q000033:anatomy & histology* / Q000196:embryology* / Q000502:physiology; D004268:DNA-Binding Proteins / Q000378:metabolism; D004627:Embryonic Induction; D004742:Enhancer Elements, Genetic; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D053823:Hedgehog Proteins; D051542:Hepatocyte Nuclear Factor 3-beta; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D017403:In Situ Hybridization; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D011485:Protein Binding; D016415:Sequence Alignment; D015398:Signal Transduction / Q000502:physiology; D015534:Trans-Activators / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism
D004268:DNA-Binding Proteins; C496502:FOXA2 protein, human; C496503:Foxa2 protein, mouse; D053823:Hedgehog Proteins; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; C465570:Shh protein, mouse; D015534:Trans-Activators; D014157:Transcription Factors; D051542:Hepatocyte Nuclear Factor 3-beta