Subscriber access provided by University of Florida | Smathers Libraries
Perspective
Vertebrate Bitter Taste Receptors: Keys for Survival in Changing Environments Maik Behrens, and Wolfgang Meyerhof J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.6b04835 • Publication Date (Web): 25 Dec 2016 Downloaded from http://pubs.acs.org on December 27, 2016
Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.
Journal of Agricultural and Food Chemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.
Page 1 of 41
Journal of Agricultural and Food Chemistry
Perspectives
Vertebrate Bitter Taste Receptors: Keys for Survival in Changing Environments
Maik Behrens and Wolfgang Meyerhof German Institute of Human Nutrition Potsdam-Rehbruecke, Dept. Molecular Genetics Correspondence: Dr. Maik Behrens, German Institute of Human Nutrition PotsdamRehbruecke, Dept. Molecular Genetics, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany. Fax: +49 33200 88 2384; Phone: +49 33200 88 2545; e-mail:
[email protected] 1 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 2 of 41
1
Abstract
2
Research on bitter taste receptors has made enormous progress during the recent years. While
3
in the early period after the discovery of this highly interesting receptor family special
4
emphasis was placed on the deorphanization of mainly human bitter taste receptors, the
5
research focus has shifted to sophisticated structure-function analyses, the discovery of small
6
molecule interactors, and the pharmacological profiling of non-human bitter taste receptors.
7
These findings allowed novel perspectives on e.g. evolutionary and ecological questions that
8
have arisen and that are discussed.
9
10
Key words
11
Bitter taste perception; G protein-coupled receptors
12
2 ACS Paragon Plus Environment
Page 3 of 41
Journal of Agricultural and Food Chemistry
13
Introduction
14
The ability of vertebrates to sense bitterness is thought to be important for the avoidance of
15
potentially toxic compounds occurring frequently in nature, although a clear correlation
16
between bitterness and toxicity is lacking.1 The detection of these substances is mediated by
17
G protein-coupled receptors belonging to the taste 2 receptor (TAS2R) family that are present
18
in specialized taste receptor cells located on the tongue and in the oral cavity. Following their
19
discovery in the year 2000,2-4 enormous progress has been made including the functional
20
characterization, the establishment of intra- and extraoral expression patterns, the
21
determination of structure-function relationships and other biochemical as well as cell
22
biological details. More recently, the identification of bitter taste receptor repertoires of a
23
larger collection of vertebrates and the acquisition of the agonist profiles detected by some of
24
these receptors allowed better insights in the evolutionary processes shaping these highly
25
interesting proteins. However, the answers to many of the early questions resulted in new, so
26
far unanswered questions, which need to be addressed in the future. Rather than reviewing all
27
aspects of bitter taste research, the present article will highlight only some of the past
28
developments and achievements in the field and how they shaped current views and, likely,
29
future research directions.
30
31
Bitter taste receptors
32
The human bitter taste receptor repertoire- The enormous variety of bitter substances is
33
detected by G protein-coupled receptors of the taste 2 receptor (gene symbol = TAS2R
34
(human), Tas2r (mouse)) family. The first functionally characterized receptors, the mouse
35
Tas2r105 (mT2R5), mouse Tas2r108 (mT2R8), and human TAS2R4 (hT2R4) were shown to
36
respond to one or maximally two of 55 diverse bitter compounds used for functional 3 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 4 of 41
37
screening, suggesting that bitter taste receptor genes could be specialized for the detection of
38
distinct agonists.3 Although the subsequently deorphaned human TAS2R16 was demonstrated
39
to respond to numerous chemically closely related β-D-glucopyranosides,5 the pronounced
40
specificity of this receptor again pointed towards a narrow range of substances detected by
41
these proteins raising the obvious question of how can so few receptors facilitate the detection
42
of almost countless and chemically diverse bitter agonists? A reasonable solution to this
43
problem came from the observation that human TAS2Rs are able to form homo- and
44
heterodimers with each other in vitro and, since the possible combinations appeared
45
unrestricted, 325 homo- and heterodimeric receptors could exist.6 However, it is still
46
unknown if bitter taste receptor heterodimers contribute to a broadening of the detectable
47
agonist spectrum as it was not possible to identify functional consequences of the
48
oligomerization despite considerable efforts.6 To date, all reported bitter taste receptor
49
responses in vitro can be ascribed to monomeric or homodimeric receptors. The discovery of
50
the much broader tuning properties of the human TAS2R14, which responded to about a
51
quarter of the tested compounds7 hinted at another possible solution for the apparent
52
discrepancy between receptor number and the plethora of bitter tastants, as some receptors
53
may contribute to the overall bitter taste profile of humans more than others. Indeed, after the
54
deorphanization of 21 of the ~25 putative functional human bitter taste receptors,8,9 it appears
55
that the number of TAS2Rs is fully sufficient to facilitate the detection of that many bitter
56
substances. In general, the human TAS2Rs can be categorized into 4 groups, the 3 receptor
57
“generalists” with extensive agonist spectra comprising of TAS2R10, TAS2R14, and
58
TAS2R46, each able to respond to about one-third of the bitter substances (their combined
59
activities suffice for the detection of about half of the bitter substances tested so far), a
60
number of narrowly tuned receptor “specialists” that detect few bitter compounds, the
61
intermediately tuned receptors representing the majority, as well as two receptors, the
4 ACS Paragon Plus Environment
Page 5 of 41
Journal of Agricultural and Food Chemistry
62
TAS2R165 and TAS2R38,10 which exhibit pronounced selectivity for defined classes of
63
chemicals (Fig. 1).
64 65
TAS2R gene variants- Shortly after the discovery of human TAS2R genes it was recognized
66
that numerous genetic polymorphisms of these genes exist with high frequencies in the human
67
population.11,12 Some of the TAS2R variants resulting from these polymorphisms were
68
subsequently shown to affect the function of the corresponding receptors contributing to
69
individual bitter taste perception. Whereas some of the genetic variations result in the
70
complete loss of receptor function due to incapacitating changes of the receptors’ polypeptide
71
chains10,13,14 or the genomic deletion of entire TAS2R genes,15-18 other variants exhibit more
72
subtle changes leading to reduced receptor responsiveness.19 The best investigated genetic
73
polymorphism in a TAS2R gene affects the receptor TAS2R38.12 The two major alleles occur
74
with rather similar frequencies in most populations and determine the ability to taste the
75
synthetic bitter substances phenylthiocarbamide (PTC) and 6-n-propyl-thiouracil (PROP).
76
The functional taster variant exhibits 3 amino acid sequence differences at positions 49, 262,
77
and 296 compared to the non-functional non-taster variant.14 Whereas the taster variant,
78
TAS2R38-P49A262V296
79
TAS2R38A49V262I296 shows no response in vitro.10 Also natural compounds activating human
80
TAS2R38 are plentiful and may thus influence food choice20 and innate immunity, since
81
TAS2R38 has been reported to respond to bacterial quorum sensing molecules and is
82
implicated in pathogen defense reactions.21 Other variations resulting in non-functional bitter
83
taste receptors affect TAS2R9 (missense mutation),13 TAS2R46 (nonsense mutation),11,16
84
TAS2R43 and TAS2R45 (whole gene deletions).15-18 Additional TAS2R variants affect the
85
receptors TAS2R16,19 TAS2R31 (former gene symbol TAS2R44), and TAS2R43,15,17
86
however, these receptors do not lose their function completely. A highly interesting case is
87
presented by receptor TAS2R16, which occurs as a low-sensitive variant with high frequency
confers
exquisite
sensitivity
for
PTC
and
PROP,
the
5 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 6 of 41
88
in some areas of the African continent, whereas outside of Africa exclusively the high-
89
sensitive variant is found.19 Since this receptor responds to various cyanogenic β-D-
90
glucopyranosides such as amygdalin from bitter almonds and linamarin from manioc, it was
91
suggested that the low-sensitive variant dominates in regions with an elevated malaria risk,
92
because a lower sensitivity for bitter vegetables containing cyanogenic-β-D-glucopyranosides
93
in humans could exert antimalarial activity causing protective sickle cell-like symptoms.19 A
94
recent report, however, challenged the regional correlation between the occurrence of low-
95
sensitive TAS2R16 alleles and malaria risk.22 Among the mentioned functional
96
polymorphisms two have been associated with non-gustatory TAS2R functions. Whereas the
97
non-functional TAS2R9V187 is associated with an elevated diabetes mellitus risk, which could
98
be attributable to its expression in enteroendocrine L-cells secreting blood glucose regulating
99
incretin hormones,13 the non-functional TAS2R38-A49V262I296, whose expression in human
100
sinonasal epithelia was detected, correlated with an increased frequency of upper-airway
101
infections.21,23 As more and more reports on extraoral expression of TAS2Rs emerge, it
102
appears likely that TAS2R-polymorphisms have profound physiological consequences apart
103
from perceptual differences.
