bitter taste genetics: ptc and other stories lynn jorde and steve wooding department of human...

Bitter Taste Genetics:Bitter Taste Genetics: PTC and Other Stories PTC and Other Stories

Lynn Jorde and Steve WoodingLynn Jorde and Steve WoodingDepartment of Human GeneticsDepartment of Human Genetics

University of Utah School of MedicineUniversity of Utah School of Medicine

OverviewOverview

PTC: evidence for balancing selectionPTC: evidence for balancing selection

PTC in chimps and humansPTC in chimps and humans

The other bitter taste receptor genesThe other bitter taste receptor genes

Michael Bamshad, MD Stephen Wooding, PhD

Molecules involved in taste perceptionMolecules involved in taste perceptionSourSour Ion channelsIon channels

SaltySalty Ion channelsIon channels

SweetSweet TAS1R family of G-TAS1R family of G-protein coupled protein coupled receptorsreceptors

UmamiUmami TAS1R family of G-TAS1R family of G-protein coupled protein coupled receptorsreceptors

BitterBitter TAS2R family (25 TAS2R family (25 functional members)functional members)

Sweet and Umami (T1R)Sweet and Umami (T1R)

T1R1 T1R2 T1R3

11

22 33 High-affinity sugar receptorHigh-affinity sugar receptor

33 High-affinity umami receptorHigh-affinity umami receptor

33 Low-affinity umami receptorLow-affinity umami receptor

Cell types Cell types observed observed in vivoin vivo

The Bitter DraughtAdriaen Brouwer, 1635

Many plants contain toxinsMany plants contain toxins

Ricin

AbrinStrychnine

Digitoxin

Many plant toxins taste bitterMany plant toxins taste bitter

Quinine Nicotine

PapaverineSalicin

Biomedical relevance of bitter tasteBiomedical relevance of bitter taste

Bitter-tasteBitter-taste sensitivity

SmokingSmoking

Body Mass IndexBody Mass Index

Diet ChoiceDiet Choice

CancerCancer

Bitter-taste (TAS2R) ReceptorsBitter-taste (TAS2R) Receptors

Bitter-taste (TAS2R) ReceptorsBitter-taste (TAS2R) Receptors

TAS2R

G

x

Ca+

Ca+Ca+

Ca+

Bitter-taste receptors bind plant toxinsBitter-taste receptors bind plant toxins

TAS2R10TAS2R10 strychninestrychnine

TAS2R14TAS2R14 -thujones-thujones

TAS2R16TAS2R16 salicin, salicin, cyanogenic compoundscyanogenic compounds

Bufe et al., Nat. Gen., 2002; Behrens et al., BBRC, 2004

L’absintheEdgar Degas, 1876

Phenylthiocarbamide (PTC)Phenylthiocarbamide (PTC)

SyntheticSynthetic• Bitter or tastelessBitter or tasteless

Fox 1932Fox 1932• Variable sensitivityVariable sensitivity• Similar in structure to plant compounds Similar in structure to plant compounds

Blakeslee 1932Blakeslee 1932• Taste blindness is Mendelian recessiveTaste blindness is Mendelian recessive

Fisher et al., 1939Fisher et al., 1939• Variable sensitivity in chimpVariable sensitivity in chimp

Thousands of individuals have been typed for Thousands of individuals have been typed for PTC taster statusPTC taster status

PTCPTC Gene Gene

Drayna Drayna et al., et al., 20032003• Genome-wide linkage analysisGenome-wide linkage analysis

• Utah CEPH families• PTC gene localized to chromosome 7 and subsequently

cloned

How have natural selection and population historyaffected the PTC gene?

• PTCPTC Gene ( Gene (TAS2R38TAS2R38))• Haplotypes strongly associated with phenotype• Accounted for ~50-85% of phenotypic variance

PTCPTC resequencing in humans resequencing in humans

Sequenced entire coding region (1,002 bp)Sequenced entire coding region (1,002 bp)

• 174 individuals (348 chromosomes)174 individuals (348 chromosomes)• 40 African• 69 Asian• 55 European• 10 Native American

- Not selected for PTC sensitivity

- “Anthropological” sampling

Wooding et al., 2004, Am. J. Hum. Genet. 74: 637-46

Diversity in Diversity in PTCPTC

6 nucleotide substitutions in humans6 nucleotide substitutions in humans

Diversity in Diversity in PTCPTC

6 nucleotide substitutions6 nucleotide substitutions• 7 haplotypes7 haplotypes

PAV AVI

Diversity in Diversity in PTCPTC

TasterAllele

NontasterAllele

6 nucleotide substitutions6 nucleotide substitutions• 7 haplotypes7 haplotypes

PAV: AVI:

Allele frequenciesAllele frequencies

African Asian EuropeanNative

American

T

t

FST = 0.05

Has selection been acting?Has selection been acting?

