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Characterization of Xenopus Phox2a and Phox2b defines expression domains within the embryonic nervous system and early heart field.
Talikka M
,
Stefani G
,
Brivanlou AH
,
Zimmerman K
.
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The closely related homeodomain containing genes, Phox2a and Phox2b, are essential for neuronal specification and differentiation within discrete subsets of neurons during vertebrate embryogenesis. We have isolated Xenopus Phox2 homologs, termed Xphox2a and Xphox2b, and characterized their expression during early development. In addition, we have characterized a Phox2a splice variant, termed Xphox2a.2, which lacks homeo- and C-terminal protein coding domains. Xphox2a, Xphox2a.2 and Xphox2b transcripts are expressed in dynamic temporal and regional patterns during nervous system development. The expression of Xphox2a and Xphox2b is only partially overlapping and includes cranial motor and interneuron populations as well as peripheral sympathetic and cranial ganglion neurons, sites linked to Phox2 expression in other species. In addition, we have identified an early domain of Xphox2a and subsequent Xphox2b expression in ventral regions of the embryo, within the developing heart field. XPhox2 expression within this domain is preceded by the gastrula-stage expression of the proneural basic helix-loop-helix transcription factor, Xash1, pointing to a new region of action for this group transcription factors during vertebrate development.
Fig. 2. Characterization of Xphox2a, Xphox2a.2 and Xphox2b expression in the Xenopus embryo. Whole-mount in situ hybridization was used to characterize Xphox2 gene expression during Xenopus development. Xphox2a (A–G) and Xphox2a.2(H) expression in the developing embryo. Similar results were obtained with both full-length and 3′ specific Xphox2a and Xphox2a.2 probes as outlined in the text and Experimental Procedures. Panels (A–G) show staining with full-length Xphox2a probe. Arrow marks ventralmidbrain domain of Xphox2a expression at stage 27 (B) and stage 35 (E). This ventral domain is further documented in transverse sections at these stages (C,F). Asterisks mark zones of Xphox2a and Xphox2a.2 expression in the hindbrain (B,H). Transverse sections through posteriorhindbrain domains of Xphox2a stained embryos are shown at stage 27 (D) and stage 35 (G). Note that the stage 27 embryo was imbedded asymmetrically and staining in (D) is therefore asymmetric due to the combination of thick sections (30 μm) and discontinuous zones of Xphox2a expression within this region. (I–O) Xphox2b expression in the developing embryo. Asterisk marks anteriorhindbrain domain of Xphox2b expression (J) also shown in transverse section at stage 26 (K) and stage 35 (N). Arrow marks posteriorhindbrainXphox2b staining (J) also shown at stage 26 (L) and stage 35 (O). In (P) a cross-section through the hindbrain of an embryo double hybridized with Xphox2a and Xphox2b probes is shown. The Xphox2a domain is blue and the Xphox2b domain is magenta/purple.
Fig. 3. Ventral and lateral domains of Xphox2a and Xash1 expression. Patterns of gene expression were characterized as indicated by whole-mount in situ hybridization at neural plate/fold (A–E, G–L), neurula (F) and early tailbud stages (M–O). The ventral side of the embryo is shown in (A,E–G,I). In (B,H,J), both dorsal and ventral domains of XPhox2a, N-tubulin and Xash1 expression are visible in a neural plate/fold stage embryo. In (F), Xnkx2.5 staining is blue and Xphox2a positive cells are magenta. A transverse section through the neural plate of embryos probed for Xphox2a(C,D) or Xash1(K,L) is shown at low (C,K) and high (D,L) magnifications.
ascl1 (achaete-scute complex homolog 1) gene expression in Xenopus laevis embryos, NF stage 14, as assayed by in situ hybridization. Anterior view: dorsal up.
