1. Using a Single Nucleotide Polymorphism to Predict Bitter-Tasting Ability
Can you Taste PTC ?

2. Important concepts drug efficacy
Science evolves from past discoveries.
Modern biological research merges genetics, biochemistry, comparative studies and bioinformatics.
Receptors: Genetic differences in taste and smell
drug efficacy

3. Taste in Mammals Mammals can distinguish only five basic tastes Sweet
Sour
Bitter
Salty
Umami (the taste of monosodium gluatmate)

4. Taste in Mammals Taste perception is a two-step process
1st…A taste molecule binds to a specific receptor on the surface of a taste cell
Question - WHAT IS A RECEPTOR??
Question - WHAT DETERMINES THE STUCTURE OF A RECEPTOR
2nd …The taste cell generates a nervous impulse, which is interpreted by the brain

6. An Example: Taste in Mammals
Stimulation of “sweet cells” generates a perception of sweetness in the brain
Taste sensation is ultimately determined by the wiring of a taste cell to the cortex in the brain
If you have a sweet cell
But it expresses a “bitter taste receptor”
Bitter molecule will be perceived as being sweet!

7. Taste in Mammals
Taste recognition is mediated by specialized taste cells that communicate with several brain regions through direct connections to sensory neurons

8. Taste in Mammals
While there are only 5 tastes there are thousand more olfactory (smell) receptors (OR)
Smell is like taste—a receptor – a protein that binds to a molecule that we smell.
Similar also to how many drugs work (the drug binds to a cell protein—or receptor)
All are coded by specific genes

9. A Serendipitous Observation
The genetic basis of taste first observed by accident in 1930’s
PTC = phenylthiocarbamide
Prepared by Arthur Fox at Du Pont Company in late 1920s
Lab partner C.R. Noller complained of bitter taste but Fox felt no taste

10. Albert Blakeslee with Carnegie Department of Genetics, Cold Spring Harbor, New York, 1933
Followed up by Albert Blakeslee at Carnegie Department of Genetics showed that inability to taste is recessive.
Published in 1932

11. Albert Blakeslee, AAAS Convention, 1938

12. Inheritance pattern

13. Molecular Genetics of PTC Tasting
Gene identified in 2003 by Dennis Drayna
TAS2R38 gene
Polymorphism associated with PTC tasting
SNP--Nucleotide position 145
Taster = C Nontaster = G
Change in Amino acid 49 …. (proline)  (alanine)

14. Analysis of the Trait Isolate DNA from human cheek cells
Amplify a region of TAS2R38 gene by PCR
Primers used in the experiment:
CCTTCGTTTTCTTGGTGAATTTTTGGGATGTAGTGAAGAGGCGG
AGGTTGGCTTGGTTTGCAATCATC
Then cut with restriction enzyme (HaeIII)
RFLP-Restriction Fragment Length Polymorphism

15. Analysis of the Trait

16. Analysis of the Trait
How does the Hae III enzyme discriminate between the C-G polymorphism in the TAS2R38 gene.
HaeIII cuts at the sequence GGCC
This is at the position of the gene
The nontaster has a GGGC and won’t cut

17. Analysis by eletrophoresis

18. 2% Agarose Gel ElectrophoresisTT Tt tt
Marker U C U C U C Marker
221 bp
177 bp
44 bp

19. More Complication: More than 1 PTC Haplotypes
Postition
Taster
Nontaster
145
C (proline)
G (alanine)
785
C (alanine)
T (valine)
886
G (valine)
A (isoleucine)

20. What is the relationship between this trait and our ancestors?
What is the normal state?
To taste or to not taste?

21. Multiple Sequence Alignment

22. Advantage: Taste or not to taste?

23. Compare primer to sequence:
CCTTCGTTTTCTTGGTGAATTTTTGGGATGTAGTGAAGAGGCGG
primer anneal
TTTTTGGGATGTAGTGAAGAGGCGG
Taster TAGTGAAGAGGCAGCCACTG
Nontaster TAGTGAAGAGGCAGGCACTG

24. Compare primer to sequence:
CCTTCGTTTTCTTGGTGAATTTTTGGGATGTAGTGAAGAGGCGG
primer anneal
TTTTTGGGATGTAGTGAAGAGGCGG
3’ AAAAACCCTACATCACTTCTCCGTC
Taster ’ TAGTGAAGAGGCAGCCACTG
Nontaster TAGTGAAGAGGCAGGCACTG

25. Compare primer to sequence:
CCTTCGTTTTCTTGGTGAATTTTTGGGATGTAGTGAAGAGGCGG
primer anneal
TTTTTGGGATGTAGTGAAGAGGCGGCCACTG………..
3’AAAAACCCTA CATCACTTCTCCGTC
Taster TAGTGAAGAGGCAGCCACTG
Nontaster TAGTGAAGAGGCAGGCACTG

26. Evolution of ..
Many people are nontasters…more than what is expected if bitter taste was the ONLY trait under natural selection SO…. Is there some factor that makes this a positive outcome to balance out the negative effect of not tasting bitter? Is there an advantage to being a heterozygote (like sickle cell anemia)? Maybe….Maybe the NONTASTING form allow for individuals to taste another type of bitter molecule and so these people may know to avoid potentially toxic compounds.

27. OR Evolution Mice: 20% of ORs are inactive
Primates: 30-40% of ORs are inactive
Humans: 60% of ORs are inactive
Human-chimp comparisons:
OR genes are diverging quickly
OR genes are under natural selection