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A transient receptor potential channel expressed in taste receptor cells

Abstract

We used differential screening of cDNAs from individual taste receptor cells to identify candidate taste transduction elements in mice. Among the differentially expressed clones, one encoded Trpm5, a member of the mammalian family of transient receptor potential (TRP) channels. We found Trpm5 to be expressed in a restricted manner, with particularly high levels in taste tissue. In taste cells, Trpm5 was coexpressed with taste-signaling molecules such as α-gustducin, Gγ13, phospholipase C-β2 (PLC-β2) and inositol 1,4,5-trisphosphate receptor type III (IP3R3). Our heterologous expression studies of Trpm5 indicate that it functions as a cationic channel that is gated when internal calcium stores are depleted. Trpm5 may be responsible for capacitative calcium entry in taste receptor cells that respond to bitter and/or sweet compounds.

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Figure 1: Distribution of Trpm5 mRNA in mouse tissues.
Figure 2: Expression of Trpm5 mRNA in taste receptor cells.
Figure 3: Coexpression of Trpm5, Gγ13 and PLC-β2 in taste receptor cells.
Figure 4: Single taste receptor cell expression of Trpm5, α-gustducin, Gβ1, Gβ3, Gγ13 and PLC-β2.
Figure 5: Heterologous expression of Trpm5 in X. laevis oocytes.
Figure 6: Heterologous expression of Trpm5 in CHO cells.
Figure 7: Trpm5 facilitates Ca2+ influx in X. laevis oocytes.

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Acknowledgements

We are grateful to M. Cahalan, D. Logothetis and S. Kinnamon for critical reading of the manuscript. R.F.M. is an Associate Investigator of the Howard Hughes Medical Institute. This research was supported by grants from the U.S. National Institutes of Health: DC03055 and DC03155 (R.F.M.), MH57241 (M.M.) and DC00310 (L.H.).

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Correspondence to Robert F. Margolskee.

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Supplementary information

Supplementary Fig. 1.

Identification of differentially expressed cDNAs in taste receptor cells. Individual clones picked from a cDNA library generated from a single α-gust+ TRC were arrayed in duplicate on nylon membranes then hybridized in parallel with radiolabeled probes from an α-gust+ (Membrane 1) or an α-gust (Membrane 2) taste cell. The first row of each membrane contained the following cDNAs as internal standards: Gβ1 (A1/A'1), a serial dilution of G3PDH (A3/A'3, A4/A'4, A5/A'5, A6/A'6: 50, 5, 0.5, 0.05 ng, respectively), RGS2 (A7/A'7), α-gustducin (A8/A'8, A9/A'9: 50, 5 ng, respectively), cytokeratin 8 (A10/A'10), Gβ3 (A11/A'11), ABC transporter (A12/A'12), or no DNA (A2/A'2). Spots B1/B'1 through H12/H'12 contained 84 cDNA inserts isolated from clones from a cDNA library derived from a single α-gust+ TRC. Unless otherwise specified each spot contained 50 ng of a cDNA insert. Spot B3/B'3 contained lqseq91 (subsequently identified as TRPM5). (JPG 33 kb)

Supplementary Fig. 2.

Expressed sequence tag (est) matches to murine and human TRPM5 genes. For each panel the top line displays a region of ~25,000 bp containing the Trpm5/TRPM5 and adjacent Tssc1/TSSC1 genes, respectively. The second line displays the intron/exon pattern of the Trpm5/TRPM5 genes (green boxes represent exons; green lines indicate spliced out introns). The genomic sequence of the region of the TRPM5 genes from mouse (a) and human (b) were used as queries to search the mouse and human expressed sequence tag (est) databases, respectively. Matches to the est databases are shown as solid red or blue bars with red representing a higher degree of identity. Black hatched lines show gaps in the genomic sequence between contiguous sequences that had high identity matches. Matches with Trpm5/TRPM5 exons are boxed in light blue, while matches with presumed intronic sequences of Trpm5/TRPM5 are boxed in yellow. Matches to Trpm5/TRPM5 exons in the est database were few and for mouse (a) only found in embryonic or in a few cases neonatal tissues (see list of est hits below), suggesting that Trpm5/TRPM5 expression is not widespread. In contrast, the adjacent Tssc1/TSSC1 gene has many more "hits" in the est database than does Trpm5/TRPM5. There were, however, many hits from many different tissues for the Trpm5/TRPM5 est corresponding to the introns, suggesting that an expressed repetitive element or remnant of an expressed pseudogene exists within this region of Trpm5/TRPM5 genomic DNA. Tissue distribution of est matches to Trpm5 exons (demarcated within blue box in a): 12 day embryo diaphragm/neck library; 15 day embryo head library; 13 day embryo stomach library; 16 day embryo head library; 13 day embryo lung library; 14 day embryo liver library; 13 day embryo forelimb library; 0 day neonate lung; 0 day neonate eye. Tissue distribution of est matches to TRPM5 exons (demarcated within blue box in b): CGAP library, Clontech pancreas library, colon library. (GIF 32 kb)

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Pérez, C., Huang, L., Rong, M. et al. A transient receptor potential channel expressed in taste receptor cells. Nat Neurosci 5, 1169–1176 (2002). https://doi.org/10.1038/nn952

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