Gustducin
Encyclopedia
Gustducin is a G protein
associated with basic taste and the gustatory system
. Due to its relatively recent discovery and isolation, not all is known about its nature and its associated pathways. It is known that it plays a large role in the transduction of bitter, sweet and umami stimuli and that its pathways (especially for detecting bitter stimuli) are many and diverse. Perhaps the most intriguing feature of gustducin is its similarity to transducin
. These two G proteins have been shown to be structurally and functionally similar, leading researchers to believe that the sense of taste evolved in a similar fashion to the sense of sight
.
Gustducin is a heterotrimeric protein composed of the products of the GNAT3
(α-subunit), GNB1
(β-subunit) and GNG13
(γ-subunit) genes.
primers were synthesized and mixed with a taste tissue [cDNA] library. The DNA
products were amplified by the polymerase chain reaction
method and eight positive clones were shown to encode the α subunits of G-proteins, which interact with G-protein-coupled receptors. Of these eight units, two had previously been shown to encode rod
and cone
α-transducin
. The eighth clone, α-gustducin, was unique to the gustatory tissue.
Phosphodiesterase
, which leads to the breakdown of cGMP. Similarly, α-gustducin binds the inhibitory units subunits of taste cell camp PDE which also causes a decrease in cAMP levels. Also, the terminal 38 amino acids of α-gustducin and α-transducin are identical. This suggests that gustducin and interact with opsin
and opsin-like G-coupled receptors. Conversely, this also suggests that transducin can interact with taste receptors. The structural similarities between gustducin and transducin are so great that comparison with transducin were used to propose a model of gustducin's role and functionality in taste transduction.
Due to its structural similarity to transducin, gustducin was predicted to activate a phosphodiesterase
(PDE). Phosphodieterases were found in taste tissues and their activation was tested in vitro
with both gustducin and transducin. This experiment revealed transducin and gustducin were both expressed in taste tissue (1:25 ratio) and that both G proteins are capable of activating retinal
PDE. Furthermore, when present with denatonium
and quinine, both G proteins can activate taste specific PDEs. This indicated that both gustducin and transducin are important in the signal transduction of denatonium and quinine.
Finally, Margolskee’s group investigated the role of gustducin in bitter taste reception by using “knock-out” mice lacking the gene for α-gustducin. A taste test with knock-out and control mice revealed that the knock-out mice showed no preference between bitter and regular food in most cases. When the α-gustducin gene was re-inserted into the knock-out mice, the original taste ability returned. However, the loss of the α-gustducin gene did not completely remove the ability of the knock-out mice to taste bitter food. This indicates that α-gustducin is not the only mechanism for tasting bitter food. It was thought, though unconfirmed, that an alternative mechanism of bitter taste detection could be associated with the βγ subunit of gustducin. This theory was validated when it was discovered that both peripheral and central gustatory neurons typically respond to more than one type of taste stimulant, although a neuron typically would favor one specific stimulant over others. This suggests that, while many neurons favor bitter taste stimuli, neurons that favor other stimuli such as sweet and umami may be capable of detecting bitter stimuli in the absence of bitter stimulant receptors, as with the knock-out mice.
results from βγ-gustducin. Although the following steps of the α-gustducin pathway are unconfirmed, it is suspected that decreased cNMPs may act on protein kinases which would regulate taste receptor cell ion channel activity. It is also possible that cNMP levels directly regulate the activity of cNMP-gated channels and cNMP-inhibited ion channels expressed in taste receptor cell. The βγ-gustducin pathway continues with the activation of IP3 receptors and the release of Ca2+ followed by neurotransmitter
release.
Bitter taste transduction models
Several models have been suggested for the mechanisms regarding the transduction of bitter taste signals.
It is thought that these five diverse mechanisms have developed as defense mechanisms. This would imply that many different poisonous or harmful bitter agents exist and these five mechanisms exist to prevent humans from eating or drinking them. It is also possible that some mechanisms can act as backups should a primary mechanism fail. One example of this could be quinine, which has been shown to both inhibit and activate PDE in bovine taste tissue.
of K+ channels, closing the channels, depolarizing the taste cell causing, voltage-gated Ca2+ channels to open and causing neurotransmitter release. The second pathway is a GPCR-Gq/Gβγ-IP3 pathway which is used with artificial sweeteners. Artificial sweeteners bind and activate GPCRs coupled to PLCβ2 by either α-Gq or Gβγ. The activated subunits activate PLCβ2 to generate IP3 and DAG. IP3 and DAG elicit Ca2+ release and cause cellular depolarization. In influx of Ca2+ trigger neurotransmitter release. While these two pathways coexist in the same TRCs, it is unclear how the receptors selectively mediate cAMP responses to sugars and IP3 responses to artificial sweeteners.