104 105
Structure-function analyses- The thorough characterization of human TAS2Rs, on the one
106
hand raised questions about the architecture of the binding pockets that enable these receptors
107
to accommodate so many diverse bitter substances, yet maintaining an astonishing degree of
108
specificity, and, on the other hand provided the basis for careful structure-function analyses.
109
Consequently, in the recent years several studies have been devoted to elucidate structural
110
features of TAS2Rs involved in agonist activation. As these studies were subject of detailed
111
reviews24-26 only some facets of the findings shall be presented here. Already the first detailed
112
structure-function study devoted to one of the broadly tuned human bitter taste receptors, the
113
TAS2R46, found an answer to the question whether large ligand profiles may require the 6 ACS Paragon Plus Environment
Page 7 of 41
Journal of Agricultural and Food Chemistry
114
existence of multiple binding pockets rather than relying on a single binding site. By a
115
combination of functional calcium-mobilization assays, extensive site-directed mutagenesis as
116
well as in silico homology modeling and ligand docking experiments, it was shown that
117
agonists interact with the receptor in a single orthosteric binding pocket with overlapping, but
118
individual contact points.27 Recently, a subsequent study found evidence that agonists before
119
entering the orthosteric binding pocket transiently occupy a vestibular binding site, which
120
may act as a “specificity filter” for agonists.28 In light of the complex and concentrated
121
mixtures of chemicals to which TAS2Rs are exposed during eating this seems to represent an
122
appealing mechanism to enhance detection accuracy. In another study investigating the
123
likewise broadly tuned human TAS2R10 two intriguing observations were presented.29
124
Firstly, it was demonstrated that several amino acid residues located in the binding pocket of
125
this receptor were highly agonist selective, supporting the interaction with some agonists,
126
while perturbing optimal interaction with other agonists, suggesting that this receptor is
127
optimized to interact with many agonists at the expense of potentially higher affinities for
128
individual agonists (Fig. 2). Secondly, the finding that the binding mode for the toxic alkaloid
129
strychnine in TAS2R10 differs from that of the same molecule in TAS2R46 indicates that the
130
ability of different TAS2Rs to respond to the same bitter substances is not necessarily the
131
result of conserved pharmacological features “inherited” from common ancestral bitter
132
receptors, but rather evolved independently during evolution. Moreover, the above mentioned
133
studies agree with structure-function analyses of other TAS2Rs such as the chemical class-
134
specific TAS2R1630 and TAS2R3831 with respect to the location of the orthosteric binding
135
pocket rather deeply buried in the upper one-third of the transmembrane domain area (Fig. 3),
136
although the involved transmembrane domains may slightly differ among these TAS2Rs.
137
Other reports suggested a more pronounced involvement of extracellular loops in ligand
138
binding of TAS2R4,32 TAS2R31, and TAS2R4333 and it remains to be seen whether these
139
residues indeed contribute to the formation of the orthosteric binding site or rather indicate the 7 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 8 of 41
140
general presence of vestibular sites in TAS2Rs. Comparing the location of binding sites of
141
TAS2Rs with those of class A GPCRs it is eminent to stress that similarities prevail25 and
142
hence, despite low overall sequence homology of TAS2Rs with other GPCRs, the structures
143
and functional principles of TAS2Rs are far less exotic than initially thought.
144 145
Tas2r repertoires of other vertebrates- From an evolutionary perspective the bitter taste
146
receptor gene family represents a rather recent addition to the GPCR superfamily traceable
147
back to teleostean fish (bony fish), but absent in cartilaginous fish such as elephant sharks.34
148
Even though the history of bitter taste receptor genes is not as long as those for many other
149
GPCRs, their evolution has been more dynamic leading to rapid diversification of the Tas2r
150
genes. This is indicated by both, substantial sequence variation among Tas2r paralogs and
151
considerable differences in the sizes of Tas2r gene repertoires among vertebrates, which range
152
from 0-1 putatively functional genes in penguins35 and cetaceans (including e.g. whales)36-40
153
over ~25 in humans41 to almost 80 in the coelacanth.42 Not surprisingly, the number of
154
pseudogenes is also subject to intense variation. Some hypotheses that could explain the
155
considerable variability of the numbers of potentially intact bitter receptors have been
156
formulated and may help to understand why humans fit right in between the extremes,
157
although throughout human history dietary habits, including the acceptance of bitter food
158
items, were clearly influenced by changing sociocultural factors as well (for reviews see43,44).
159
One such hypothesis is that a low number of intact bitter taste receptor genes indicates
160
inferior bitter tasting abilities or even the complete loss of the sense of taste. Indeed, some
161
animals that swallow their prey whole such as dolphins and other cetaceans have lost all or
162
almost all of their taste receptors. Similarly, it has been speculated that chickens, which do not
163
possess a functional sweet receptor and carry only 3 intact bitter taste receptor genes in the
164
genome, have inferior tasting abilities (for a review see45).
8 ACS Paragon Plus Environment
Page 9 of 41
Journal of Agricultural and Food Chemistry
165
Several recent reports addressed the relationship between the numbers of bitter taste receptor
166
genes in broader set of vertebrates with the corresponding dietary habits.37,39,46,47 In general, it
167
seems that herbivores, who more frequently encounter bitter substances than carnivores
168
possess more Tas2r genes. Whereas some studies found a positive correlation between diet
169
and Tas2r gene numbers,37,39,46 other studies failed to obtain significant differences.47 Several
170
reasons may exist for a somewhat skewed relationship between dietary habits and the number
171
of bitter taste receptors. Firstly, at least in some herbivore species the tolerance for the
172
consumption of bitter plant constituents may result from improved degradation mechanisms
173
that have co-evolved.48 Secondly, there is not a strict correlation between bitterness and
174
toxicity1 and therefore some variability in the receptor numbers may not immediately affect
175
the chances for survival of species in particular in highly specific habitats. Thirdly, some
176
bitter substances have even beneficial health effects, e.g. in cases of infections with worms or
177
other pathogens, which would suggest a role of Tas2rs in active seeking behavior for
178
medicinal plants49,50 and therefore a selective benefit beyond nutritional needs appears likely.
179
Fourthly, and related to the last point, it is still a matter of debate whether the vertebrates’
180
bitter sensing system has some discriminative capacity (c.f.51 and references therein) and thus,
181
some bitter substances could be tolerated, while others lead to rejection behavior. If
182
discrimination among bitter substances is possible and, in turn, connected with specialized
183
Tas2rs for, e.g. rejection, the simple counting of functional Tas2rs would insufficiently
184
describe dietary preferences. Fifthly, and perhaps most importantly- bitter taste receptor
185
expression is not restricted to the oral cavity, an ever growing number of non-gustatory
186
tissues were reported indicating roles beyond taste (for recent reviews see23,52,53). While some
187
of the expression sites such as the gastrointestinal tract may indicate an interaction with food
188
derived xenobiotics analogous to the role of Tas2rs in the oral cavity, their expression in other
189
tissues such as respiratory tract (for a recent review see23) , brain,54-57 mast cells58 and white
9 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 10 of 41
190
blood cells,59-62 testis (for a recent review see63), or heart,64,65 to name just a few are difficult
191
to correlate at present with dietary habits.
192
Some of the above speculations were nourished by the fact that the knowledge about the
193
functions of bitter taste receptors were strongly human biased since no comprehensive
194
analyses of other vertebrate Tas2r was performed until recently. The functional
195
characterization of a number of non-human Tas2rs shed some light on the functional
196
relationships among Tas2rs of different clades. Whereas previous studies concentrated on the
197
characterization of single or few Tas2rs from other species such as rodents, fish, and primates,
198
recently more comprehensive analyses were published on avian, amphibian,66 carnivores67,68
199
and mouse (Fig. 4).69
200
An important outcome of the characterization of chicken and turkey Tas2rs was that very
201
small bitter taste receptor repertoires represented by the 3 chicken and 2 turkey receptors do
202
not necessarily indicate inferior bitter tasting abilities.66 It was shown that the Tas2rs of
203
chicken and turkey are on average very broadly tuned and therefore, their low number is at
204
least partially compensated by tuning breadth. On the other hand, a large number of Tas2rs as
205
in the cases of mice and the Western clawed-frog X. tropicalis apparently allows the
206
development of highly specialized receptors.66,69 The Tas2r repertoire of the domestic cat (F.
