RESUMO
We have identified a retinoic acid response element (RARE) within a neural enhancer located 3' to the Hoxd4 gene. This RARE is required for the initiation and maintenance of Hoxd4 transgene expression in neurectoderm, and for full anteriorized expression upon retinoic acid (RA) treatment. Mutations within the sequence TTTTCTG, located 2 bp downstream of the RARE, posteriorized transgene activity. However, the onset of transgene expression and its response to RA were indistinguishable from wild type. While the TTTTCTG motif resembles a CDX binding site, human CDX1 protein did not interact with this element in vitro. Three additional regions were also shown to control transgene expression in neurectoderm, establishing that multiple elements constitute the Hoxd4 neural enhancer.
Assuntos
Proteínas Aviárias , Sistema Nervoso Central/metabolismo , Proteínas de Homeodomínio/biossíntese , Proteínas de Homeodomínio/genética , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética , Tretinoína/metabolismo , Animais , Sequência de Bases , Sítios de Ligação , Sistema Nervoso Central/embriologia , Mapeamento Cromossômico , DNA/metabolismo , Ectoderma/metabolismo , Eletroforese em Gel de Poliacrilamida , Embrião de Mamíferos/metabolismo , Elementos Facilitadores Genéticos , Deleção de Genes , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Modelos Genéticos , Dados de Sequência Molecular , Crista Neural/embriologia , Crista Neural/metabolismo , Neurônios/metabolismo , Elementos de Resposta , Homologia de Sequência do Ácido Nucleico , Fatores de Tempo , Transgenes , Tretinoína/farmacologia , beta-Galactosidase/metabolismoRESUMO
One action of retinoids in development is the regulation of Hox gene expression. Hoxd4 and Hoxb4 expression in the embryonic hindbrain is anteriorized by retinoic acid (RA) treatment of mid-gestation mouse embryos. Here we demonstrate that retinoic acid receptor beta (Rarb) deficient mice present only a partial anteriorization of either Hoxd4 or Hoxb4 in response to RA treatment. Our results strongly suggest that RARbeta is a mediator of their RA-response, and reveal anteroposterior polarity within a single rhombomere (r). Additionally, we generated mice doubly mutated for Hoxd4 and Rarb in an attempt to identify common morphogenetic pathways between these two genes. We conclude that there are no synergistic interactions between Hoxd4 and Rarb in the specification of the cervical vertebrae.
