XB-ART-6711Dev Growth Differ August 1, 2002; 44 (4): 257-71.
Conserved expression control and shared activity between cognate T-box genes Tbx2 and Tbx3 in connection with Sonic hedgehog signaling during Xenopus eye development.
Tbx2 and Tbx3 are considered to be cognate genes within a Tbx2/3/4/5 subfamily of T-box genes and are expressed in closely overlapping areas in a variety of tissues, including the eye. Herein, we show that misexpression of Tbx2 and Tbx3 in Xenopus embryos gave rise to defective eye morphogenesis, which was reminiscent of the defect caused by attenuated Sonic hedgehog (Shh) signaling. Indeed, Tbx2/3 misexpression suppressed Gli1, Gli2, Ptc2 and Pax2, mediators or targets of Hedgehog (Hh) signals. From these data, Tbx2/3 may have a shared function in inhibiting Gli-dependent Shh signaling during eye development. Conversely, the expression of Tbx2/3 was severely affected by both Shh and a putative dominant negative form of Hh, as well as by both transactivator and transrepressor forms of Gli-fusion proteins, suggesting that the expression of Tbx2/3 may be regulated by a Gli-dependent Hh signal transduction pathway. Because the Shh signal has been considered to play crucial roles in the formation of the proximal-distal and dorsal-ventral axes in the eyes, these findings about the mutual regulatory mechanism between Tbx2/3 and Gli-dependent Hh signaling provide valuable insight into the cause of the localized expression of Tbx2/3 and their role during the formation of these axes. In addition, our findings also imply the conserved regulation and shared activity between the cognate genes of Tbx2 and Tbx3.
PubMed ID: 12175361
Article link: Dev Growth Differ
Genes referenced: bmp4 gli1 gli2 pax2 pax6 ptch2 shh tbx2 tbx3 tbx5
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|Fig. 1. Expression patterns of Tbx2/3 and the genes involved in Xenopus eye morphogenesis. Whole-mount in situ analyses were performed following single staining for Gli2 (Q–T; the digoxigenin-labeled probe), double staining for Pax6 (A–L,O,P; the fluoresceinlabeled probe; light blue) and the digoxigenin-labeled Tbx2 (A–D), Tbx3 (E–H), Tbx5 (I,J), Pax2 (K,L) or Gli1 (O,P), and triple staining for the fluorescein-labeled probe of Pax6 and the digoxigenin-labeled probes of Sonic hedgehog (Shh) and bone morphogenetic protein- 4 (BMP4; M,N). Except for (T), which is a lateral view, the other figures show anterior views. Arrowheads in (B) and (I) indicate the initial faint expression of Tbx2 and Tbx5, respectively, in the eye field. Arrowheads in (F–H) indicate the position of Tbx3 expression in a region of the forebrain. Arrowheads and arrows in (M) indicate the expression of Shh and the initial faint expression of BMP4 in the eye field, respectively. Arrowheads in (O,P) indicate the expression of Gli1 in the ventral outline of the eye field.|
|Fig. 2. Morphologic defects caused by misexpression of Tbx2 and Tbx3. (A–F) Tbx2 or (G–L) Tbx3 mRNA was injected unilaterally into the animal pole of two-cell stage embryos. LacZ mRNA was coinjected as a tracer (A–D,G–J). With 100 pg Tbx2/3 mRNA injected, embryos at stage 45 typically show defective eye morphogenesis (B–E,H–K). (A,B) Lateral views of uninjected and injected sides, respectively. (C) Dorsal view of the same Tbx2-injected embryo shown in (A,B). (G–I) A Tbx3- injected embryo. (D,J) Dorsal views of the fused eyes caused by ectopic expression of Tbx2 and Tbx3, respectively. (E,F) Transverse sections through the eyes and otic vesicles of the Tbx2-injected embryo; (K,L) transverse sections through the eyes and otic vesicles of the Tbx3-injected embryo.|
|Fig. 3. Alterations in the expression of Pax2 and Pax6 in embryos misexpressing Tbx2 or Tbx3. (A–C) 100 pg Tbx2 or (D–F) Tbx3 mRNA was injected unilaterally into the animal pole of two-cell stage embryos. LacZ mRNA was coinjected as a tracer. Pax2 and Pax6 were stained purple and light blue, respectively. (A,D) Anterior views of early tailbud embryos. (B,C,E,F) Lateral views of uninjected (B,E) and injected (C,F) sides of the same tadpole embryo. Green arrowheads in (A,D) indicate the midline position.|
|Fig. 4. Tbx2/3 suppressed Gli1/2 and Gli-responsive genes in the neural plate. 100 pg Tbx2 or Tbx3 mRNA was injected unilaterally into the animal pole of two-cell stage embryos and cultured until stage 13. LacZ mRNA was coinjected as a tracer. Injected mRNA and probes are indicated at the top and left, respectively. Control indicates uninjected control embryos.|
|Fig. 5. VP16-Tbx2-GR and VP16-Tbx3-GR induced the ectopic tail-like structure and Gli2. 10 pg VP16-Tbx2-GR (A,C,E) or VP16- Tbx3-GR (B,G) mRNA and 100 pg Tbx2 (D) or Tbx3 (F) mRNA was injected unilaterally into the animal pole of two-cell stage embryos. LacZ mRNA was coinjected as a tracer. (A–C) Lateral and (D–G) anterior views. Embryos were treated with dexamethasone (dex) at stage 10.25 (A–C) or stage 12.5 (E,G). (A) Ectopic tail-like structure induced by VP16-Tbx2-GR. (B,C) Ectopic Gli2 expression in the protrusion. While Gli2 was suppressed by Tbx2 (D; n = 23/25) and Tbx3 (F; n = 13/20) in the eye field, it was weakly upregulated by VP16-Tbx2-GR (E; n = 11/19) and VP16-Tbx3-GR (G; n = 4/9) around the eye field (E,G, arrows).|
|Fig. 6. Overexpression of Sonic hedgehog (Shh) and N–C changed the expression of Tbx2 and Tbx3. (A–F) 1 ng Shh or (G–L) 2 ng N–C mRNA was injected unilaterally into the animal pole of two-cell stage embryos. LacZ mRNA was coinjected as a tracer. (B,D,H,J) Lateral views of the uninjected side of the embryos in (A,C,G,I), respectively. (E,F,K,L) Anterior views of the injected embryos. Tbx3 was both suppressed (E,K) and expanded (F,L) by Shh or N–C when embryos were analyzed at the neurula and early tailbud stages (E, stage 18; F, stage 16; K, stage 18; L, stage 23).|
|Fig. 7. Alterations in the expression of Tbx2 and Tbx3 in embryos overexpressing VP16-Gli2zfd-GR, EnR-Gli2zfd-GR, Tbx2 or Tbx3. (A–D) 100 pg VP16-Gli2zfd-GR, (E H) 1 ng EnR-Gli2zfd-GR, (I–K) 100 pg Tbx2 or (L) 100 pg Tbx3 mRNA was injected unilaterally into the animal pole of two-cell stage embryos. LacZ mRNA was coinjected as a tracer. (A–H) Embryos were treated with dexamethasone (dex) at stage 16. (A,C,E,G,J) Lateral views of the uninjected side of the embryos in (B,D,F,H,K), respectively. (L) Anterior view of a Tbx3-injected embryo. Probes were Tbx2 (A,B,E,F,L) and Tbx3 (C,D,G–K). Embryos are at stages 20 (L), 26 (A–I) and 28 (J,K).|