207
catus) is until now the only functionally characterized bitter taste receptor repertoire within
208
the order of carnivores67,68 and exhibits, albeit a relative small Tas2r gene number with 12
209
potentially intact genes, similar characteristics possessing broadly tuned as well as
210
intermediate and narrowly tuned receptors.67 The analyses of mouse Tas2rs revealed more
211
interesting details. On the one hand, it was demonstrated that among the 35 putatively
212
functional receptors only a single receptor can be considered broadly tuned, whereas more
213
narrowly tuned receptors exist. On the other hand and most surprisingly, it was reported that
214
orthologous receptors are not functionally conserved.69 In fact, for none of the compared
215
mouse and human orthologs an unambiguous functional conservation could be demonstrated 10 ACS Paragon Plus Environment
Page 11 of 41
Journal of Agricultural and Food Chemistry
216
indicating that even receptor pairs whose sequence was well conserved after the split of
217
rodent and primate lineages contribute to diversification of bitter recognition rather than the
218
detection of common agonists.69
219
An interesting possibility to investigate the evolutionary development of bitter taste receptor
220
genes results from the availability of functional data on a large number of Tas2rs and detailed
221
structure-function analyses on selected reference receptors. Combining such data Lossow and
222
colleagues69 were able to conclude that species specific Tas2r gene expansions generated
223
diversified receptor arrays by permutation of few critical positions located in the ligand
224
binding pockets of the Tas2r. This represents a highly efficient way to generate different
225
agonist selectivities with a limited number of mutations. Moreover, such comparative data can
226
be used to trace changes in receptor specificities over a range of species with a limited
227
number of functional data.
228 229
Bitter taste receptor gene expression- In the mammalian oral cavity Tas2r genes are
230
expressed in a specific subpopulation of type II taste receptor cells (TRCs), which do not
231
overlap with those TRCs that express sweet or umami taste receptors.70 It has been a matter of
232
debate whether the bitter TRCs represent uniform sensors for bitter substances expressing all
233
Tas2r genes in every cell or whether they form a heterogeneous population where each bitter
234
TRC expresses only subsets of them. On the one hand in situ hybridization data with Tas2r
235
probe mixtures2 as well as sophisticated functional complementation experiments in
236
genetically modified mouse models71 were interpreted in support of a uniform bitter TRC
237
population in rodents, on the other hand independent in situ hybridization experiments using
238
multiple probes4 and elaborate in vivo stimulation protocols on lingual slices of rats72 pointed
239
to a heterogeneous bitter TRC population. Comprehensive analyses of Tas2r mRNAs in
240
lingual tissues of human51 and mouse69 supported the existence of a heterogeneous bitter TRC
241
population. Since a non-homogeneous bitter TRC population would be a prerequisite for a 11 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 12 of 41
242
possible discrimination among different bitter compounds, these findings have important
243
implications. Although a number of studies investigated the bitter discriminatory capacity of
244
mammals, a final answer to this question is still lacking as contrasting results were obtained
245
(cf.51 and references therein). Whereas it was demonstrated that indeed all Tas2r genes are
246
expressed in gustatory tissues of the oral cavity of humans and mice51,69 and hence, support a
247
function as taste receptors, more specialized expression profiles seem to exist in extraoral
248
tissues. Whereas 7 of the 35 putative functional mouse Tas2rs and about half of the 25 human
249
TAS2Rs are expressed in cardiac tissue,65 the occurrence of these receptors in respiratory and
250
gastrointestinal epithelia seems to be more heterogeneous as evident by the differential
251
responses seen upon stimulation with different bitter compounds in rodent experiments.23,73
252
As some indications for nutrient-dependent regulation of Tas2r gene expression in
253
gastrointestinal tissues have accumulated,65,74,75 these expression profiles may exhibit
254
dynamic changes. It has to be mentioned that only few reports provided direct evidence on the
255
cell type(s) expressing Tas2rs in gastrointestinal tissues74-76 and hence, the heterogeneity of
256
bitter responsive cells could be even higher than assumed. The intriguing observation that
257
Tas2r genes expressed in cardiac tissue are clustered together on the corresponding
258
chromosomes,65 suggests that common regulatory elements in these loci exist orchestrating
259
tissue-specific expression. However, the frequent lack of identified cell types and the
260
substantial overlap in agonist profiles identified for the individual Tas2rs does currently not
261
allow conclusions about the biological meaning of the specific arrays of receptors found in
262
non-gustatory tissues.
263
Future directions- Despite the incredible gain in knowledge about bitter taste receptors
264
over the last 16 years open questions remained or have emerged in the course of the research.
265
While the architecture of the binding pockets of the broadly tuned TAS2Rs is quite well
266
understood, it is not clear how a narrow tuning breadth is achieved. Hence, structure-function 12 ACS Paragon Plus Environment
Page 13 of 41
Journal of Agricultural and Food Chemistry
267
experiments on narrowly tuned TAS2Rs and subsequent comparison with TAS2Rs possessing
268
large agonist panels are important to understand which mechanism(s) govern limited agonist
269
spectra. The prediction of bitter taste receptor structures today relies exclusively on homology
270
modeling with template structures that derive from GPCRs with low amino acid homology.
271
Hence, the corresponding models cannot be considered to represent high resolution structures.
272
Such structures would greatly improve the prediction of novel agonists, but also help to
273
design selective antagonists, which are urgently needed for basic research as well as for use in
274
pediatric medicinal formulations. Therefore, attempts to obtain experimental structures for at
275
least some of the TAS2Rs would significantly accelerate research in this area. Another rather
276
poorly understood process concerns the ligand-induced conformational changes occurring
277
during receptor activation. To date just a single study has been devoted to investigate the
278
mechanism of bitter taste receptor activation at the example of human TAS2R1.77 It would be
279
important to intensify research devoted to the activation mechanism of TAS2Rs as the gained
280
knowledge could aid rational design of small molecule modulators.
281
Only recently the relationship between the level of TAS2R38 mRNA in taste cells and human
282
bitter perception was established.78 As the expression strength of genes could be modulated
283
by epigenetic mechanisms and food items have been associated with epigenetic modifications
284
(for a review see79), it seems warranted to devote research efforts to study this interesting
285
field, especially since TAS2R gene expression is not limited to the oral cavity.
286
Bitter taste receptors cannot longer be seen solely as taste receptors, because their expression
287
and functional roles in other tissues such as the respiratory epithelia, the gastrointestinal tract,
288
testis, brain, heart, make them prime targets for drug design. Clarification of the role(s) that
289
bitter taste receptors play outside the gustatory system is of outmost importance for many
290
open and urgent questions. Whereas some of the activities exerted by the activation of bitter
291
taste receptors in epithelia such as the gastrointestinal tract or the airways might be related to
292
xenobiotic detection as well, other tissues such as brain or heart are not directly accessible by 13 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 14 of 41
293
bitter compounds from the outside environment. Here, the detection of yet unknown
294
endogenously formed ligands might be conceivable. It would be a high priority to identify
295
such endogenous ligands for bitter taste receptors and to elucidate the function of this
296
detection system. Moreover, studies aiming at the evolutionary sequence of tissues acquiring
297
bitter compound responsiveness would not only allow to identify the original function of
298
bitter taste receptors in vertebrates, it would also shed light on the interdependence of food
299
resources and the evolutionary development of bitter taste receptor pharmacology.53 Of
300
course, if bitter taste receptors have dominant functions aside from taste, the existence of a
301
gene sharing-like mechanism like that proposed for lens crystalline genes
302
impact the evolutionary flexibility of the Tas2r gene family.
80
would greatly
303
304
Bitter compounds
305
Bitter substances occur plentiful in nature and cover a wide variety of chemicals that can
306
differ in size, polarity, and chemical structures (cf.81 and references therein). Whereas many
307
bitter substances represent plant, fungal, or animal metabolites, other rich sources of bitter
308
compounds are chemical processes occurring during cooking, fermentation, or chemical
309
syntheses (Fig. 5). At present, it is impossible to judge how many bitter compounds may exist
310
in nature. Current data including synthetic compounds document over 680 bitter substances
311
based on published functional receptor screenings and information about perceptual properties
312
of chemicals.82 It appears reasonable to assume that the number of identified bitter substances
313
will increase considerably with time and may exceed one thousand easily. It is important to
314
note that the term “bitter” for the taste of these substances cannot be simply extended to
315
species other than humans. While there is usually a good overlap between substances that
316
represent aversive stimuli to other species and their bitter taste in humans, differences should
317
be anticipated. These differences could be due to different bitter taste receptor gene 14 ACS Paragon Plus Environment
Page 15 of 41
Journal of Agricultural and Food Chemistry
318
repertoires shaped to meet the corresponding ecological niches of the species, as well as
319
alternative routes leading to orosensorically-mediated aversive reactions such as irritants
320
activating TRP channels residing in the oral cavity83,84 or compounds eliciting a dry, tightened
321
mouthfeel called astringency.85,86 Not all substances that taste bitter to humans have been
322
matched with one of the 25 human TAS2Rs in vitro.8 One explanation for this unanticipated
323
outcome could be technical issues preventing the deorphanization of the remaining 4 orphan
324
human TAS2Rs which could possess the necessary recognition spectra to close this gap.
325
Another possibility is the existence of TAS2R-independent bitter detection routes. Indeed,
326
direct interaction of cell permeable bitter compounds with intracellular signaling
327
molecules87,88 as well as alternative receptors such as the nicotinic acetylcholine receptor,
328
which has been identified in taste receptor cells of rodents89 have been proposed.
