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Sox17 and ß-catenin cooperate to regulate the transcription of endodermal genes
Development Sinner et al. 131: 3069

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Figure S1 - Fig. S1. Sox and Tcf target specificity. RNA encoding the indicated Sox or Tcf (200 pg) was injected into the animal region of two-cell embryos. Animal caps were isolated at stage 9 and assayed at stage 11-12 by real-time RT-PCR. The results are normalized and shown as a ratio to ODC. This experiments was repeated three times and a representative is shown. With the exception that Sox3 and Lef induced Gata6 and Gsc, none of the other HMG box factors reproducibly induced significant expression of Sox17 targets.
Figure S2 - Fig. S2. Tcf3 does not regulate Sox17 target genes in vivo. Embryos were injected at the two-cell stage with antisense morpholino oligos to b-catenin (20 ng) or Tcf3 (20 ng), as previously described (Heasman et al., 2000; Houston et al., 2002). At gastrula stage, five embryos from each condition were pooled and assayed by real-time RT-PCR and relative gene expression was normalized and presented as a ratio to ODC. As a control, the expression of the ubiquitously expressed gene plakoglobin was unchanged. As expected, Hex was downregulated by b-catenin depletion, but upregulated by Tcf3 depletion. This is the predicted result observed from all Tcf3-regulated genes (Houston et al., 2002) and is consistent with de-repressing Tcf3 activity. By contrast, the Sox17 target genes Hnf1b, Edd and Foxa2 were not upregulated in Tcf3-depleted embryos but were repressed in b-catenin depleted embryos. This indicates that they are not Tcf3 targets but are b-catenin targets. Consistent with the results presented in the paper, Foxa1 was not significantly regulated by b-catenin levels.
Figure S3 - Fig. S3. Direct Sox17b targets. This experiment is a repeat of that presented in Fig. 2B, but analyzed by real time RT-PCR to better show the quantitation of the induced transcription. Isolated blastula animal cap explants either injected with mRNA encoding GR:Sox17b (150 pg) or uninjected were each cultured in three conditions; 1´MBS, 1´MBS + 10-6 dexamethasone (DEX) or in 1´MBS with 10-6 dexamethasone + 10 mg/ml cycloheximide (CHX). Dexamethasone activates the GR:Sox17b fusion protein and cycloheximide blocks translation. Control animal caps were treated with 5 ng/ml human activin A either with or without 10 mg/ml cycloheximide. At gastrula (stage 11), the explants were assayed by RT-PCR. Houston, D. W., Kofron, M., Resnik, E., Langland, R., Destree, O., Wylie, C. and Heasman, J. (2002). Repression of organizer genes in dorsal and ventral Xenopus cells mediated by maternal XTcf3. Development 129, 4015-4025.

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