tastes) are mediated by receptors from the G protein-coupled receptor family. Mammalian bitter taste receptors (T2Rs) are encoded by a gene family of only a few dozen members. It is believed that bitter taste receptors evolved as a mechanism to avoid ingesting poisonous and harmful substances. If this is the case, one might expect different species to develop different bitter taste receptors based on dietary and geographical constraints. With the exception of T2R1 (which lies on chromosome
5) all human bitter taste receptor genes can be found clustered on chromosome 7 and chromosome 12. Analyzing the relationships between bitter taste receptor genes show that the genes on the same chromosome are more closely related to each other than genes on different chromosomes. Furthermore, the genes on chromosome 12 have higher sequence similarity than the genes found on chromosome 7. This indicated that these genes evolved via tandem gene duplications and that chromosome 12, as a result of its higher sequence similarity between its genes, went through these tandem duplications more recently than the genes on chromosome 7.
and gastrointestinal tract. His work suggests that gustducin is present in these areas as a defense mechanism. It is widely known that some drugs and toxins can cause harm and even be lethal if ingested. It has already been theorized that multiple bitter taste reception pathways exist to prevent harmful substances from being ingested, but a person can choose to ignore the taste of a substance. Ronzegurt suggests that the presence of gustducin in epithelial cells in the stomach and gastrointestinal tract are indicative of another line of defense against ingested toxins. Whereas taste cells in the mouth are designed to compel a person to spit out a toxin, these stomach cells may act to force a person to spit up the toxins in the form of vomit.
G protein
G proteins are a family of proteins involved in transmitting chemical signals outside the cell, and causing changes inside the cell. They communicate signals from many hormones, neurotransmitters, and other signaling factors. G protein-coupled receptors are transmembrane receptors...
associated with basic taste and the gustatory system
Gustatory system
The gustatory system is the sensory system for the sense of taste.- Importance :The gustatory system allows humans to distinguish between safe and harmful food. Bitter and sour foods we find unpleasant, while salty, sweet, and meaty tasting foods generally provide a pleasurable sensation...
. Due to its relatively recent discovery and isolation, not all is known about its nature and its associated pathways. It is known that it plays a large role in the transduction of bitter, sweet and umami stimuli and that its pathways (especially for detecting bitter stimuli) are many and diverse. Perhaps the most intriguing feature of gustducin is its similarity to transducin
Transducin
Transducin is a heterotrimeric G protein that is naturally expressed in vertebrate retina rods and cones .- Mechanism of action :...
. These two G proteins have been shown to be structurally and functionally similar, leading researchers to believe that the sense of taste evolved in a similar fashion to the sense of sight
Visual perception
Visual perception is the ability to interpret information and surroundings from the effects of visible light reaching the eye. The resulting perception is also known as eyesight, sight, or vision...
.
Gustducin is a heterotrimeric protein composed of the products of the GNAT3
GNAT3
Guanine nucleotide-binding protein G subunit alpha-3, also known as gustducin alpha-3 chain, is a protein that in humans is encoded by the GNAT3 gene....
(α-subunit), GNB1
GNB1
Guanine nucleotide-binding protein G/G/G subunit beta-1 is a protein that in humans is encoded by the GNB1 gene.-Further reading:...
(β-subunit) and GNG13
GNG13
Guanine nucleotide-binding protein G/G/G subunit gamma-13 is a protein that in humans is encoded by the GNG13 gene.-Further reading:...
(γ-subunit) genes.
Discovery
Gustducin was discovered by Margolskee when degenerate oligonucleotideOligonucleotide
An oligonucleotide is a short nucleic acid polymer, typically with fifty or fewer bases. Although they can be formed by bond cleavage of longer segments, they are now more commonly synthesized, in a sequence-specific manner, from individual nucleoside phosphoramidites...
primers were synthesized and mixed with a taste tissue [cDNA] library. The DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
products were amplified by the polymerase chain reaction
Polymerase chain reaction
The polymerase chain reaction is a scientific technique in molecular biology to amplify a single or a few copies of a piece of DNA across several orders of magnitude, generating thousands to millions of copies of a particular DNA sequence....
method and eight positive clones were shown to encode the α subunits of G-proteins, which interact with G-protein-coupled receptors. Of these eight units, two had previously been shown to encode rod
Rod cell
Rod cells, or rods, are photoreceptor cells in the retina of the eye that can function in less intense light than can the other type of visual photoreceptor, cone cells. Named for their cylindrical shape, rods are concentrated at the outer edges of the retina and are used in peripheral vision. On...
and cone
Cone cell
Cone cells, or cones, are photoreceptor cells in the retina of the eye that are responsible for color vision; they function best in relatively bright light, as opposed to rod cells that work better in dim light. If the retina is exposed to an intense visual stimulus, a negative afterimage will be...