329
Bitter substances exert a wide range of pharmacological activities, which include, but are not
330
limited to acute toxic effects.90 One of the most infamous toxic bitter compounds is the
331
alkaloid strychnine from the seeds of the Strychnos nux-vomica tree that acts as glycin
332
receptor antagonist leading91 to muscular convulsions. Less known, but with a similar mode
333
of action, is the sesquiterpene lactone picrotoxinin from the seeds of the plant Anamirta
334
cocculus, which inhibits GABAA-receptors92 again resulting in potentially deadly
335
convulsions. Interestingly, both compounds occur in the corresponding plants mixed with
336
structurally closely related substances called brucin and picrotin, respectively, which are
337
nevertheless less potent bitter compounds7,93 demonstrating pronounced selectivity of the
338
corresponding bitter receptors. Also the bitter substance (-)-α-thujone, the psychotropic
339
component of the liqueur absinthe, exerts its incapacitating effect via the inhibition of
340
GABAA and 5-HT receptors.94 Glycin-, GABAA-, and 5-HT-receptors are members of the
341
neurotransmitter-gated ion channels of the Cys-loop receptor family (for a review see95) and
342
hence, unrelated to Tas2rs belonging to the GPCR-family. Therefore, the protective
343
interaction of these substances with Tas2rs evolved independently. Other bitter substances 15 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 16 of 41
344
possess rather beneficial effects as they act as antimalarial agents in case of artemisinin96 or
345
can be used as analgesic, anti-inflammatory, and antipyretic drug such as D-(-)-salicin from
346
willow bark.97 As different as the pharmacological activities are, the chemical classes of bitter
347
compounds span a wide spectrum including amino acids and peptides, amines and amides,
348
esters and lactones, ketones, fatty acids, phenols, alkaloids, metal ions, N-heterocyclic
349
compounds, crown ethers, azacycloalkanes as well as urea and related substances (for a more
350
detailed list see81 and references therein).
351
Somewhat surprisingly, naturally occurring bitter substances do not appear to activate the
352
receptors better than synthetic compounds despite their presumed co-evolution. Among the
353
compounds that taste most bitter to humans we find the plant metabolite amarogentin as well
354
as the synthetic substance denatonium benzoate indicating that maximal bitterness may not be
355
a particular strong natural selector for human receptor-substance co-evolution. Since the
356
number of potent natural bitter compounds, which activate TAS2Rs already at low
357
concentrations, exceeds the number of potent synthetic substances,8 one could speculate that
358
the dominant trait affecting the evolution of the system is detection sensitivity. The number of
359
human bitter taste receptors activated by natural or synthetic bitter compounds (cf.8,9,69)
360
appears quite similar with 17 TAS2Rs being responsive to natural compounds and 20
361
receptors responding to synthetic compounds. Sixteen receptors were actually activated by
362
both. This is also true for individual compound receptor combinations. Whereas the synthetic
363
substance diphenidol with 15 cognate receptors activates the largest number of human
364
TAS2Rs, the natural substance quinine is able to elicit responses from 9 receptors. Moreover,
365
there are numerous examples for both, natural and synthetic chemicals that activate several
366
different TAS2Rs.8 Evidence is still missing to demonstrate if the number of cognate bitter
367
receptors determines the perceived bitterness of a given compound. Since overall bitter
368
chemicals stimulate their various cognate TAS2Rs with different potencies and efficacies it is
369
likely that some receptors contribute more to the perceived bitterness of a compound than 16 ACS Paragon Plus Environment
Page 17 of 41
Journal of Agricultural and Food Chemistry
370
others. Recent research started to address the question whether the ability of certain bitter
371
compounds to activate many bitter receptors is intrinsic to the substance or dependent on the
372
individual bitter taste receptor repertoire of a biological species and whether specific chemical
373
features identify broad acting bitter substances compared to compounds with a limited
374
spectrum of responding bitter taste receptors. It was shown that some substances such as the
375
antibiotic chloramphenicol, which activates 9 human TAS2Rs,8,9 also activated all 3 chicken,
376
all 2 turkey, 1 of 3 tested zebra finch, and 4 of 6 tested frog Tas2rs66 as well as two mouse
377
TAS2Rs,69 similarly diphenidol, which acted most broadly on human TAS2Rs,8 activated
378
chicken (3 of 3), turkey (2 of 2), zebra finch (2 of 3), frog (1 of 6),66 and mouse Tas2rs (6 of
379
34).69 According to in silico analyses of molecular properties of broadly versus narrowly
380
acting bitter substances it is suggested that, among other descriptors, small and globular
381
substances behave rather promiscuous, whereas large and flat molecules tend to be bitter taste
382
receptor selective.98
383
Not all molecules that bind to TAS2Rs act as agonists, rather some represent antagonists. The
384
first receptor selective but rather broad acting antagonist was discovered in a high throughput
385
screening devoted to discover small molecules able to block the bitter off-taste of the artificial
386
sulfonyl amide sweeteners saccharin and acesulfame K.99 It was demonstrated that the
387
compound 4-(2,2,3-trimethylcyclopentyl)butanoic acid, also known as GIV3727, potently
388
inhibited the activation of the two dominant bitter receptors for saccharin’s and acesulfame
389
K’s bitterness, the TAS2R31 and TAS2R43, by a competitive mode of action. Soon
390
thereafter, the first natural bitter inhibitors were identified.100 These compounds were
391
strikingly similar to bona fide agonists of the receptor TAS2R46, however, they did not
392
activate,
393
hydroxydihydrocostunolide and 3β-hydroxypelenolide, occur in the same plants that produce
394
TAS2R46 activating sesquiterpen lactones, a previously unanticipated level of complexity for
395
the bitterness of plants became evident. Moreover, all inhibitors, GIV3727 as well as the
but
rather
inhibited
the
receptor.
Since
these
substances,
3β-
17 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 18 of 41
396
natural bitter blockers, surprisingly exhibited a bivalent feature by acting as bitter inhibitors
397
on some TAS2Rs and as bitter agonists on others. After these initial reports a growing number
398
of bitter blockers were identified (e.g.32,101,102), indicating that such molecules may not be
399
rare.
400
Future directions- There are many open questions concerning bitter substances that
401
remain to be answered in the future. One of these is the existence of common chemical (core-)
402
structures that identify bitter substances. This question was already asked, and partially
403
answered, by Tancredi and colleagues many years before bitter taste receptor genes were
404
identified.103 Based on previous models predicting that the common core structure of sweet
405
molecules would consist of a hydrogen bond donor and –acceptor site at a distance of about
406
3Å,104,105 it was suggested that bitter compounds have a similar structure but with a reverse
407
orientation.103 Whereas this model indeed explained why D- and L-enantiomers of some
408
amino acids elicit pronounced sweet or bitter taste, the finding that so many differently tuned
409
TAS2Rs exist in human argues for a more complex interaction pattern. For two of the 25
410
receptors, the TAS2R16 and the TAS2R38, such common chemical structures, namely the β-
411
D-glucopyranose moiety and the isothiocyanate/thiourea moiety, respectively, were
412
identified.5,10 However, other receptors seem to respond to a larger variety of chemicals.
413
There are several possible explanations for the difficulties associated with the identification of
414
common pharmacophores for TAS2Rs: Firstly, recent evidence was provided that not only
415
bitter agonists can be found in nature but also antagonists. Since the majority of reports so far
416
relied on assays measuring receptor activation in vitro and not ligand binding, the
417
identification of the chemical core structure required for binding to the receptors without
418
leading to activation was usually not implemented. Recent publications using competitive
419
approaches making use of agonists/antagonist mixtures32,99-102,106,107 could close this gap since
420
classical binding assays suffer from the rather low affinities of most bitter compound-receptor
421
interactions and, as a consequence, no such data exist. Secondly, some bitter receptor binding 18 ACS Paragon Plus Environment
Page 19 of 41
Journal of Agricultural and Food Chemistry
422
sites may actually be composed of sub-sites, which interact with subsets of chemically diverse
423
bitter agonists. An indication for agonists based on multiple distinct chemical scaffolds was
424
recently provided for in silico predicted novel TAS2R14 agonists107 and could apply to other
425
receptors as well. Of course, this obscures the identification of individual pharmacophores
426
considerably.
427
Most likely only some (if at all) of the identified bitter activators of present day’s bitter taste
428
receptors are identical to those that shaped the evolution of the receptors. Major vertebrate
429
classes were already formed prior to the neophyticum/upper Cretaceous period when
430
angiosperm plants, which dominate our planet since then, started to conquer the earth.
431
Clearly, the early development of nowadays bitter receptors was not driven by bitter
432
substances originating from “modern” plants, but rather by toxic compounds from other
433
sources. Also climate and habitat changes that early humans faced when they started to
434
migrate out of Africa obscure the bitter substance-receptor co-evolution because plants
435
growing ~2 million years ago in Africa dominantly shaped our TAS2R gene repertoire.
