Brändli AW and Kirschner MW (1995), Molecular cloning of tyrosine kinases in the ea...

XB-ART-19649

Dev Dyn 1995 Jun 01;2032:119-40. doi: 10.1002/aja.1002030202.

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Molecular cloning of tyrosine kinases in the early Xenopus embryo: identification of Eck-related genes expressed in cranial neural crest cells of the second (hyoid) arch.

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Species referenced: Xenopus laevis
Genes referenced: csk egr2 en2 epha2 epha8 ephb4 fgfr1 fgfr4 g67 jak3 mt-tr odc1 pdgfra ptk7 trna tyk2

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	Fig. 4. A-L: Developmental expression profile of Xenopus TK genes. RNase protection analysis of Xenopus TK genes was performed using 10 p,g of total RNA (2.2 embryo equivalents) from oocytes (VI), unfertilized eggs (E), and embryos of blastula (st. 5, 8, 9), gastrula (st. 10.5, 12), neurula (st. 15, 19), tailbud (st. 21, 24), and hatching stage (st. 34). The expression of ornithine decarboxylase (ODC), and EF-la are included as controls. ["PI Labeled anti-sense probes were synthesized from the partial TK cDNAs. The undigested probe and a background control (hybridization with yeast tRNA) were included in each RNase protection analysis experiment (not shown). Protected fragments were quantitated using a Phosphorlmager (Molecular Dynamics), and the number of TK mRNA molecules per embryo was estimated as described in Experimental Procedures. The y-axis represents the TK mRNA levels displayed as units of 106 mRNA molecules per embryo.
	Fig. 5. Spatial expression of E59 (FGFR-Al), G62, and E30 transcripts in tailbud Xenopus embryos. Whole-mount in situ hybridizations were performed on albino Xenopus embryos using digoxigenin-labeled antisense RNA probes. Hybridization signals were visualized as alkaline phosphatase chromogenic reaction products. A: Lateral view of a stage 29/30 embryo stained for E59 mRNA expression. E59 transcripts are detected mainly in the brain, pronephros, and somites. Note also the high levels of expression in unsegmented somitogenic mesoderm (sm). 9: Close-up view of A at higher magnification showing details of E59 mRNA expression in the anterior half of the embryo. Extensive expression is seen in the fore-, mid-, and hindbrain (fb, mb, and hb). The otic vesicle (ov) is indicated as a reference point for the posterior limit of the E59 expression in the hindbrain. Staining is also associated with visceral arches (arrows), the pronephros (pn), and the pronephric duct (pd). C: Lateral view of a stage 29/30 embryo stained for G62 mRNA. Similar to E59, G62 transcripts are mainly detected in the brain, pronephroi, and somites. No expression was detected in unsegmented somitogenic mesoderm as with E59. D: Close-up view of C at higher magnification. In the brain, expression of G62 transcripts is seen predominantly in the eye vesicle (ev), midbrain (mb), and hindbrain (hb). The midbrain/hindbrain junction (j) is devoid of G62 transcripts. Other areas of expression include the pronephros (pn), the pronephric duct (pd), and the tissues in the region of the liver rudiment (Ir). Faint expression is also seen in the visceral arches (arrows). E: Lateral view of a stage 28 embryo stained for E30 mRNA expression. Extensive expression of E30 transcripts is seen in the brain, visceral arches, and at lower levels in somites. F: Close-up view of E at higher magnification to illustrate staining of the olfactory placode (op), the wall of the otic vesicle (ov), the forebrain (fb), and visceral arches (arrows). All embryos were rendered transparent with Murray's clear prior to hybridization. Anterior ends of embryos are oriented to the left. Scale bars: A = 288 pm; B = 144 pm; C, E = 225 km; D, F = 112.5 pm.
