RESUMEN
Retinoic acid receptor beta 2 (RARß2) has been proposed as an important receptor mediating retinoid-induced axonal growth and regeneration in developing mammalian spinal cord and brain. In urodele amphibians, organisms capable of extensive central nervous system (CNS) regeneration as adults, this receptor had not been isolated, nor had its function been characterized. We have cloned a full-length RARß2 cDNA from adult newt CNS. This receptor, NvRARß2, is expressed in various adult organs capable of regeneration, including the spinal cord. Interestingly, both the NvRARß2 mRNA and protein are up-regulated during the first 2 weeks after amputation of the tail, primarily in the ependymoglial and meningeal tissues near the rostral cut surface of the cord. Treatment with LE135, a RARß-selective antagonist, caused a significant inhibition of ependymal outgrowth and a decrease in tail regenerate length. These data support an early role for this receptor in caudal spinal cord and tail regeneration in this amphibian.
Asunto(s)
Proteínas Anfibias/biosíntesis , Regulación de la Expresión Génica/fisiología , Receptores de Ácido Retinoico/biosíntesis , Regeneración/fisiología , Médula Espinal/fisiología , Cola (estructura animal)/fisiología , Proteínas Anfibias/antagonistas & inhibidores , Proteínas Anfibias/genética , Animales , Clonación Molecular , ADN Complementario/genética , ADN Complementario/metabolismo , Dibenzazepinas/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Notophthalmus viridescens , Especificidad de Órganos/efectos de los fármacos , Especificidad de Órganos/fisiología , Ratas , Receptores de Ácido Retinoico/antagonistas & inhibidores , Receptores de Ácido Retinoico/genética , Regeneración/efectos de los fármacos , Médula Espinal/patología , Traumatismos Vertebrales/genética , Traumatismos Vertebrales/metabolismo , Traumatismos Vertebrales/patología , Cola (estructura animal)/lesiones , Cola (estructura animal)/patologíaRESUMEN
The proximodistal identity of a newt limb regeneration blastema is respecified by exposure to retinoic acid, but its molecular basis is unclear. We identified from a differential screen the cDNA for Prod 1, a gene whose expression in normal and regenerating limbs is regulated by proximodistal location and retinoic acid: Prod 1 is the newt ortholog of CD59. Prod 1/CD59 was found to be located at the cell surface with a GPI anchor which is cleaved by PIPLC. A proximal newt limb blastema engulfs a distal blastema after juxtaposition in culture, and engulfment is specifically blocked by PIPLC, and by affinity-purified antibodies to two distinct Prod 1/CD59 peptides. Prod 1 is therefore a cell surface protein implicated in the local cell-cell interactions mediating positional identity.
Asunto(s)
Antígenos CD59/fisiología , Extremidades/fisiología , Regeneración/fisiología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , ADN Complementario/genética , Glicosilfosfatidilinositoles/fisiología , Datos de Secuencia Molecular , Notophthalmus viridescensRESUMEN
Members of the hedgehog family have been shown to play a key role in many developmental processes, including limb patterning and chondrogenesis. We have therefore investigated whether members of this family are also expressed during regeneration of the adult urodele limb and are regulated by retinoic acid (RA), since this derivative induces proximodistal duplications in regenerating limbs, and has been shown to regulate sonic hedgehog (shh) in the developing limbs of birds and mammals. We report here that a newt homologue of Xenopus banded hedgehog, called N-bhh, is uniformly expressed by mesenchymal blastemal cells from the initial stages of regeneration and is up-regulated by RA. In addition, we show that N-bhh is uniformly expressed in the early limb bud of the newt embryo. Since bhh has not been detected in developing limbs of higher vertebrates, its expression in developing and regenerating newt limbs may be related to the regenerative capability of urodeles.