436
Hence, it is anticipated that important contributions on the structures of relevant bitter
437
substances may come from botanical and paleo botanical experts who join the field of bitter
438
taste research. Finally, the relationship between bitterness and toxicity is less tight than
439
frequently assumed.1 It would be highly relevant to investigate this question from an
440
ecological point of view to see if bitterness is mostly indicating potential toxins or if it is also
441
guiding seeking behavior for therapeutic substances in case of illness49,50 and whether plants
442
may not only synthesize toxins to defend herbivores but also, as a kind of chemical
443
mimicry,108 non-toxic substances.
444
The occurrence of natural bitter receptor inhibitors raises the question of how abundant such
445
molecules in nature are and if the existence of such modulatory compounds has influenced the
446
evolution bitter taste receptor gene repertoires. It may well be that the expansion of bitter taste
447
receptor genes was not dominantly influenced by the need to recognize more and more bitter 19 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 20 of 41
448
substances, but rather to avoid as much as possible completely overlapping activities of
449
activators and inhibitors with potentially fatal outcomes.100 In the future large screening
450
campaigns to identify more natural bitter inhibitors may help to clarify this question.
451
20 ACS Paragon Plus Environment
Page 21 of 41
Journal of Agricultural and Food Chemistry
452
References
453
1.
1994, 56, 1217-1227.
454 455
2.
3.
4.
Matsunami, H.; Montmayeur, J. P.; Buck, L. B. A family of candidate taste receptors in human and mouse. Nature 2000, 404, 601-604.
460 461
Chandrashekar, J.; Mueller, K. L.; Hoon, M. A.; Adler, E.; Feng, L.; Guo, W.; Zuker, C. S.; Ryba, N. J. T2Rs function as bitter taste receptors. Cell 2000, 100, 703-711.
458 459
Adler, E.; Hoon, M. A.; Mueller, K. L.; Chandrashekar, J.; Ryba, N. J.; Zuker, C. S. A novel family of mammalian taste receptors. Cell 2000, 100, 693-702.
456 457
Glendinning, J. I. Is the bitter rejection response always adaptive? Physiol. Behav.
5.
Bufe, B.; Hofmann, T.; Krautwurst, D.; Raguse, J. D.; Meyerhof, W. The human
462
TAS2R16 receptor mediates bitter taste in response to beta-glucopyranosides. Nat.
463
Genet. 2002, 32, 397-401.
464
6.
receptors. Chem. Senses 2010, 35, 395-406.
465 466
Kuhn, C.; Bufe, B.; Batram, C.; Meyerhof, W. Oligomerization of TAS2R bitter taste
7.
Behrens, M.; Brockhoff, A.; Kuhn, C.; Bufe, B.; Winnig, M.; Meyerhof, W. The
467
human taste receptor hTAS2R14 responds to a variety of different bitter compounds.
468
Biochem. Biophys. Res. Commun. 2004, 319, 479-485.
469
8.
Meyerhof, W.; Batram, C.; Kuhn, C.; Brockhoff, A.; Chudoba, E.; Bufe, B.;
470
Appendino, G.; Behrens, M. The molecular receptive ranges of human TAS2R bitter
471
taste receptors. Chem. Senses 2010, 35, 157-170.
472
9.
Thalmann, S.; Behrens, M.; Meyerhof, W. Major haplotypes of the human bitter taste
473
receptor TAS2R41 encode functional receptors for chloramphenicol. Biochem.
474
Biophys. Res. Commun. 2013, 435, 267-273.
475 476
10.
Bufe, B.; Breslin, P. A.; Kuhn, C.; Reed, D. R.; Tharp, C. D.; Slack, J. P.; Kim, U. K.; Drayna, D.; Meyerhof, W. The molecular basis of individual differences in 21 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 22 of 41
477
phenylthiocarbamide and propylthiouracil bitterness perception. Curr. Biol. 2005, 15,
478
322-327.
479
11.
Kim, U.; Wooding, S.; Ricci, D.; Jorde, L. B.; Drayna, D. Worldwide haplotype
480
diversity and coding sequence variation at human bitter taste receptor loci. Hum.
481
Mutat. 2005, 26, 199-204.
482
12.
Wooding, S.; Kim, U. K.; Bamshad, M. J.; Larsen, J.; Jorde, L. B.; Drayna, D. Natural
483
selection and molecular evolution in PTC, a bitter-taste receptor gene. Am. J. Hum.
484
Genet. 2004, 74, 637-646.
485
13.
Dotson, C. D.; Zhang, L.; Xu, H.; Shin, Y. K.; Vigues, S.; Ott, S. H.; Elson, A. E.;
486
Choi, H. J.; Shaw, H.; Egan, J. M.; Mitchell, B. D.; Li, X.; Steinle, N. I.; Munger, S.
487
D. Bitter taste receptors influence glucose homeostasis. PLoS ONE 2008, 3, e3974.
488
14.
Kim, U. K.; Jorgenson, E.; Coon, H.; Leppert, M.; Risch, N.; Drayna, D. Positional
489
cloning of the human quantitative trait locus underlying taste sensitivity to
490
phenylthiocarbamide. Science 2003, 299, 1221-1225.
491
15.
Pronin, A. N.; Xu, H.; Tang, H.; Zhang, L.; Li, Q.; Li, X. Specific alleles of bitter
492
receptor genes influence human sensitivity to the bitterness of aloin and saccharin.
493
Curr. Biol. 2007, 17, 1403-1408.
494
16.
Roudnitzky, N.; Behrens, M.; Engel, A.; Kohl, S.; Thalmann, S.; Hubner, S.; Lossow,
495
K.; Wooding, S. P.; Meyerhof, W. Receptor Polymorphism and Genomic Structure
496
Interact to Shape Bitter Taste Perception. PLoS Genet. 2015, 11, e1005530.
497
17.
Roudnitzky, N.; Bufe, B.; Thalmann, S.; Kuhn, C.; Gunn, H. C.; Xing, C.; Crider, B.
498
P.; Behrens, M.; Meyerhof, W.; Wooding, S. P. Genomic, genetic and functional
499
dissection of bitter taste responses to artificial sweeteners. Hum. Mol. Genet. 2011, 20,
500
3437-3449.
22 ACS Paragon Plus Environment
Page 23 of 41
501
Journal of Agricultural and Food Chemistry
18.
Roudnitzky, N.; Risso, D.; Drayna, D.; Behrens, M.; Meyerhof, W.; Wooding, S. P.
502
Copy Number Variation in TAS2R Bitter Taste Receptor Genes: Structure, Origin,
503
and Population Genetics. Chem. Senses 2016, 41, 649-659.
504
19.
Soranzo, N.; Bufe, B.; Sabeti, P. C.; Wilson, J. F.; Weale, M. E.; Marguerie, R.;
505
Meyerhof, W.; Goldstein, D. B. Positive Selection on a High-Sensitivity Allele of the
506
Human Bitter-Taste Receptor TAS2R16. Curr. Biol. 2005, 15, 1257-1265.
507
20.
we taste toxins in food. Curr. Biol. 2006, 16, R792-794.
508 509
Sandell, M. A.; Breslin, P. A. Variability in a taste-receptor gene determines whether
21.
Lee, R. J.; Xiong, G.; Kofonow, J. M.; Chen, B.; Lysenko, A.; Jiang, P.; Abraham, V.;
510
Doghramji, L.; Adappa, N. D.; Palmer, J. N.; Kennedy, D. W.; Beauchamp, G. K.;
511
Doulias, P. T.; Ischiropoulos, H.; Kreindler, J. L.; Reed, D. R.; Cohen, N. A. T2R38
512
taste receptor polymorphisms underlie susceptibility to upper respiratory infection. J.
513
Clin. Invest. 2012, 122, 4145-4159.
514
22.
Campbell, M. C.; Ranciaro, A.; Zinshteyn, D.; Rawlings-Goss, R.; Hirbo, J.;
515
Thompson, S.; Woldemeskel, D.; Froment, A.; Rucker, J. B.; Omar, S. A.; Bodo, J.
516
M.; Nyambo, T.; Belay, G.; Drayna, D.; Breslin, P. A.; Tishkoff, S. A. Origin and
517
differential selection of allelic variation at TAS2R16 associated with salicin bitter
518
taste sensitivity in Africa. Mol. Biol. Evol. 2014, 31, 288-302.
519
23.
in health and disease. J. Mol. Med. (Berl.) 2014, 92, 1235-1244.
520 521
24.
Behrens, M.; Meyerhof, W. Bitter taste receptor research comes of age: from characterization to modulation of TAS2Rs. Semin. Cell Dev. Biol. 2013, 24, 215-221.
522 523
Lee, R. J.; Cohen, N. A. Bitter and sweet taste receptors in the respiratory epithelium
25.
