Compare and contrast how the signaling cascade of olfactory receptors with T1Rs and T2Rs and their ...

(A) T1Rs (left; shown in red and blue) and T2Rs (right; shown in green) function as taste receptors (TRs) for sweet (T1R2+T1R3), umami (T1R1+T1R3) and bitter (T2R) tasting stimuli. T1R2 and T1R3 contain many allosteric binding sites for sweeteners and sweet taste inhibitors [29,63,81–83], although sugars such as glucose and sucrose bind to the clamshell domains at the amino end of each subunit [74]. (B) The current model for the transduction of sweet, umami and bitter stimuli in the gustatory system is shown. When a tastant binds to a T1R or T2R receptor it activates a G protein-coupled signaling cascade that leads to the production of the second messenger IP3, the release of Ca2+ from intracellular stores, the opening of Ca2+-gated TRPM5 cation channels, and depolarization of the taste cell. The subunits of the heterotrimeric G-protein are represented by α, β and γ.

Gβγ-mediated signaling

In general, T2Rs and T1Rs ... are expressed in largely non-overlapping subsets of Type II taste cells. However, T1Rs and T2Rs generally activate the same downstream signaling effectors in Type II taste cells. Taste receptor binding leads to activation of a heterotrimeric G protein, which consists in most cells of Gα–gustducin (McLaughlin et al., 1992) and it’s βγ partners, β3γ13 (Huang et al., 1999). The dominant leg of the pathway is mediated by the βγ partners, which activate phospholipase Cβ2 (PLCβ2) (Rossler et al., 1998) to convert the membrane lipid PIP2 into the second messengers 1,4,5- inositol trisphosphate (IP3) and diacylglycerol (DAG). While the function of DAG remains unclear, IP3 binds to the Type III IP3 receptor (IP3R3), causing release of Ca2+ from intracellular stores and subsequent Ca2+-dependent activation of a monovalent selective cation channel, transient receptor potential channel M5 (TrpM5) (Perez et al., 2002, Zhang et al., 2007). This leads to membrane depolarization, action potential generation, and release of ATP through gap junction hemichannels, likely composed of pannexin-1. Recent evidence suggests that Type II taste cells also express the vesicular ATP transporter, VNUT. This leaves open the possibility that ATP may also be released in a vesicular manner (Iwatsuki et al., 2009). Evidence for this signaling pathway comes from immunocytochemical and molecular studies showing that the component signaling effectors are co-expressed in both bitter and sweet/umami responsive Type II taste cells (Clapp et al., 2001, Clapp et al., 2004, DeFazio et al., 2006). Further, stimulation of isolated Type II taste cells with bitter, sweet, or umami taste stimuli elicits increases in intracellular Ca2+ that do not require extracellular Ca2+, are blocked by the PLC inhibitor U73122, and are sensitive to thapsigargin, which inhibits the Ca2+ ATPase that refills intracellular Ca2+ stores (Ogura et al., 2002, Hacker et al., 2008). Finally, knockout of PLCβ2, TrpM5 or IP3R3 strongly reduces or eliminates afferent nerve responses to most bitter, sweet, and umami taste stimuli (Zhang et al., 2003, Damak et al., 2006, Hisatsune et al., 2007).

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Gα-mediated signaling

All T2R receptors, and the T1R receptors in the anterior tongue and palate, are co-expressed with the Gα subunit, gustducin (Adler et al., 2000, Kim et al., 2003, Stone et al., 2007). Gα-gustducin (α-gust) was the first protein to be molecularly identified in taste cells (McLaughlin et al., 1992), but its role in taste signal transduction is still not completely understood. Gustducin has considerable sequence homology to transducin, which is also expressed in taste buds (McLaughlin et al., 1994). By analogy to the visual system, both α-gust and α-transducin are expected to activate a phosphodiesterase (PDE) and decrease intracellular cAMP levels. Biochemical studies have shown that bitter stimuli do decrease intracellular cAMP levels, and the decrease is inhibited by antibodies to α-gust (Yan et al., 2001). Cyclic AMP is also decreased in taste tissue in response to umami stimuli (Abaffy et al., 2003). However, many studies have shown that sugars increase cAMP levels in taste tissue (Bernhardt et al., 1996) and the increase is not simply a secondary consequence of Gβγ mediated release of Ca2+ from intracellular stores (Trubey et al., 2006). Gustducin knockout mice are significantly compromised to bitter, sweet, and umami stimuli, but the effect for sweet is much less than for bitter and umami (Wong et al., 1996, Ruiz et al., 2003, Glendinning et al., 2005). Thus the role of α-gust in sweet taste is much less clear than for bitter and umami taste. Part of the lack of effect of the gustducin knockout on sweet taste is that the sweet receptor T1R3 is not generally co-expressed with α-gust in posterior tongue. Instead, T1R3 is usually co-expressed with a Gq family protein, Gα14 in circumvallate and foliate taste buds (Tizzano et al., 2008, Shindo et al., 2008). Whether Gα14 mediates sweet transduction in these taste fields awaits studies in Gα14 knockout mice.

What is the role of the decreased cAMP in taste signaling? Although molecular evidence has indicated expression of a cyclic nucleotide gated channel in taste buds (Misaka et al., 1997), there is no physiological evidence for cyclic nucleotide-gated currents in taste cells. To determine other possible functions of the gustducin-mediated decrease in cAMP, biochemical assays were performed on isolated circumvallate taste buds of gustducin knockout mice. Interestingly, the knockout mice were found to have highly elevated levels of cAMP relative to wildtype mice (Clapp et al., 2008). These levels were elevated in the absence of any taste stimuli, suggesting that the taste receptors and/or G protein have tonic activity in the absence of taste ligands. The elevated cAMP likely activates Protein Kinase A to phosphorylate and inhibit PLC signaling effectors, since H-89, a specific Protein Kinase A inhibitor, rescued responses to bitter stimuli in the taste cells of gustducin knockout mice (Clapp et al., 2008). These data suggest that gustducin tonically regulates cAMP levels in taste cells to keep phosphorylation levels low and prevent chronic adaptation to bitter taste stimuli. Whether α-gust plays a similar role in the transduction of umami and sweet taste has not been determined.