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Lecture 22 : Taste II. 11/18/09. Today or Monday. Final project: 1 pg of background. Each taste modality relies on different receptors. pH sensing. T1R2+T1R3. Cats don’t taste sweet. Search for T1R2 and T1R3. Screen cat BAC library Sequence positive clones. Find complete T1R3 gene.

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today or monday
Today or Monday
  • Final project:
    • 1 pg of background
search for t1r2 and t1r3
Search for T1R2 and T1R3
  • Screen cat BAC library
  • Sequence positive clones
slide13

Human

Mouse

Dog

Cow

Opossum

Chicken

t2r genes in mouse and human
T2R genes in mouse and human
  • Mouse
    • 35 functional 6 pseudogenes
  • Humans
    • 25 functional 11 pseudogenes
  • Other primates
slide18

Human

OWM

NWM

Prosimians

Mouse

X marks pseudogenes

Amany : one

Human:mouse

Bone : many

Human:mouse

Cone : one

Human:mouse

human variation in pseudogenes
Human variation in pseudogenes

T2R62

T2R2

Functional

Hosa - Homo sapiens Hyag - Hylobates agilis gibbon

Patr - Pan troglodytes chimp Mamu - Macaca mulatta rhesus

Gogo - Gorilla gorilla Caja - Callithrix jacchus marmoset

Popy - Pongo pygmaeus orangutan Ceap - Cebus apella capuchin

convergence of pseudogenes
Convergence of pseudogenes

T2R64

T2R55

Hosa - Homo sapiens Hyag - Hylobates agilis gibbon

Patr - Pan troglodytes chimp Mamu - Macaca mulatta rhesus

Gogo - Gorilla gorilla Caja - Callithrix jacchus marmoset

Popy - Pongo pygmaeus orangutan Ceap - Cebus apella capuchin

slide23
PTC
  • 75% of humans taste as bitter
    • Discovered in 1930’s
  • “Single” gene trait
  • Originally thought was Mendelian recessive
    • AA, Aa = sensitive
    • aa = not sensitive
  • Probably 2 loci
give people increasing concentrations of ptc till they tasted bitter
Give people increasing concentrations of PTC till they tasted bitter

x2

x2

x2

x2

x2

x2

x2

1 2 3 4 5 6 7

x2

x2

x2

x2

x2

x2

8 9 10 11 12 13 14

1M

Increasing concentration

quantitative trait locus qtl study
Quantitative trait locus (QTL) study
  • Link phenotype (sensitivity to PTC) to genotype in Utah pedigree
  • Found link to 4 Mb region on chromosome 7
    • Are 9 T2R and 7 OR genes in this region
  • Finer scale
    • Genotyped 50 more SNPs in this region
    • Added broader diversity of taster and non-taster individuals
tas2r38 variations
Tas2R38 variations

A V

V I

262 296

P

A

49

European EAsian

P A V 49 31 Taster

A A V 3

A V I 47 69 Non-taster

phenotype is determined by cup at which tasted ptc
Phenotype is determined by cup # at which tasted PTC

x2

x2

x2

x2

x2

x2

x2

1 2 3 4 5 6 7

x2

x2

x2

x2

x2

x2

8 9 10 11 12 13 14

1M

Increasing concentration

genotype vs phenotype
Genotype vs phenotype

Nontaster

Taster

human origin of nontaster
Human origin of nontaster

Human PAV /AAV/ AVI

Chimp PAV

Gorilla PAV

Orangutan PAV

Macaque PAV

bitter taste transduction
Bitter taste transduction

Sheridan Nature Biotech 2004

sour and salty
Sour and salty
  • Ionotropic channels
    • Salty - Na+
    • Sour - H+
salt detection likely a channel
Salt detection likely a channel
  • Membrane responds to Na+ gradient
capsicum
Capsicum

Capsaicin detected by vanilloid receptor ??

approach
Approach

30,000 genes

Screen mouse genome for genes containing TM

Screen EST libraries for genes which are not common

5 genes

884

PCR genes in papillae vs tongue

In situ hybridization to identify ones uniquely expressed

98

approach1
Approach
  • Assume TRP channels are important in taste
    • TRPM5 known to be channel in sweet, umami and bitter
  • Use in situ hybridization
    • Screen taste papillae for all 33 TRP channels
    • TrpM - 40%
    • PKD1L - 20% of circumvallate
    • No other Trp’s label
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