Lau GT et al. (2001), Embryonic XMab21l2 expression is required for g...

XB-ART-9654

Biochem Biophys Res Commun 2001 Feb 09;2805:1378-84. doi: 10.1006/bbrc.2001.4290.

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Embryonic XMab21l2 expression is required for gastrulation and subsequent neural development.

Lau GT , Wong OG , Chan PM , Kok KH , Wong RL , Chin KT , Lin MC , Kung HF , Chow KL .

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Cell fate determining gene mab-21 regulates the proper establishment of neural cell fate and sensory organ identity in nematode. Mammalian homologs of mab-21 have also been implicated to play critical roles in mid-, hindbrain and craniofacial differentiation. We report here the isolation of a mab-21 homolog, XMab21l2, from Xenopus. We showed that its expression in Xenopus was initiated at gastrulation and prominent signal was detected in neurulating embryos at the neural tube, the optic tissue, the developing midbrain, and the pharyngeal pouches. We demonstrated by RNA interference (RNAi), together with other antisense approaches, that XMab21l2 expression is required for the completion of gastrulation and subsequent neural development.

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Species referenced: Xenopus
Genes referenced: mab21l2 pax6 tbxt

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	FIG. 1. Sequence alignment of Mab21 homologs in both invertebrate and vertebrates; worm (Ce), human (H), mouse (M), and Xenopus (X). Identical amino acids are boxed.
	FIG. 2. RT-PCR analysis of XMab21l2 expression in Xenopus laevis embryos from stage 0 to 14.
	FIG. 3. Expression patterns of Xenopus XMab21l2. (A) Viewed from the vegetal pole. B, C, E, and L are dorsal views and D and F–H are lateral views. All whole-mount embryos and sections have the anterior side to the left, except for frontal cross sections in I and J where the dorsal side is on top. For sagittal section in K and coronal section in L, anterior is on the left. (A) Stage 12, (B) stage 14, (C) stage 15, (D) stage 17/18, (E, I) stage 25, (F, J, L) stage 28/29, (G, K) stage 32, (H) stage 37/38. (EY, eye; DI, diencephalon; HB, hindbrain; LN, lens; MB, midbrain; NF, neural fold; NT, neural tube; OV, optic vesicle; PH, pharyngeal pouch; RE, retinal epithelium; SO, somitic derivatives; YP, yolk plug). Scale bars 5 1 mm for A to H and J; scale bars 5 0.4 mm, for I, K, and L.
	FIG. 4. Suppression of XMab21l2 expression induces a neural tube closure failure in Xenopus embryos. Posterior abnormality was also observed from stage 16 to stage 40 (arrows). (A) Uninjected embryo at stage 26. (B and C) Embryos injected with 1 ng XMab21l2 antisense RNA, shown at stage 26 with different neural tube closure abnormality. (D and E) Embryos injected with 1 ng antisense oligo, shown at stage 26 and stage 40, respectively. (F, G) Embryos injected with 0.5 and 1 ng dsRNA, respectively, shown at stage 40. (F, G, H) Embryos under dsRNA interference did not develop normal eye (arrowhead). (I) Uninjected control embryo at stage 40.
	FIG. 5. Antisense XMab21l2 RNA injection experiment results in embryos arrested at late-gastrula/early-neurula stage exhibiting yolk plug and abnormal endoderm (A), which could be rescued by coinjecting sense XMab21l2 mRNA (B).
	FIG. 6. b-Galactosidase activity assayed on embryo extracts demonstrated that XMab21l2 dsRNA mixture at neural defect inducing dosage (0.1 to 1 ng) did not significantly inhibit the translation of coinjected b-galactosidase mRNA.
	FIG. 7. RT-PCR analysis of the expression of notochordal (Xbra) and eye (Pax-6) marker genes in buffer-injected (C) and XMab21l2 dsRNA-injected embryos (R).