Neutrality Not Rejected, p > 0.06

Has selection been acting?Has selection been acting?

Wooding et al., Am. J. Hum. Genet., 2004

Neutrality rejected

Exceeds 99.5% of D values reported for Exceeds 99.5% of D values reported for >1,600 genes>1,600 genes

How has selection acted?How has selection acted?

Nontaster alleles are not

premature stop codons or frameshifts

HYPOTHESIS: Nontaster allele is a functional receptor that binds ligands other than PTC.

Tt +

What happened in chimps?What happened in chimps?

TAS2R38 Resequencing in 37 Wild-born Chimpanzees

AGG StartATG Start

Wooding et al., 2006, Wooding et al., 2006, Nature Nature 440: 930-4440: 930-4

Hypothesized PTC Tasting in Chimpanzees

“Taster” “Non-taster”

Wooding et al., 2006, Wooding et al., 2006, Nature Nature 440: 930-4440: 930-4

How does AGG make a nontaster allele?How does AGG make a nontaster allele?

ATG AGG

Full-length Protein: PTC response Truncated Protein: no PTC response

Phenotyping ChimpanzeesPhenotyping Chimpanzees

Is AGG the nontaster allele in chimps?Is AGG the nontaster allele in chimps? Genotype-phenotype association analysisGenotype-phenotype association analysis

- 40 chimpanzee subjects40 chimpanzee subjects- Genotyped

- Phenotyped- Apples soaked in H2O or 4.0 mM PTC

- Response on a 1-5 scale (1 = Readily Accepted; 5 = Strongly Rejected)

- Fisher’s Exact Test (2x2)Fisher’s Exact Test (2x2)

Taster Nontaster

Taster

Nontaster

23 1

7 8

Expected

Observed

Rejected no association (p < 10-3)AGG/AGG “broken start” chimps eat PTC apples

Humans versus ChimpsHumans versus Chimps

HumansHumans ChimpsChimps

T, t allelesT, t alleles

50:50 frequencies50:50 frequencies

3 amino acid changes3 amino acid changes

T, t allelesT, t alleles

50:50 frequencies50:50 frequencies

One start codon changeOne start codon change

Same gene, same phenotype, same alleleSame gene, same phenotype, same allelefrequencies… different mechanism.frequencies… different mechanism.

Fisher’s HypothesisFisher’s Hypothesis

“Taster” “Non-Taster”

Hu Ch Hu Ch

Ancestral Gene

Chimp-HumanDivergence

Human Chimpanzee

T t T t

Ancestral Gene

Resequencing all 25 Resequencing all 25 TAS2RTAS2R genes in 55 humans genes in 55 humans

Do all TAS2Rs have two clusters (i.e., T and t)?Do all TAS2Rs have two clusters (i.e., T and t)?• No – some have very low levels of variation.No – some have very low levels of variation.

Are all TAS2R alleles found at similar frequencies across Are all TAS2R alleles found at similar frequencies across populations?populations?• No – Some have dramatically different frequenciesNo – Some have dramatically different frequencies• On average populations differ more at TAS2Rs than at other genesOn average populations differ more at TAS2Rs than at other genes

TAS2R49

Kim et al., 2005, Kim et al., 2005, Hum. Mutation Hum. Mutation 26: 199-20426: 199-204

FST = 0.33

Population Differentiation Implies Local Population Differentiation Implies Local AdaptationAdaptation

FST

PTC ObservedSimulated

TAS2Rs (p < 0.01)

Kim et al., 2005, Kim et al., 2005, Hum. Mutation Hum. Mutation 26: 199-20426: 199-204

TAS2R genes are involved in the TAS2R genes are involved in the perception of many substancesperception of many substances

AlcoholAlcohol• Subjects with PTC taster haplotype experience more Subjects with PTC taster haplotype experience more

bitterness and have lower alcohol intake bitterness and have lower alcohol intake (Duffy et al., 2004, (Duffy et al., 2004, Alcohol Clin. Exp. Res. Alcohol Clin. Exp. Res. 28: 1629-37)28: 1629-37)

• Variant in Variant in TAS2R16TAS2R16 is strongly associated with risk of is strongly associated with risk of alcohol dependence alcohol dependence (Hinrichs et al., 2006, (Hinrichs et al., 2006, Am. J. Hum. Genet. Am. J. Hum. Genet. 78: 103-11)78: 103-11)

