XB-ART-2400Dev Biol February 15, 2005; 278 (2): 526-41.
In zebrafish, the divergent F-type SOX casanova acts downstream of Nodal signaling to specify endoderm. While no casanova orthologs have been identified in tetrapods, the F-type SOX, SOX7, is supplied maternally in Xenopus (Fawcett and Klymkowsky, 2004. GER 4, 29). Subsequent RT-PCR and section-based in situ hybridization analyses indicate that SOX7 mRNA is localized to the vegetal region of the blastula-stage embryo. Overexpression and maternal depletion studies reveal that the T-box transcription factor VegT, which initiates mesoendodermal differentiation, directly regulates SOX7 expression. SOX7, but not SOX17 (another F-type SOX), binds to sites within the Xnr5 promoter and SOX7, but not SOX17, induces expression of the Nodal-related genes Xnr1, Xnr2, Xnr4, Xnr5, and Xnr6, the homeodomain transcription factor Mixer, and the endodermal marker SOX17beta; both SOX7 and SOX17 induce expression of the pan-endodermal marker endodermin. SOX7''s induction of Xnr expression in animal caps is independent of Mixer and Nodal signaling. In animal caps, VegT''s ability to induce Mixer and Edd appears to depend upon SOX7 activity. Whole embryo experiments suggests that vegetal factors partially compensate for the absence of SOX7. Based on the antagonistic effects of SOX7 and SOX3 (Zhang et al., 2004. Dev. Biol. 273, 23) and their common binding sites in the Xnr5 promoter, we propose a model in which competitive interactions between these two proteins are involved in refining the domain of endodermal differentiation.
PubMed ID: 15680368
Article link: Dev Biol
Genes referenced: a2m mixer myc nodal nodal1 nodal2 nodal5 nodal5.4 nodal6 odc1 sox11 sox17a sox17b.1 sox17b.2 sox3 sox7 tbx2 tbxt vegt
Morpholinos: sox7 MO1
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|Fig. 2. Localization of SOX7 RNA. In situ hybridization of longitudinally bisected blastula stage embryos reveals the animal hemisphere concentration of SOX3 (A) RNA, while SOX17 RNA is localized to the vegetal region (B). While staining for SOX7 RNA was consistently quite weak, it was clearly restricted to the vegetal hemisphere (C). Embryos at stage 8 were dissected into animal caps (AC) and vegetal masses (VM) (indicated by white line in part A) and then analyzed for SOX7 or SOX11 mRNA by RT-PCR (D) either immediately (st8) or after they had been cultured until control embryos reached stage 16 (st16). Primers for ornithine decarboxylase (ODC) were used as a control for cDNA quality; the absence of reverse transcriptase (-RT) was used to control for the presence of contaminating DNA. At both time points, SOX7 mRNA was restricted to the vegetal mass; under similar conditions, SOX11 mRNA was restricted to the animal hemisphere. (E) A second set of SOX7-specific primers (Table 1 “SOX7 coding”) was used to confirm the vegetal localization of SOX7 RNA. Animal caps and vegetal masses were prepared at stage 8 and analyzed either immediately (“1”), after 2 h (“2”), or when control embryos had reached stage 27 (“3”); these primers amplified a fragment of the expected size (1kb) that was again restricted to the vegetal mass.|
|Fig. 3. SOX7 induces the endodermal marker endodermin. Fertilized eggs were injected with RNA encoding SOX7-GFP (B and C), allowed to develop to stage 11, and were then stained for Edd RNA. Compared to uninjected embryos (A), the extent and intensity of Edd staining was greatly increased in SOX7-GFP RNA-injected embryos. We compared the effects of injecting embryos with RNAs encoding VegT-EnR (D and G), Mixer-EnR (E and H), and SOX7δC-EnR-myc (F and I). All three EnR-repressor chimeras produce a similar decrease in the level and extent of Edd expression (D–F). In contrast, while VegT-EnR suppressed Xbra expression (G), both Mixer-EnR (H) and SOX7δC-EnR-myc (I) appeared to increase the level of Xbra expression; this increase was confirmed by QPCR (data not shown). When SOX7-GFP and VegT-EnR RNAs were injected together, the level and extent of Edd expression (K and L) was increased compared to the level observed in control (A) or VegT-EnR RNA injected embryos (D and J).|