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The Xenopus laevis nuclear receptor BXR has recently been shown to be activated by a class of endogenous benzoate metabolites, indicating the presence of a novel and unsuspected benzoate ligand-dependent signalling pathway. The receptor is expressed ubiquitously in blastula and gastrula stage embryos, and its expression declines during neurula stages. In order to examine further this novel vertebrate signalling system, we have examined the expression of the BXR gene in tailbud stage embryos and adults. We show here that in Xenopus tailbud stage embryos expression is restricted to the hatching gland, suggesting a role in hatching gland function. Neither BXR nor a BXR-VP16 fusion is sufficient to specify hatching gland in neurally-induced tissue. In adults, BXR expression is abundant in the brain and gonads. This expression pattern in adults is distinct from any of the putative mammalian homologues. A nuclear receptor that mediates benzoate signalling has yet to be found in mammals.
Fig. 1. Homology of BXR to other nuclear receptors. Schema of Xenopus BXR showing the DNA-binding zinc finger domain and the ligand binding domain, and homology within these domains to other members of the nuclear receptor superfamily. The closely-related human CARα, PXR.1, SXR and VDR receptors are shown. The human RARα is included as a representative of more distantly-related members of the superfamily. Percentages refer to amino acid identity in the DNA and ligand binding domains.
Fig. 2. Whole-mount in situ hybridisation with a BXR probe. X. laevis embryos were hybridised with a BXR antisense mRNA probe, and the embryos stained according to Harland (1991). (A) Head of a tailbud stage albino embryo (stage 25) showing inverted Y staining of the hatching gland. (B) Schema of the embryohead region showing the positions of the hatching gland and cement gland (adapted from Drysdale and Ellinson 1991).
Fig. 3. BXR expression in neurally-induced ectoderm does not specify hatching gland tissue. (A) Schema of the wild -type BXR and chimeric derivatives. DNA constructs were made which encoded the activator and repressor domains of VP16 and Drosophila Engrailed fused to the carboxy terminus of BXR. Amino acid positions are numbered. (B) Synthetic noggin mRNA was injected into embryos at the two-cell stage, either alone or together with wild-type BXR, BXR-VP16, or BXR-EnR mRNA. Animal cap explants were taken at stage 9, and half were treated with bFGF. Caps were incubated to stage 20 for RT-PCR analysis of Xhe gene expression. ODC was assayed to show that comparable amounts of RNA were isolated for each sample.
A range of tissue RNAs was used as templates for reverse transcriptase. The resultant cDNAs, along with no RNA, no reverse transcriptase and no cDNA negative controls, were used as a template for radioactive PCR, with a primer pair designed against the hinge region of BXR. The resulting fragments were visualised by PAGE and autoradiography. EF1α was assayed to show that comparable amounts of RNA were isolated for each sample.
nr1i2 (nuclear receptor subfamily 1 group I member 2) gene expression in Xenopus laevis embryos, NF stage 25, as assayed by in situ hybridization, lateral view, anteriorleft, dorsal up.
