The total dependence of amphibian metamorphosis on thyroid hormone (T 3 ) provides a unique vertebrate model for studying the molecular mechanism of T 3 receptor (TR) function in vivo. In vitro transcription and developmental expression studies have led to a dual function model for TR in amphibian development, i.e., TRs act as transcriptional repressors in premetamorphic tadpoles and as activators during metamorphosis. We examined molecular mechanisms of TR action in T3-induced metamorphosis by using dominant-negative receptors (dnTR) ubiquitously expressed in transgenic Xenopus laevis. We showed that T 3 -induced activation of T 3 target genes and morphological changes are blocked in dnTR transgenic animals. By using chromatin immunoprecipitation, we show that dnTR bound to target promoters, which led to retention of corepressors and continued histone deacetylation in the presence of T 3 . These results thus provide direct in vivo evidence for the first time for a molecular mechanism of altering gene expression by a dnTR. The correlation between dnTR-mediated gene repression and inhibition of metamorphosis also supports a key aspect of the dual function model for TR in development: during T 3 -induced metamorphosis, TR functions as an activator via release of corepressors and promotion of histone acetylation and gene activation.

The matrix metalloproteinases are a family of proteases capable of degrading various components of the extracellular matrix. Expression studies have implicated the involvement of the matrix metalloproteinase stromelysin-3 (ST3) in tissue remodeling and pathogenesis. However, the in vivo role of ST3 has been difficult to study because of a lack of good animal models. Here we used intestinal remodeling during thyroid hormone-dependent metamorphosis of Xenopus laevis as a model to investigate in vivo the role of ST3 during postembryonic organ development in vertebrates. We generated transgenic tadpoles expressing ST3 under control of a heat shock-inducible promoter. We showed for the first time in vivo that wild type ST3 but not a catalytically inactive mutant was sufficient to induce larval epithelial cell death and fibroblast activation, events that normally occur only in the presence of thyroid hormone. We further demonstrated that these changes in cell fate are associated with altered gene expression in the intestine and remodeling of the intestinal basal lamina. These results thus suggest that ST3 regulates cell fate and tissue morphogenesis through direct or indirect ECM remodeling.

Advances in vertebrate genetics have allowed studies of gene function in developing animals through gene knockout and transgenic analyses. These advances have encouraged the development of gene-based therapies through introduction of exogenous genes to enhance and/or replace dysfunctional or missing genes. However, in vertebrates, such analyses often involve tedious screening for transgenic animals, such as PCR-based genotype determinations. Here, we report the use of double-promoter plasmids carrying the transgene of interest and the crystallin-promotor-driven Green fluorescent protein (GFP) in transgenic Xenopus laevis tadpoles. This strategy allows a simple examination for the presence of GFP in the eyes to identify transgenic animals. PCR-based genotyping and functional characterization confirms that all animals expressing GFP in the eyes indeed carry the desired promoter/transgene units. Thus, the use of this and other similar vectors should dramatically improve current transgenesis protocols and reduce the time and cost for identifying transgenic animals.