	Fig. 6. Spatial expression patterns of G56 mRNA in neurula and tailbud Xenopus embryos. A: Lateral view of a stage 20 embryo. The expression of G56 mRNA is predominantly associated with the developing brain, the cement gland anlage (cg), and somites that have undergone rotation (arrowheads 1-6). B: Dorsal view of a stage 21 embryo illustrating G56 mRNA expression in somites (arrowheads 1-8), and the unsegmented somitogenic mesoderm (sm). C: Lateral view of a stage 26 embryo. G56 mRNAs are localized to segmented somites (arrowheads), and to different parts of the hindbrain (hb). Expression in the cement gland (cg) has ceased by this stage. D: Dorsal view of a stage 26 embryo at a higher magnification to illustrate expression of G56 mRNA in the differentiating somites. G56 mRNAs are present around somite nuclei, which are aligned vertically in each somite. Thus, the staining appears as a stripe (arrowheads) in each somite block. Scale bars: A,B,D = 144 pm; C = 225 pm.
	Fig. 7. Neural crest expression of G37 in neurula and tailbud Xenopus embryos. A: Lateral view of a stage 15 embryo. Arrowheads indicate staining of cranial neural crest. B: Dorsal view of A. G37 expression is seen in the most anterior cranial neural crest. Segregation into individual neural crest segments (arrowheads) has begun. C: Lateral view of a stage 21 embryo. G37 mRNAs are now found in cranial (m, mandibular crest; h, hyoid crest; ab, anterior branchial; pb, posterior branchial), and truncal (1) neural crest segments. The periphery of the eye vesicle (ev) is populated by mandibular (m) neural crest expressing G37 mRNA. D: Dorsal view of C to illustrate expression of G37 mRNAs in the posterior branchial arch segment (pb), and in truncal (1) neural crest. E: Lateral view of a stage 27 embryo. Expression of G37 mRNAs in all cranial neural crest segments and in truncal (1) neural crest of the dorsal fin. F: Close-up view of E at higher magnification to illustrate expression in cranial and truncal neural crest (see C for labeling). Neural crest derivatives were labeled according to the nomenclature of Sadaghiani and Thiebaud (1987). Scale bars: A, B = 160 pm; C, D = 144 pm; E = 225 pm; F = 112.5 pm.
	Fig. 8. Whole-mount in situ hybridization analysis of G50 expression in neurula and early tailbud stage Xenopus embryos. A: Lateral view of a stage 17 embryo stained for G50 mRNA expression. Transcripts are detected in two bilateral stripes of cells in the head region (arrowheads), and diffusely in the posterior end of the embryo. 6: Dorsal view of A at higher magnification showing in greater detail the localization of G50 mRNAs in premigratory (arrow) and early migratory neural crest cell populations (arrowhead). C: Lateral view of a stage 21 embryo stained for Xenopus En-2 to localize the midbrain-hindbrain junction. D: Lateral view of a stage 21 embryo stained for G50. Expression of G50 mRNAs is seen in a stripe of cells at the height of the presumptive rhombomere 3, and in neural crest of the hyoid arch (h). E: Lateral view of a stage 21 embryo stained for Xenopus Krox-20. Two stripes of cells in the hindbrain corresponding to the presumptive rhombomeres 3 and 5, and neural crest of anterior branchial arch (ab) are expressing XKroxPO mRNAs. F Lateral view of a stage 21 embryo stained simultaneously for G50 and XKroxPO. Labeling of stained tissues is as in D and E, respectively. The intensity of the background stain varies with respect to the length of incubation of the embryos in the chromogenic reaction. Embryos in C and E were developed for 1 hr, while the other embryos were incubated 16 hr to obtain optimal staining. The stars in C and F indicate staining artifacts. Scale bars: A = 160 pm; 0 = 80 pm; C F = 144 pm.
	epha2 (EPH receptor A2) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 21, lateral view, anterior left, dorsal up.
	pdgfra (platelet-derived growth factor receptor, alpha polypeptide) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 27, lateral view, anterior left, dorsal up.
	ptk7 (protein tyrosine kinase 7) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 29 and 30, lateral view, anterior left, dorsal up.
	pdgfra (platelet-derived growth factor receptor, alpha polypeptide) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 21, lateral view, anterior left, dorsal up.