α-transducin
Transducin
Transducin is a heterotrimeric G protein that is naturally expressed in vertebrate retina rods and cones .- Mechanism of action :...
. The eighth clone, α-gustducin, was unique to the gustatory tissue.
Comparisons with transducin
Upon analyzing the amino-acid sequence of α-gustducin, it was discovered that a-gustducins and a-transducins were closely related. α-gustducin's protein sequence gives it 80% identity to both rod and cone a-transducin. Despite the structural similarities, the two proteins have very different functionalities. This is not to say the two proteins do not have similar mechanism and capabilities. Transducin removes the inhibition from cGMPCyclic guanosine monophosphate
Cyclic guanosine monophosphate is a cyclic nucleotide derived from guanosine triphosphate . cGMP acts as a second messenger much like cyclic AMP...
Phosphodiesterase
Phosphodiesterase
A phosphodiesterase is any enzyme that breaks a phosphodiester bond. Usually, people speaking of phosphodiesterase are referring to cyclic nucleotide phosphodiesterases, which have great clinical significance and are described below...
, which leads to the breakdown of cGMP. Similarly, α-gustducin binds the inhibitory units subunits of taste cell camp PDE which also causes a decrease in cAMP levels. Also, the terminal 38 amino acids of α-gustducin and α-transducin are identical. This suggests that gustducin and interact with opsin
Opsin
Opsins are a group of light-sensitive 35–55 kDa membrane-bound G protein-coupled receptors of the retinylidene protein family found in photoreceptor cells of the retina. Five classical groups of opsins are involved in vision, mediating the conversion of a photon of light into an electrochemical...
and opsin-like G-coupled receptors. Conversely, this also suggests that transducin can interact with taste receptors. The structural similarities between gustducin and transducin are so great that comparison with transducin were used to propose a model of gustducin's role and functionality in taste transduction.
Evolution of the gustducin-mediated signaling model
While gustducin was known to be expressed in taste cells, studies with rats showed that gustducin was also present in a limited subset of cells lining the stomach and intestine. These cells appear to share several feature of taste receptor cells. Another study with humans brought to light two immunoreactive patterns for α-gustducin in human circumavallate and foliate taste cells: plasmalemmal and cytosolic. These two studies showed that gustducin is distributed through gustatory tissue and some gastric and intestinal tissue and gustducin is presented in either a cytoplasmic or apical pattern.Due to its structural similarity to transducin, gustducin was predicted to activate a phosphodiesterase
Phosphodiesterase
A phosphodiesterase is any enzyme that breaks a phosphodiester bond. Usually, people speaking of phosphodiesterase are referring to cyclic nucleotide phosphodiesterases, which have great clinical significance and are described below...
(PDE). Phosphodieterases were found in taste tissues and their activation was tested in vitro
In vitro
In vitro refers to studies in experimental biology that are conducted using components of an organism that have been isolated from their usual biological context in order to permit a more detailed or more convenient analysis than can be done with whole organisms. Colloquially, these experiments...
with both gustducin and transducin. This experiment revealed transducin and gustducin were both expressed in taste tissue (1:25 ratio) and that both G proteins are capable of activating retinal
Retinal
Retinal, also called retinaldehyde or vitamin A aldehyde, is one of the many forms of vitamin A . Retinal is a polyene chromophore, and bound to proteins called opsins, is the chemical basis of animal vision...
PDE. Furthermore, when present with denatonium
Denatonium
Denatonium, usually available as denatonium benzoate and as denatonium saccharide, is the bitterest chemical compound known; with bitterness thresholds of 0.05 ppm for the benzoate and 0.01 ppm for the saccharide.It was discovered in 1958 during research on local anesthetics by Macfarlan...
and quinine, both G proteins can activate taste specific PDEs. This indicated that both gustducin and transducin are important in the signal transduction of denatonium and quinine.