Cigarette smokeCigarette smoke• Some evidence that PTC tasters are aversive to cigarette Some evidence that PTC tasters are aversive to cigarette

smoke and less likely to become smokers smoke and less likely to become smokers (Enoch et al., 2001, (Enoch et al., 2001, Addict. Behav. Addict. Behav. 26: 399-404)26: 399-404)

Saccharin and acesulfame KSaccharin and acesulfame K• Activate TAS2R43 and TAS2R44 bitter taste receptorsActivate TAS2R43 and TAS2R44 bitter taste receptors

(Kuhn et al., 2004, (Kuhn et al., 2004, J. Neurosci.J. Neurosci. 10: 10260-5) 10: 10260-5)

SummarySummary

Polymorphism in the human Polymorphism in the human PTCPTC gene is gene is maintained by balancing natural selectionmaintained by balancing natural selection

Humans and chimps both have taster and Humans and chimps both have taster and non-taster alleles, but these alleles arose non-taster alleles, but these alleles arose independentlyindependently

Substantial inter-population differentiation Substantial inter-population differentiation of of TAS2RTAS2R genes exists, consistent with genes exists, consistent with local adaptation to environmental toxinslocal adaptation to environmental toxins

University of Utah: Mike Bamshad, Steve Wooding, Mike Bamshad, Steve Wooding, Diane Dunn, Bob Weiss, Mike Howard Diane Dunn, Bob Weiss, Mike Howard

NIH: Dennis Drayna, U. KimDennis Drayna, U. Kim

Potsdam: Bernd Bufe, Wolfgang Meyerhof Bernd Bufe, Wolfgang Meyerhof

University of Arizona: Anne StoneAnne Stone

University of Utah: Mike Bamshad, Steve Wooding, Mike Bamshad, Steve Wooding, Diane Dunn, Bob Weiss, Mike Howard Diane Dunn, Bob Weiss, Mike Howard

NIH: Dennis Drayna, U. KimDennis Drayna, U. Kim

Potsdam: Bernd Bufe, Wolfgang Meyerhof Bernd Bufe, Wolfgang Meyerhof

University of Arizona: Anne StoneAnne Stone

AcknowledgmentsAcknowledgments

How has selection acted?How has selection acted?

How does AGG make a nontaster allele?How does AGG make a nontaster allele?

ATG AGG

AUG AGG

Met Met Met

Transcription

Translation

TAC TCC

DNA

RNA

Protein

Has selection been acting?Has selection been acting?

Tajima’s D statisticTajima’s D statistic• Compares SCompares S

S = Number of nucleotide substitutions

Has selection been acting?Has selection been acting?

Tajima’s D statisticTajima’s D statistic• Compares S and Compares S and

= Mean pairwise nucleotide difference

Has selection been acting?Has selection been acting?

Tajima’s D statisticTajima’s D statistic• Compares S and Compares S and • Affected by “shape” of haplotype networkAffected by “shape” of haplotype network

Has selection been acting?Has selection been acting?

Tajima’s D statisticTajima’s D statistic• Compares S and Compares S and • Affected by “shape” of haplotype networkAffected by “shape” of haplotype network

S = 6 S = 6

Has selection been acting?Has selection been acting?

Tajima’s D statisticTajima’s D statistic• Compares S and Compares S and • Affected by “shape” of haplotype networkAffected by “shape” of haplotype network

S = 6 S = 6 = 0.1 = 3.0

Has selection been acting?Has selection been acting?

Tajima’s D statisticTajima’s D statistic• Compares S and Compares S and • Affected by “shape” of haplotype networkAffected by “shape” of haplotype network

S = 6 S = 6 = 0.1 = 3.0

D = -2.0 D = 3.6

Has selection been acting?Has selection been acting?

Tajima’s D statisticTajima’s D statistic• Compares S and Compares S and • Affected by “shape” of haplotype networkAffected by “shape” of haplotype network

Positive selection Balancing selectionor

Local adaptation

Human Variation

Habitats

Genes and dietGenes and diet

GeneGene PhenotypePhenotype

HemochromatosisHemochromatosis Iron absorptionIron absorption

AngiotensinogenAngiotensinogen Sodium retentionSodium retention

LactaseLactase Lactose toleranceLactose tolerance

Alcohol dehydrogenaseAlcohol dehydrogenase Ethanol metabolismEthanol metabolism

Environmentally Responsive GenesEnvironmentally Responsive Genes

Genes controlling the active interface betweenthe human body and its environment

Genes involved in:

Sensory perception

Immune system

Toxin metabolism

Bitter-taste sensitivity