Scholnick J et al. (1997), Differential expression of Xenopus ribosomal pr...

XB-ART-15686

Biochim Biophys Acta 1997 Oct 09;13541:72-82.

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Differential expression of Xenopus ribosomal protein gene XlrpS1c.

Scholnick J , Sinor C , Oakes J , Outten W , Saha M .

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Components of the translational machinery of the cell, including ribosomal proteins, are generally considered to be clear examples of housekeeping genes with a spatially ubiquitous distribution of messenger RNA during embryonic development. Here we present data based upon in situ hybridization experiments as well as RNase protection assays, demonstrating that Xenopus ribosomal protein gene S1 is differentially expressed in a complex and spatially distinct pattern during embryogenesis. We observed dramatically high levels of expression in some tissues, such as the branchial arches, otic vesicles, optic vesicles and somites and virtually no expression in other tissues, such as the cement gland, epidermis and notochord. Moreover, ribosomal protein genes S22, L1, and L5 display expression patterns nearly identical to S1. Our data is consistent with a model of ribosomal gene expression according to which ribosomal protein genes (or perhaps a subset of ribosomal protein genes) may be expressed at low levels in all tissues, but are abundantly expressed in other cell types reflecting a dynamic and complex pattern of transcriptional control throughout embryonic development.

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Species referenced: Xenopus
Genes referenced: eef1a2 krt12.4 rpl4 rpl5 rps15a rps3 xa-1

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	Fig. 1. Nucleic acid sequence of XlrpS1c and the deduced amino acid sequence. The nucleotide sequence of the XlrpS1c cDNA clone is presented along with a conceptual translation and comparisons to the XlrpS1a and XlrpS1b genes. Letters above the XlrpS1c sequence denote base changes; those in brackets indicate changes which result in the same amino acid or conservative changes.
	Fig. 2. Whole-mount in situ hybridization. Whole mount in situ hybridization with XlrpS1c was performed on Xenopus embryos at different developmental stages. (A) Neurula (stage 20), lateral view; (B) Early tailbud (stage 23) dorsal view; (C) Late tailbud (stage 27); (D, E) Hatching (stage 32), lateral view; (F) dorsal view of tailbud and hatching embryos; (G) Swimming tadpole (stage 35); (H) Swimming tadpole (stage 35 with double color in situ for XlrpS1c (red) and xJ1 (blue). (I, J) Whole mount in situ hybridization was performed on hatching stage (stage 32) embryos with ribosomal protein cDNA S22 (I), L1 (J, upper embryo) and L5 (J, lower embryo). (K) Negative control with sense XlrpS1c probe; early tailbud (stage 23) and late tailbud (stage 27), lateral views. so, somite; ov, optic vesicle; ba, branchial arches; cg, cement gland; ot, otic vesicle; he, heart; nm, nephrogenic mesoderm; hg, hatching gland. Arrowheads point to cement gland. Magnifications are as follows: A, B (30×); C, D, I, J (15×); E, F (10×); G, H (20×).
	Fig. 3. Histological analysis of XlrpS1c expression. Following completion of the in situ hybridization procedure with an XlrpS1c probe (A–E), a cytokeratin probe (F) or an EF-1a probe (G–I), embryos were sectioned transversely at the level of the anterior neural tube/eye region (A, C, G, H), otic vesicles (B), mid-spinal cord (D, F), or tailbud (E, I) and the resulting sections photographed with bright-field optics. Stages of embryos were as follows: A, B, G, H, late tailbud (stage 25–27); C, D, E, F, I, hatching (stages 32–34). J depicts an anterior (optic level) section through a hatching stage (stage 32) embryo which was hybridized with a the sense XlrpS1c probe as a negative control. Nt, neural tube; ov, optic vesicle, ep, epidermis, ot, otic vesicle, ba, branchial arches; le, lens; ey, eye; cg, cement gland; no, notochord, so, somite. Magnifications are as follows: A, B, C, H (50×); D, G, I (60×); E (70×); F (40×).
	Fig. 4. RNase protection analysis of XlrpS1c expression. (A) XlrpS1c is present at low levels in eggs, increases throughout development until late swimming tadpole stages when levels decline. Nieuwkoop and Faber stages [19]are as follows: Blastula, stage 8; Gastrula, stage 10; Neurula, stage 18; Tailbud, stage 25; Hatching, stage 30, Swimming tadpole, stage 35; Limb bud, stage 55. (B) RNase protection was also used to determine if low levels of XlrpS1c were expressed in a given region. Embryos at tailbud stages were dissected into three regions: head; dorsal; and ventral (see text for details of dissections). XlrpS1c levels were highest in the head region and lowest in ventral pieces. At hatching, stages pieces from the region of the neural retina (which showed intense XlrpS1c signal in situ hybridization experiments) and the mid-ventral region (which appeared negative as assayed by in situ hybridization) were subjected to RNase protection analysis. While XlrpS1c levels were clearly higher in the lane with pieces from the retinal region, XlrpS1c signal is also present in the lane containing the ventral pieces, thus suggesting that XlrpS1c may be present ubiquitously at low to moderate levels. Arrows indicate the protected fragment for XlrpS1c. Actin and EF-1a were used as internal controls for the presence and amount of RNA in each sample.
	rps3 (ribosomal protein S3) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 20, lateral view, anterior right, dorsal up.
	rps3 (ribosomal protein S3) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 27, lateral view, anterior right, dorsal up.
	rps3 (ribosomal protein S3) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 32, lateral view, anterior right, dorsal up.
	rps15a (ribosomal protein S15a) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 32, lateral view, anterior right, dorsal up.
	rpl4 (ribosomal protein L4) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 32, lateral view, anterior right, dorsal up.
	eef1a2 (eukaryotic translation elongation factor 1 alpha 2) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 27, transverse section, dorsal up.
	eef1a2 (eukaryotic translation elongation factor 1 alpha 2) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 33-34, transverse section, dorsal up.