Finally, Margolskee’s group investigated the role of gustducin in bitter taste reception by using “knock-out” mice lacking the gene for α-gustducin. A taste test with knock-out and control mice revealed that the knock-out mice showed no preference between bitter and regular food in most cases. When the α-gustducin gene was re-inserted into the knock-out mice, the original taste ability returned. However, the loss of the α-gustducin gene did not completely remove the ability of the knock-out mice to taste bitter food. This indicates that α-gustducin is not the only mechanism for tasting bitter food. It was thought, though unconfirmed, that an alternative mechanism of bitter taste detection could be associated with the βγ subunit of gustducin. This theory was validated when it was discovered that both peripheral and central gustatory neurons typically respond to more than one type of taste stimulant, although a neuron typically would favor one specific stimulant over others. This suggests that, while many neurons favor bitter taste stimuli, neurons that favor other stimuli such as sweet and umami may be capable of detecting bitter stimuli in the absence of bitter stimulant receptors, as with the knock-out mice.
Gustducin and its second messengers
Until recently, the nature of gustducin and its second messengers was unclear. It was clear, however, that gustducin acted like transducin, and transduced intracellular signals. Spielman was one of the first to look at the speed of taste reception, utilizing the quenched-flow technique. When the taste cells were exposed to the bitter stimulants denatonium and sucrose octaacetate, the intracellular response was a transient increase of IP3 occurred within 50-100 millisecond of stimulation. This was not unexpected, as it was known that transducin was capable of sending signals within rod and cone cells at similar speeds. This indicated that IP3 was one of the second messengers used in bitter taste transduction. It was later discovered that cAMP also causes an influx of cations during bitter and some sweet taste transduction, leading to the conclusion that it also acted as a second messenger to gustducin.Bitter transduction
When bitter-stimulated T2R/TRB activate gustducin heterotrimers, gustducin acts to mediate two responses in taste receptor cells. A decrease in cNMPs is triggered by α-gustducin and a rise in IP3(Inositol trisphosphate)/DAGDag
-Places:*Dág , a village*Dąg, a village in Poland*DAG, the British National Rail station code for Dalgety Bay railway station, Scotland*DAG, the IATA code for Barstow-Daggett Airport, California, USA...
results from βγ-gustducin. Although the following steps of the α-gustducin pathway are unconfirmed, it is suspected that decreased cNMPs may act on protein kinases which would regulate taste receptor cell ion channel activity. It is also possible that cNMP levels directly regulate the activity of cNMP-gated channels and cNMP-inhibited ion channels expressed in taste receptor cell. The βγ-gustducin pathway continues with the activation of IP3 receptors and the release of Ca2+ followed by neurotransmitter
Neurotransmitter
Neurotransmitters are endogenous chemicals that transmit signals from a neuron to a target cell across a synapse. Neurotransmitters are packaged into synaptic vesicles clustered beneath the membrane on the presynaptic side of a synapse, and are released into the synaptic cleft, where they bind to...
release.
Bitter taste transduction models
Several models have been suggested for the mechanisms regarding the transduction of bitter taste signals.
- Channel blockage: Patch clamping experiments have shown that several bitter ions act directly on potassium channels, blocking them. This suggests that the potassium channels would be located in the apical region of the taste cells. While this theory seems valid, it has only been identified in mudpuppyMudpuppyMudpuppies or waterdogs are aquatic salamanders of the family Proteidae. Their name originates from the misconception that they make a dog-like barking sound. The range of the genus Necturus runs from southern central Canada, through the midwestern United States, east to North Carolina and south to...
taste cells. - Cell-surface receptors: Patch clamping experiments have shown evidence that bitter compounds such as denatoniumDenatoniumDenatonium, usually available as denatonium benzoate and as denatonium saccharide, is the bitterest chemical compound known; with bitterness thresholds of 0.05 ppm for the benzoate and 0.01 ppm for the saccharide.It was discovered in 1958 during research on local anesthetics by Macfarlan...
and sucrose octaacetate act directly on specific cell-surface receptors. - Direct activation of G proteins: Certain bitter stimulants such as quinineQuinineQuinine is a natural white crystalline alkaloid having antipyretic , antimalarial, analgesic , anti-inflammatory properties and a bitter taste. It is a stereoisomer of quinidine which, unlike quinine, is an anti-arrhythmic...
have been show to activate G proteins directly. While these mechanisms have been identified, the physiologic relevance of the mechanism has not yet been established. - PDE activation: Other bitter compounds, such as thioacetamide and propylthiouracil, have been shown to have stimulatory effects on PDEs. This mechanism has been recognized in bovine tongue epithelium contains fungiform papillae.