Di Pizio, A.; Levit, A.; Slutzki, M.; Behrens, M.; Karaman, R.; Niv, M. Y. Comparing
524
Class A GPCRs to bitter taste receptors: Structural motifs, ligand interactions and
525
agonist-to-antagonist ratios. Methods Cell Biol. 2016, 132, 401-427.
23 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
526
26.
Page 24 of 41
Pydi, S. P.; Upadhyaya, J.; Singh, N.; Pal Bhullar, R.; Chelikani, P. Recent advances
527
in structure and function studies on human bitter taste receptors. Curr. Protein Pept.
528
Sci. 2012, 13, 501-508.
529
27.
bitter taste receptor activation. Proc. Natl. Acad. Sci. U S A 2010, 107, 11110-11115.
530 531
Brockhoff, A.; Behrens, M.; Niv, M. Y.; Meyerhof, W. Structural requirements of
28.
Sandal, M.; Behrens, M.; Brockhoff, A.; Musiani, F.; Giorgetti, A.; Carloni, P.;
532
Meyerhof, W. Evidence for a Transient Additional Ligand Binding Site in the
533
TAS2R46 Bitter Taste Receptor. J. Chem. Theory Comput. 2015, 11, 4439-4449.
534
29.
Born, S.; Levit, A.; Niv, M. Y.; Meyerhof, W.; Behrens, M. The human bitter taste
535
receptor TAS2R10 is tailored to accommodate numerous diverse ligands. J. Neurosci.
536
2013, 33, 201-213.
537
30.
Sakurai, T.; Misaka, T.; Ishiguro, M.; Masuda, K.; Sugawara, T.; Ito, K.; Kobayashi,
538
T.; Matsuo, S.; Ishimaru, Y.; Asakura, T.; Abe, K. Characterization of the beta-D-
539
glucopyranoside binding site of the human bitter taste receptor hTAS2R16. J. Biol.
540
Chem. 2010, 285, 28373-28378.
541
31.
Marchiori, A.; Capece, L.; Giorgetti, A.; Gasparini, P.; Behrens, M.; Carloni, P.;
542
Meyerhof, W. Coarse-grained/molecular mechanics of the TAS2R38 bitter taste
543
receptor: experimentally-validated detailed structural prediction of agonist binding.
544
PLoS One 2013, 8, e64675.
545
32.
Pydi, S. P.; Sobotkiewicz, T.; Billakanti, R.; Bhullar, R. P.; Loewen, M. C.; Chelikani,
546
P. Amino acid derivatives as bitter taste receptor (T2R) blockers. J. Biol. Chem. 2014,
547
289, 25054-25066.
548
33.
human bitter T2R receptors. Chem. Senses 2004, 29, 583-593.
549 550 551
Pronin, A. N.; Tang, H.; Connor, J.; Keung, W. Identification of ligands for two
34.
Grus, W. E.; Zhang, J. Origin of the genetic components of the vomeronasal system in the common ancestor of all extant vertebrates. Mol. Biol. Evol. 2009, 26, 407-419. 24 ACS Paragon Plus Environment
Page 25 of 41
552
Journal of Agricultural and Food Chemistry
35.
penguins. Curr. Biol. 2015, 25, R141-142.
553 554
36.
Feng, P.; Zheng, J.; Rossiter, S. J.; Wang, D.; Zhao, H. Massive losses of taste receptor genes in toothed and baleen whales. Genome Biol. Evol. 2014, 6, 1254-1265.
555 556
Zhao, H.; Li, J.; Zhang, J. Molecular evidence for the loss of three basic tastes in
37.
Jiang, P.; Josue, J.; Li, X.; Glaser, D.; Li, W.; Brand, J. G.; Margolskee, R. F.; Reed,
557
D. R.; Beauchamp, G. K. Major taste loss in carnivorous mammals. Proc. Natl. Acad.
558
Sci. U S A 2012, 109, 4956-4961.
559
38.
the evolution of smell and taste in whales. Zoological Lett. 2015, 1, 9.
560 561
Kishida, T.; Thewissen, J.; Hayakawa, T.; Imai, H.; Agata, K. Aquatic adaptation and
39.
Liu, Z.; Liu, G.; Hailer, F.; Orozco-terWengel, P.; Tan, X.; Tian, J.; Yan, Z.; Zhang,
562
B.; Li, M. Dietary specialization drives multiple independent losses and gains in the
563
bitter taste gene repertoire of Laurasiatherian Mammals. Front. Zool. 2016, 13, 28.
564
40.
genes in cetaceans. BMC Evol. Biol. 2014, 14, 218.
565 566
41.
Shi, P.; Zhang, J.; Yang, H.; Zhang, Y. P. Adaptive diversification of bitter taste receptor genes in Mammalian evolution. Mol. Biol. Evol. 2003, 20, 805-814.
567 568
Zhu, K.; Zhou, X.; Xu, S.; Sun, D.; Ren, W.; Zhou, K.; Yang, G. The loss of taste
42.
Syed, A. S.; Korsching, S. I. Positive Darwinian selection in the singularly large taste
569
receptor gene family of an 'ancient' fish, Latimeria chalumnae. BMC Genomics 2014,
570
15, 650.
571
43.
23, R409-418.
572 573
44.
576
Drewnowski, A. The science and complexity of bitter taste. Nutr. Rev. 2001, 59, 163169.
574 575
Breslin, P. A. An evolutionary perspective on food and human taste. Curr. Biol. 2013,
45.
Roura, E.; Baldwin, M.; Klasing, K. 23 rd ANNUAL AUSTRALIAN POULTRY SCIENCE SYMPOSIUM, Sydney, p 97-104.
25 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
577
46.
47.
48.
Freeland, W. J.; Janzen, D. H. Strategies in Herbivory by Mammals: The Role of Plant Secondary Compounds. Am. Nat. 1974, 108, 269-289.
582 583
Li, D.; Zhang, J. Diet shapes the evolution of the vertebrate bitter taste receptor gene repertoire. Mol. Biol. Evol. 2014, 31, 303-309.
580 581
Hu, L. L.; Shi, P. Smallest bitter taste receptor (T2Rs) gene repertoire in carnivores. Zool. Res. 2013, 34, E75-81.
578 579
Page 26 of 41
49.
Koshimizu, K.; Ohigashi, H.; Huffman, M. A. Use of Vernonia amygdalina by wild
584
chimpanzee: possible roles of its bitter and related constituents. Physiol. Behav. 1994,
585
56, 1209-1216.
586
50.
Villalba, J. J.; Miller, J.; Ungar, E. D.; Landau, S. Y.; Glendinning, J. Ruminant self-
587
medication against gastrointestinal nematodes: evidence, mechanism, and origins.
588
Parasite 2014, 21, 31.
589
51.
Behrens, M.; Foerster, S.; Staehler, F.; Raguse, J. D.; Meyerhof, W. Gustatory
590
expression pattern of the human TAS2R bitter receptor gene family reveals a
591
heterogenous population of bitter responsive taste receptor cells. J. Neurosci. 2007,
592
27, 12630-12640.
593
52.
bitter in the oral cavity. Acta Physiol. (Oxf.) 2016, 216, 407-420.
594 595
53.
Behrens, M.; Prandi, S.; Meyerhof, W. Taste Receptor Gene Expression Outside the Gustatory System. Springer Berlin Heidelberg: Berlin, Heidelberg, 2014; pp 1-34.
596 597
Avau, B.; Depoortere, I. The bitter truth about bitter taste receptors: beyond sensing
54.
Ansoleaga, B.; Garcia-Esparcia, P.; Llorens, F.; Moreno, J.; Aso, E.; Ferrer, I.
598
Dysregulation of brain olfactory and taste receptors in AD, PSP and CJD, and AD-
599
related model. Neuroscience 2013, 248C, 369-382.
600 601
55.
Dehkordi, O.; Rose, J. E.; Fatemi, M.; Allard, J. S.; Balan, K. V.; Young, J. K.; Fatima, S.; Millis, R. M.; Jayam-Trouth, A. Neuronal expression of bitter taste
26 ACS Paragon Plus Environment
Page 27 of 41
Journal of Agricultural and Food Chemistry
602
receptors and downstream signaling molecules in the rat brainstem. Brain Res. 2012,
603
1475, 1-10.
604
56.
Garcia-Esparcia, P.; Schluter, A.; Carmona, M.; Moreno, J.; Ansoleaga, B.; Torrejon-
605
Escribano, B.; Gustincich, S.; Pujol, A.; Ferrer, I. Functional genomics reveals
606
dysregulation of cortical olfactory receptors in Parkinson disease: novel putative
607
chemoreceptors in the human brain. J. Neuropathol. Exp. Neurol. 2013, 72, 524-539.
608
57.
are expressed in brain cells. Biochem. Biophys. Res. Commun. 2011, 406, 146-151.
609 610
Singh, N.; Vrontakis, M.; Parkinson, F.; Chelikani, P. Functional bitter taste receptors
58.
