Figure 1.
Structure and conservation of EVI and MDS in humans, X. laevis, and X. tropicalis.
There are 16 exons for human and X. tropicalis EVI and 3 exon for human MDS. In X. laevis, as in humans, exon 2 of MDS can be spliced to exon 2 of EVI to form a larger, fusion transcript MDS/EVI. The % identity of the PR (PRDI-BF1 and RIZ homology domain containing) domain, zinc finger domains (ZF-D) 1 and 2, C-terminal binding protein (CtBP) domain, and acidic domain (AD) are indicated. For an alignment of the EVI amino acid sequence, see [21]. Note that despite an exhaustive search by using RACE and in silico methods, no MDS exon 3 could be found in X. laevis or X. tropicalis, although a 3′ end of MDS exon 2 that is not present in MDS/EVI fusion transcript was cloned by using RACE (not shown). Thus, the existence of the MDS transcript and encoding protein remains unknown as indicated with a question mark in the figure.
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 (n = 3). * indicates transcript levels lower than the peak expression of the specific transcript in the indicated tissue (p≤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 0–7 days. Error bars indicate SEM (n = 3). * indicates transcript levels lower than the peak expression of the specific transcript (p≤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 (n = 3). * indicates transcript levels lower than the peak expression (p≤0.05).