- PDE inhibition: Other bitter compounds have been shown to inhibit PDE. Bacitracin and hydrochloride have been show to inhibit PDE in bovine taste tissue
It is thought that these five diverse mechanisms have developed as defense mechanisms. This would imply that many different poisonous or harmful bitter agents exist and these five mechanisms exist to prevent humans from eating or drinking them. It is also possible that some mechanisms can act as backups should a primary mechanism fail. One example of this could be quinine, which has been shown to both inhibit and activate PDE in bovine taste tissue.
Sweet Transduction
There are currently two models proposed for sweet taste transduction. The first pathway is a GPCRGs-cAMP pathway. This pathway starts with sucrose and other sugars activating Gs through GPCR. The activated Gas activates adenylyl cyclase to generate cAMP. From this point, one of two pathways can be taken. cAMP may act directly to cause an influx of cations through camp gated channels or cAMP can activate protein kinase A, which cases phosphorylationPhosphorylation
Phosphorylation is the addition of a phosphate group to a protein or other organic molecule. Phosphorylation activates or deactivates many protein enzymes....
of K+ channels, closing the channels, depolarizing the taste cell causing, voltage-gated Ca2+ channels to open and causing neurotransmitter release. The second pathway is a GPCR-Gq/Gβγ-IP3 pathway which is used with artificial sweeteners. Artificial sweeteners bind and activate GPCRs coupled to PLCβ2 by either α-Gq or Gβγ. The activated subunits activate PLCβ2 to generate IP3 and DAG. IP3 and DAG elicit Ca2+ release and cause cellular depolarization. In influx of Ca2+ trigger neurotransmitter release. While these two pathways coexist in the same TRCs, it is unclear how the receptors selectively mediate cAMP responses to sugars and IP3 responses to artificial sweeteners.
Evolution of bitter taste receptors
Of the five basic tastes, three (sweet, bitter and umamiUmami
Umami , popularly referred to as savoriness, is one of the five basic tastes together with sweet, sour, bitter, and salty.-Etymology:Umami is a loanword from the Japanese meaning "pleasant savory taste". This particular writing was chosen by Professor Kikunae Ikeda from umai "delicious" and mi ...
tastes) are mediated by receptors from the G protein-coupled receptor family. Mammalian bitter taste receptors (T2Rs) are encoded by a gene family of only a few dozen members. It is believed that bitter taste receptors evolved as a mechanism to avoid ingesting poisonous and harmful substances. If this is the case, one might expect different species to develop different bitter taste receptors based on dietary and geographical constraints. With the exception of T2R1 (which lies on chromosome
Chromosome
A chromosome is an organized structure of DNA and protein found in cells. It is a single piece of coiled DNA containing many genes, regulatory elements and other nucleotide sequences. Chromosomes also contain DNA-bound proteins, which serve to package the DNA and control its functions.Chromosomes...
5) all human bitter taste receptor genes can be found clustered on chromosome 7 and chromosome 12. Analyzing the relationships between bitter taste receptor genes show that the genes on the same chromosome are more closely related to each other than genes on different chromosomes. Furthermore, the genes on chromosome 12 have higher sequence similarity than the genes found on chromosome 7. This indicated that these genes evolved via tandem gene duplications and that chromosome 12, as a result of its higher sequence similarity between its genes, went through these tandem duplications more recently than the genes on chromosome 7.
Gustducin in the stomach
Recent work by Enrique Ronzengurt has shed some light on the presence of gustducin in the stomachStomach
The stomach is a muscular, hollow, dilated part of the alimentary canal which functions as an important organ of the digestive tract in some animals, including vertebrates, echinoderms, insects , and molluscs. It is involved in the second phase of digestion, following mastication .The stomach is...
and gastrointestinal tract. His work suggests that gustducin is present in these areas as a defense mechanism. It is widely known that some drugs and toxins can cause harm and even be lethal if ingested. It has already been theorized that multiple bitter taste reception pathways exist to prevent harmful substances from being ingested, but a person can choose to ignore the taste of a substance. Ronzegurt suggests that the presence of gustducin in epithelial cells in the stomach and gastrointestinal tract are indicative of another line of defense against ingested toxins. Whereas taste cells in the mouth are designed to compel a person to spit out a toxin, these stomach cells may act to force a person to spit up the toxins in the form of vomit.