RESUMO
The administration of the teratogen retinoic acid (and other retinoids) to vertebrate embryos causes a range of developmental abnormalities. It remains to be shown how these teratogenic effects are mediated, and whether or not they reflect any morphogenetic roles retinoids may have in normal development. The most intensively studied cellular action of retinoids has been the activation of retinoid receptors (RARs and RXRs), which are members of the steroid/thyroid family of ligand-modulated transcription factors. Here we report experiments designed to investigate whether the teratogenic effects of retinoic acid on early Xenopus embryos are mediated transcriptionally by receptors, and if these receptors are necessary for normal early Xenopus development. We have demonstrated transcriptional activation of injected reporter genes by exogeneously supplied retinoic acid in Xenopus embryos, presumably as a consequence of the activation of endogenous retinoid receptors. This assay system has been used to demonstrate functional expression, from injected mRNA of (1) a wild-type RARγ (2) a domain-swapped receptor in which the retinoic acid binding domain has been replaced by a thyroid hormone domain to create a thyroid hormone responsive receptor, and (3) a dominant negative from of the RARγ. The wild-type RARγ increases the severity of retinoic acid-mediated defects. In the presence of thyroid hormone the domain-swapped receptor causes abnormalities of gastrulation. The dominant negative decreases the severity of retinoic acid-mediated defects. We conclude that the teratogenic effects of exogenous retinoic acid on Xenopus embryos are mediated, at least in part, transcriptionally via retinoid receptors. It is notable that the dominant negative has no effect on normal development in the absence of exogenous retinoic acid. This is despite observations that this receptor completely blocks transcriptional activation of reporter genes by exogenous retinoic acid up to the beginning of gastrulation, and substantially relieves the teratogenic effects of retinoic acid.
RESUMO
We have investigated the effects of aXenopus alpha thyroid hormone receptor (TR) upon early development ofX. laevis. After deleting the 5'-untranslated region of a cloned TR cDNA, we synthesised TR transcripts that can be translated in oocytes and embryos. When embryos are supplied with this RNA by direct injection, functional TR can be detected through gastrula and neurula stage embryos, considerably in advance of the normal onset of TR expression in larval development. TR mRNA-injected embryos are precociously responsive to thyroid hormone (T3). In the absence of T3 such embryos develop completely normally, but addition of T3 to the medium bathing the embryos results in abnormal embryos with graded anterior (head) deficiencies, losing forehead, eyes and cement gland, progressively. The degree of abnormality is dependent upon the dose of T3 and the stage of development at which it is applied, embryos treated at stage 6 (32 cell) becoming more abnormal than those treated at stage 121/2 (late gastrula). Embryos injected with TR mRNA and treated with T3 are similar, but not identical, to embryos treated with retinoic acid in early development. As is the case with retinoic acid treatment, we show that the T3-dependent effects are due, at least in part, to effects on gastrulation movements.
