XB-ART-3774J Comp Neurol April 26, 2004; 472 (2): 246-56.
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Expression of vomeronasal receptor genes in Xenopus laevis.
In the course of evolution, the vomeronasal organ (VNO) first appeared in amphibians. To understand the relationship between the VNO and the vomeronasal receptors, we isolated and analyzed the expression of the vomeronasal receptor genes of Xenopus laevis. We identified genes of the Xenopus V2R receptor family, which are predominantly expressed throughout the sensory epithelium of the VNO. The G-protein Go, which is coexpressed with V2Rs in the rodent VNO, was also extensively expressed throughout the vomeronasal sensory epithelium. These results strongly suggest that the V2Rs and Go are coexpressed in the vomeronasal receptor cells. The predominant expression of the Xenopus V2R families and the coexpression of the V2Rs and Go imply that V2Rs play important roles in the sensory transduction of Xenopus VNO. We found that these receptors were expressed not only in the VNO, but also in the posterolateral epithelial area of the principal cavity (PLPC). Electron microscopic study revealed that the epithelium of the PLPC is more like that of the VNO than that of the principal and the middle cavity. These results suggest that in adult Xenopus the V2Rs analyzed so far are predominantly expressed in the vomeronasal and vomeronasal-like epithelium. The analysis of V2R expression in Xenopus larvae demonstrates that V2Rs are predominantly expressed in the VNO even before metamorphosis.
PubMed ID: 15048691
Article link: J Comp Neurol
Species referenced: Xenopus laevis
Genes referenced: casr gnai2 gnao1 v2ra-1 v2rb-1 v2rc-1 v2rd-1 XB5879808 [provisional:vmn2r26] xv2r1
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|Fig. 1. Deduced amino acid sequence of the newly isolated Xenopus vomeronasal receptor cDNA. Aligned deduced amino acid sequences of goldfish 5-24 (Speca et al., 1999), mouse V2R1 (Matsunami and Buck, 1997), and newly isolated Xenopus xV2R1 vomeronasal receptors. Predicted positions of the seven transmembrane domains are underlined. The identical residues are shown in white letters on a black background. Extracellular cysteine residues that are also present in mGluRs and mouse V2Rs are marked (*).|
|Fig. 2. Expression of Xenopus V2Rs in various tissues. Expression of Xenopus vomeronasal receptors (V2Rs) was analyzed by RT-PCR as described in Materials and Methods. RT-PCR products derived from total RNA isolated from the olfactory epithelium and vomeronasal organ (lanes 3, 4), olfactory bulb (lane 5), brain (lane 6), heart (lane 7), intestine (lane 8), stomach (lane 9), and kidney (lane 10) were compared to the PCR product derived from Xenopus genomic DNA (positive control) (lane 2). As a negative control, the cDNA synthesis was performed without reverse transcriptase (lane 4). The 440-bp and 160-bp PCR products (indicated by arrows) correspond to Xenopus V2Rs and -actin cDNA, respectively. The marker used was the HaeIII-digested fX174 DNA (lane 1).|
|Fig. 3. Analysis of Xenopus V2R expression in the vomeronasal organs by in situ hybridization. Cross sections of Xenopus vomeronasal epithelium were stained with hematoxylin (A). The cross sections were hybridized to digoxigenin-labeled antisense probes which were derived from clones A-1 (B), B-1 (C), C-1 (D), E-1 (E), xV2R1 (F), a mixture of A-1, C-1, and E-1 (G), and sense probes derived from a mixture of A-1, C-1, and E-1 (H). RE, receptor cell layer; SU, supporting cell layer; VL, lumen of the VNO. Black spots around the outside of the VNO in A–H (for example, arrow in F) are melanocyte aggregates. Scale bars 50 m.|
|Fig. 4. Expression of G-proteins in the vomeronasal organ (VNO), and expression of the Xenopus V2R in the principal cavity (PC) and the middle cavity (MC) of the nasal capsule. VNO cross-sections were hybridized to Go (A) and Gi2 (B) antisense probes, and to Go (C) and Gi2 (D) sense probes (negative controls). Some cells in the MC were hybridized to the Gi2-antisense probe (B’) (positive control for the Gi2-antisense probe). The antisense Xenopus V2R probes derived from clones A-1, C-1, and E-1 were mixed, and sections of Xenopus PC (E) and MC (F) epithelia were hybridized to these probes. An arrow indicates the V2R-expressing cell (F). Scale bars 50 m.|
|Fig. 5. Expression of Xenopus V2Rs in the posterolateral epithelial area of the principal cavity. Photographs of hematoxylinstained transverse sections of Xenopus laevis upper jaw at intervals of 400 m (A). Arrow indicates the V2R-expressing region (A-3). PC, Principal cavity; MC, middle cavity; VNO, vomeronasal organ. Cross-sections of Xenopus nose were hybridized to digoxigeninlabeled antisense probes prepared from clones E-1 (B), xV2R1 (C), and Go (D). Arrows indicate the xV2R1-expressing cells (C). Scale bars 1 mm in A; 100 m in B–D.|
|Fig. 6. Electron micrographs of the posterolateral epithelial area of the principal cavity. A: Microvillar receptor cells (re) and ciliated supporting cells (sp) were observed. Microvilli (arrows) on a microvillar receptor cells are indicated. Scale bar 1 m. B: Higher magnification of luminal surface of microvillar receptor cell. Microvilli are indicated (arrows). Dense border present in cell-to-cell junction (arrowhead) indicates the junctional complex. Scale bar 500 nm. C: Higher magnification of luminal surface of ciliated supporting cells. Microvillus (large arrow), kinocilia (arrowheads), ciliary rootlets (double arrow), and ciliary basal bodies (small arrows) are indicated. Scale bars 500 nm.|
|Fig. 7. Expression of Xenopus V2Rs in the olfactory and vomeronasal epithelial cells of Xenopus larvae. Cross sections of Xenopus larvae at stage 45 (A) and stage 50 (B–D) were hybridized to digoxigenin-labeled antisense probes prepared from clones E-1 (A,B,D) and xV2R1 (C). Arrows indicate the receptor-expressing cells. OE, olfactory epithelium; VNO, vomeronasal organ. Scale bars 50 um.|
|v2ra-1 (vomeronasal receptor A-1) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 66, sagittal sections of the olfactory region.|
|v2rb-1 (vomeronasal receptor B-1) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 66, sagittal sections of the olfactory region.|
|v2rc-1 (vomeronasal receptor C-1) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 66, sagittal sections of the olfactory region.|
|xv2r1 (pheromone receptor-like) gene expression in Xenopus laevis embryo, assayed via in situ hybridization, NF stage 66, sagittal sections of the olfactory region.|