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Intestinal remodeling during amphibian metamorphosis resembles the maturation of the adult intestine during mammalian postembryonic development when the adult epithelial self-renewing system is established under the influence of high concentrations of plasma thyroid hormone (T3). This process involves de novo formation and subsequent proliferation and differentiation of the adult stem cells. The T3-dependence of the formation of adult intestinal stem cell during Xenopus laevis metamorphosis offers a unique opportunity to identify genes likely important for adult organ-specific stem cell development. We have cloned and characterized the ectopic viral integration site 1 (EVI) and its variant myelodysplastic syndrome 1 (MDS)/EVI generated via transcription from the upstream MDS promoter and alternative splicing. EVI and MDS/EVI have been implicated in a number of cancers including breast, leukemia, ovarian, and intestinal cancers. We show that EVI and MDS/EVI transcripts are upregulated by T3 in the epithelium but not the rest of the intestine in Xenopus laevis when adult stem cells are forming in the epithelium. Our results suggest that EVI and MDS/EVI are likely involved in the development and/or proliferation of newly forming adult intestinal epithelial cells.
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Figure 2. Tissue specific upregulation of EVI, MDS, and MDS/EVI transcripts in the intestine during metamorphosis.The expression of the transcripts was analyzed by using transcript-specific primer sets with qPCR on total RNA from the isolated intestinal epithelium (Ep) or the rest of the intestinal tissues (Non-Ep) of tadpoles at prometamorphosis (stage 56), the climax of metamorphosis (stage 61), and the end of metamorphosis (stage 66). The relative expression was obtained after normalizing against the control gene EF1α and was presented with the level at Ep 56 set to 1, allowing for easy comparison within each transcript. However, these data do not allow for comparisons of the expression levels between different transcript variants. Error bars represent the SEM (nâ=â3). * indicates transcript levels lower than the peak expression of the specific transcript in the indicated tissue (pâ¤0.05).
Figure 3. EVI, MDS, and MDS/EVI transcripts are induced in the intestine of premetamorphic tadpoles treated with T3.The expression of the transcripts was analyzed by using transcript-specific primer sets (as in Fig. 2.) on total intestinal RNA from tadpoles at stage 54 exposed to 5 nM T3 for 0â7 days. Error bars indicate SEM (nâ=â3). * indicates transcript levels lower than the peak expression of the specific transcript (pâ¤0.05).
Figure 4. MDS/EVI is strongly upregulated in the intestine at the climax of metamorphosis.A primer set common to EVI and MDS/EVI was used for qPCR analysis on total intestinal RNA at different stages during metamorphosis as in Fig. 2. Note that the data is consistent with that in Fig. 2 if one considers the fact that there is little non-Ep in premetamorphic tadpoles at stage 56 but non-Ep increases as a percentage of the total intestine during metamorphosis. Error bars indicate SEM (nâ=â3). * indicates transcript levels lower than the peak expression (pâ¤0.05).
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