Ekoff, M.; Choi, J. H.; James, A.; Dahlen, B.; Nilsson, G.; Dahlen, S. E. Bitter taste
611
receptor (TAS2R) agonists inhibit IgE-dependent mast cell activation. J. Allergy Clin.
612
Immunol. 2014, 134, 475-478.
613
59.
Malki, A.; Fiedler, J.; Fricke, K.; Ballweg, I.; Pfaffl, M. W.; Krautwurst, D. Class I
614
odorant receptors, TAS1R and TAS2R taste receptors, are markers for subpopulations
615
of circulating leukocytes. J. Leukoc. Biol. 2015, 97, 533-545.
616
60.
Marcinek, P.; Geithe, C.; Krautwurst, D. Chemosensory G Protein-Coupled Receptors
617
(GPCR) in Blood Leukocytes. Springer Berlin Heidelberg: Berlin, Heidelberg; pp 1-
618
23.
619
61.
Maurer, S.; Wabnitz, G. H.; Kahle, N. A.; Stegmaier, S.; Prior, B.; Giese, T.; Gaida,
620
M. M.; Samstag, Y.; Hansch, G. M. Tasting Pseudomonas aeruginosa Biofilms:
621
Human Neutrophils Express the Bitter Receptor T2R38 as Sensor for the Quorum
622
Sensing Molecule N-(3-Oxododecanoyl)-l-Homoserine Lactone. Front. Immunol.
623
2015, 6, 369.
624
62.
Orsmark-Pietras, C.; James, A.; Konradsen, J. R.; Nordlund, B.; Soderhall, C.;
625
Pulkkinen, V.; Pedroletti, C.; Daham, K.; Kupczyk, M.; Dahlen, B.; Kere, J.; Dahlen,
626
S. E.; Hedlin, G.; Melen, E. Transcriptome analysis reveals upregulation of bitter taste
627
receptors in severe asthmatics. Eur. Respir. J. 2013, 42, 65-78. 27 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
628
63.
Li, F. Taste perception: from the tongue to the testis. Mol. Hum. Reprod. 2013, 19, 349-360.
629 630
Page 28 of 41
64.
Foster, S. R.; Blank, K.; See Hoe, L. E.; Behrens, M.; Meyerhof, W.; Peart, J. N.;
631
Thomas, W. G. Bitter taste receptor agonists elicit G-protein-dependent negative
632
inotropy in the murine heart. FASEB J. 2014, 28, 4497-4508.
633
65.
Foster, S. R.; Porrello, E. R.; Purdue, B.; Chan, H. W.; Voigt, A.; Frenzel, S.; Hannan,
634
R. D.; Moritz, K. M.; Simmons, D. G.; Molenaar, P.; Roura, E.; Boehm, U.;
635
Meyerhof, W.; Thomas, W. G. Expression, regulation and putative nutrient-sensing
636
function of taste GPCRs in the heart. PLoS One 2013, 8, e64579.
637
66.
Behrens, M.; Korsching, S. I.; Meyerhof, W. Tuning properties of avian and frog bitter
638
taste receptors dynamically fit gene repertoire sizes. Mol. Biol. Evol. 2014, 31, 3216-
639
3227.
640
67.
Lei, W.; Ravoninjohary, A.; Li, X.; Margolskee, R. F.; Reed, D. R.; Beauchamp, G.
641
K.; Jiang, P. Functional Analyses of Bitter Taste Receptors in Domestic Cats (Felis
642
catus). PLoS One 2015, 10, e0139670.
643
68.
Sandau, M. M.; Goodman, J. R.; Thomas, A.; Rucker, J. B.; Rawson, N. E. A
644
functional comparison of the domestic cat bitter receptors Tas2r38 and Tas2r43 with
645
their human orthologs. BMC Neurosci. 2015, 16, 33.
646
69.
Lossow, K.; Hubner, S.; Roudnitzky, N.; Slack, J. P.; Pollastro, F.; Behrens, M.;
647
Meyerhof, W. Comprehensive Analysis of Mouse Bitter Taste Receptors Reveals
648
Different Molecular Receptive Ranges for Orthologous Receptors in Mice and
649
Humans. J. Biol. Chem. 2016, 291, 15358-15377.
650
70.
mammalian taste. Nature 2006, 444, 288-294.
651 652 653
Chandrashekar, J.; Hoon, M. A.; Ryba, N. J.; Zuker, C. S. The receptors and cells for
71.
Mueller, K. L.; Hoon, M. A.; Erlenbach, I.; Chandrashekar, J.; Zuker, C. S.; Ryba, N. J. The receptors and coding logic for bitter taste. Nature 2005, 434, 225-229. 28 ACS Paragon Plus Environment
Page 29 of 41
654
Journal of Agricultural and Food Chemistry
72.
Science 2001, 291, 1557-1560.
655 656
Caicedo, A.; Roper, S. D. Taste receptor cells that discriminate between bitter stimuli.
73.
Janssen, S.; Laermans, J.; Verhulst, P. J.; Thijs, T.; Tack, J.; Depoortere, I. Bitter taste
657
receptors and alpha-gustducin regulate the secretion of ghrelin with functional effects
658
on food intake and gastric emptying. Proc. Natl. Acad. Sci. U S A 2011, 108, 2094-
659
2099.
660
74.
Jeon, T. I.; Zhu, B.; Larson, J. L.; Osborne, T. F. SREBP-2 regulates gut peptide
661
secretion through intestinal bitter taste receptor signaling in mice. J. Clin. Invest.
662
2008, 118, 3693-3700.
663
75.
Vegezzi, G.; Anselmi, L.; Huynh, J.; Barocelli, E.; Rozengurt, E.; Raybould, H.;
664
Sternini, C. Diet-induced regulation of bitter taste receptor subtypes in the mouse
665
gastrointestinal tract. PLoS One 2014, 9, e107732.
666
76.
Prandi, S.; Bromke, M.; Hubner, S.; Voigt, A.; Boehm, U.; Meyerhof, W.; Behrens,
667
M. A subset of mouse colonic goblet cells expresses the bitter taste receptor Tas2r131.
668
PLoS One 2013, 8, e82820.
669
77.
Singh, N.; Pydi, P.; Upadhyaya, J.; Chelikani, P. Structural basis of activation of bitter
670
taste receptor T2R1 and comparison with class A GPCRs. J. Biol. Chem. 2011, 286,
671
36032-36041.
672
78.
Lipchock, S. V.; Mennella, J. A.; Spielman, A. I.; Reed, D. R. Human bitter
673
perception correlates with bitter receptor messenger RNA expression in taste cells.
674
Am. J. Clin. Nutr. 2013, 98, 1136-1143.
675
79.
Opin. Clin. Nutr. Metab. Care 2015, 18, 334-338.
676 677 678
Haggarty, P. Genetic and metabolic determinants of human epigenetic variation. Curr.
80.
Piatigorsky, J.; Wistow, G. J. Enzyme/crystallins: gene sharing as an evolutionary strategy. Cell 1989, 57, 197-199.
29 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
679
81.
82.
Wiener, A.; Shudler, M.; Levit, A.; Niv, M. Y. BitterDB: a database of bitter compounds. Nucleic Acids Res. 2012, 40, D413-419.
682 683
Meyerhof, W. Elucidation of mammalian bitter taste. Rev. Physiol. Biochem. Pharmacol. 2005, 154, 37-72.
680 681
Page 30 of 41
83.
Mancuso, G.; Borgonovo, G.; Scaglioni, L.; Bassoli, A. Phytochemicals from Ruta
684
graveolens Activate TAS2R Bitter Taste Receptors and TRP Channels Involved in
685
Gustation and Nociception. Molecules 2015, 20, 18907-18922.
686
84.
827-871.
687 688
Roper, S. D. TRPs in taste and chemesthesis. Handb. Exp. Pharmacol. 2014, 223,
85.
Fleming, E. E.; Ziegler, G. R.; Hayes, J. E. Salivary protein levels as a predictor of
689
perceived astringency in model systems and solid foods. Physiol. Behav. 2016, 163,
690
56-63.
691
86.
Schobel, N.; Radtke, D.; Kyereme, J.; Wollmann, N.; Cichy, A.; Obst, K.; Kallweit,
692
K.; Kletke, O.; Minovi, A.; Dazert, S.; Wetzel, C. H.; Vogt-Eisele, A.; Gisselmann,
693
G.; Ley, J. P.; Bartoshuk, L. M.; Spehr, J.; Hofmann, T.; Hatt, H. Astringency Is a
694
Trigeminal Sensation That Involves the Activation of G Protein-Coupled Signaling by
695
Phenolic Compounds. Chem. Senses 2014.
696
87.
Gees, M.; Alpizar, Y. A.; Luyten, T.; Parys, J. B.; Nilius, B.; Bultynck, G.; Voets, T.;
697
Talavera, K. Differential effects of bitter compounds on the taste transduction
698
channels TRPM5 and IP3 receptor type 3. Chem. Senses 2014, 39, 295-311.
699
88.
Peri, I.; Mamrud-Brains, H.; Rodin, S.; Krizhanovsky, V.; Shai, Y.; Nir, S.; Naim, M.
700
Rapid entry of bitter and sweet tastants into liposomes and taste cells: implications for
701
signal transduction. Am. J. Physiol. Cell Physiol. 2000, 278, C17-25.
702
89.
Ren, Z. J.; Mummalaneni, S.; Qian, J.; Baumgarten, C. M.; DeSimone, J. A.; Lyall, V.
703
Nicotinic Acetylcholine Receptor (nAChR) Dependent Chorda Tympani Taste Nerve
704
Responses to Nicotine, Ethanol and Acetylcholine. PLoS One 2015, 10, e0127936. 30 ACS Paragon Plus Environment
Page 31 of 41
705
Journal of Agricultural and Food Chemistry
90.
Today 2002, 37, 144-150.
706 707
91.
Young, A. B.; Snyder, S. H. Strychnine binding associated with glycine receptors of the central nervous system. Proc. Natl. Acad. Sci. U S A 1973, 70, 2832-2836.
708 709
Barratt-Fornell, A.; Drewnowski, A. The Taste of Health: Nature's Bitter Gifts. Nutr.
92.
Perret, P.; Sarda, X.; Wolff, M.; Wu, T. T.; Bushey, D.; Goeldner, M. Interaction of
710
non-competitive blockers within the gamma-aminobutyric acid type A chloride
711
channel using chemically reactive probes as chemical sensors for cysteine mutants. J.
712
Biol. Chem. 1999, 274, 25350-25354.
713
93.
Brockhoff, A.; Behrens, M.; Massarotti, A.; Appendino, G.; Meyerhof, W. Broad
714
tuning of the human bitter taste receptor hTAS2R46 to various sesquiterpene lactones,
715
clerodane and labdane diterpenoids, strychnine, and denatonium. J. Agric. Food
716
Chem. 2007, 55, 6236-6243.
717
94.
Deiml, T.; Haseneder, R.; Zieglgansberger, W.; Rammes, G.; Eisensamer, B.;
718
Rupprecht, R.; Hapfelmeier, G. Alpha-thujone reduces 5-HT3 receptor activity by an
719
effect on the agonist-reduced desensitization. Neuropharmacology 2004, 46, 192-201.
720
95.
Miller, P. S.; Smart, T. G. Binding, activation and modulation of Cys-loop receptors. Trends Pharmacol. Sci. 2010, 31, 161-174.
721 722
96.
Hien, T. T.; White, N. J. Qinghaosu. Lancet 1993, 341, 603-608.
723
97.
Vane, J. R. The fight against rheumatism: from willow bark to COX-1 sparing drugs. J. Physiol. Pharmacol. 2000, 51, 573-586.
724 725
98.
receptors. Bioorg. Med. Chem. 2015, 23, 4082-4091.
726 727
Di Pizio, A.; Niv, M. Y. Promiscuity and selectivity of bitter molecules and their
99.
Slack, J. P.; Brockhoff, A.; Batram, C.; Menzel, S.; Sonnabend, C.; Born, S.; Galindo,
728
M. M.; Kohl, S.; Thalmann, S.; Ostopovici-Halip, L.; Simons, C. T.; Ungureanu, I.;
729
Duineveld, K.; Bologa, C. G.; Behrens, M.; Furrer, S.; Oprea, T. I.; Meyerhof, W.
31 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 32 of 41
730
Modulation of bitter taste perception by a small molecule hTAS2R antagonist. Curr.
731
Biol. 2010, 20, 1104-1109.
732
100.
Brockhoff, A.; Behrens, M.; Roudnitzky, N.; Appendino, G.; Avonto, C.; Meyerhof,
733
W. Receptor agonism and antagonism of dietary bitter compounds. J. Neurosci. 2011,
734
31, 14775-14782.
735
101.
Greene, T. A.; Alarcon, S.; Thomas, A.; Berdougo, E.; Doranz, B. J.; Breslin, P. A.;
736
Rucker, J. B. Probenecid inhibits the human bitter taste receptor TAS2R16 and
737
suppresses bitter perception of salicin. PLoS One 2011, 6, e20123.
738
102.
Roland, W. S.; Gouka, R. J.; Gruppen, H.; Driesse, M.; van Buren, L.; Smit, G.;
739
Vincken, J. P. 6-Methoxyflavanones as Bitter Taste Receptor Blockers for hTAS2R39.
740
PLoS One 2014, 9, e94451.
741
103.
receptor site. Chem. Senses 1979, 4, 259-265.
742 743
104.
Shallenberger, R. S.; Acree, T. E. Molecular theory of sweet taste. Nature 1967, 216, 480-482.
744 745
Tancredi, T.; Lelj, F.; Temussi, P. A. Three-dimensional mapping of the bitter taste
105.
Shallenberger, R. S.; Acree, T. E.; Lee, C. Y. Sweet taste of D and L-sugars and
746
amino-acids and the steric nature of their chemo-receptor site. Nature 1969, 221, 555-
747
556.
748
106.
Karaman, R.; Nowak, S.; Di Pizio, A.; Kitaneh, H.; Abu-Jaish, A.; Meyerhof, W.; Niv,
749
M. Y.; Behrens, M. Probing the Binding Pocket of the Broadly Tuned Human Bitter
750
Taste Receptor TAS2R14 by Chemical Modification of Cognate Agonists. Chem.
751
Biol. Drug Des. 2016, 88, 66-75.
752
107.
Levit, A.; Nowak, S.; Peters, M.; Wiener, A.; Meyerhof, W.; Behrens, M.; Niv, M. Y.
753
The bitter pill: clinical drugs that activate the human bitter taste receptor TAS2R14.
754
FASEB J. 2014, 28, 1181-1197.
32 ACS Paragon Plus Environment
Page 33 of 41
755 756
Journal of Agricultural and Food Chemistry
108.
Dettner, K.; Liepert, C. Chemical mimicry and camouflage. Annu. Rev. Entomol. 1994, 39, 129-154.
757
33 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 34 of 41
758
Figure captions
759
Figure 1. The contribution of differently tuned bitter taste receptors to overall bitter
760
perception. More than 100 bitter substances were screened for their activation of 25 human
761
TAS2Rs. The fraction of substances detected by receptors with broad, intermediate, narrow,
762
and chemical group-specific (Group spec.) detection spectra is indicated. The human TAS2Rs
763
belonging to the four groups are indicated.
764
Figure 2. The binding pocket of human TAS2R10. Residues that were demonstrated to be
765
critical for agonist activation in the binding pocket of the human bitter taste receptor
766
TAS2R10 are shown. Three residues with pronounced agonist selectivity are depicted as bold
767
stick representations. Other residues contributing to general agonist activation are indicated as
768
think sticks. The center sphere indicates that sufficient space to accommodate large agonists is
769
available in the binding pocket.
770
Figure 3. Schematic of a TAS2R embedded in the plasma membrane with bound agonist. The
771
seven transmembrane domains connected by 3 extracellular and 3 intracellular loops of a
772
TAS2R are indicated by cylinders and connecting lines. The amino terminus points to the
773
extracellular site, the carboxy terminus (not shown) is located at the intracellular site. The
774
approximate ligand binding site in the upper third of the transmembrane domain helices is
775
indicated by a strychnine molecule (red).
776
Figure 4. Bitter taste receptor gene repertoires and tuning breadths. The bitter taste receptor
777
repertoires of seven species which have been functionally characterized are shown. The
778
putatively functional receptors are indicated by black squares and the adjacent number. The
779
tuning widths of the receptors are indicated by the sizes of the spheres and the fraction of test
780
compounds that activated the corresponding receptor is provided in percent. The receptor
781
symbols are given. The information for turkey, chicken, zebra finch, and Western clawed frog 34 ACS Paragon Plus Environment
Page 35 of 41
Journal of Agricultural and Food Chemistry
782
were derived from;66 the information for cat were derived from;67 the mouse data were taken
783
from;69 the information for human were compiled from.8,69
784
Figure 5. Chemical structures and sources of bitter substances are highly diverse. The
785
diversity of bitter compound structures range from small inorganic salts to complex organic
786
substances. The compounds originate from biological to abiotic sources and processes related
787
to food production.
788
35 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 36 of 41
Figure 1
36 ACS Paragon Plus Environment
Page 37 of 41
Journal of Agricultural and Food Chemistry
Figure 2
37 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 38 of 41
Figure 3
38 ACS Paragon Plus Environment
Page 39 of 41
Journal of Agricultural and Food Chemistry
Figure 4
39 ACS Paragon Plus Environment
Journal of Agricultural and Food Chemistry
Page 40 of 41
Figure 5
40 ACS Paragon Plus Environment
Page 41 of 41
Journal of Agricultural and Food Chemistry
For table of contents only
41 ACS Paragon Plus Environment