Professor Mark A. Tanouye April 16, 2000
1
LEARNING AND MEMORY IN ANIMALS:LEARNING MUTANTS,FORWARD GENETICS
Reading: p.189-198
NATURE VS NURTURENATURE Biological - usually called "genetic", but
actually more complicated, ill-defined.Component of intelligence that is inherited.
NURTURE Social - usually considered family background,education, socio-economic influences, ill-defined
IMPLICATIONSSocial programsHead StartSchool curriculaChild-rearing practices
"FORWARD" GENETICS(mutational analysis, functional analysis)
FUNCTION GENE PROTEIN(learning) (loss-of-function)
(stupidity)
clonemutation
LEARNING TESTMUTANT SCREEN
MAP GENECLONE GENE(positional cloning)PROTEIN SEQUENCE
IDENTIFY MUTANTMUTANT STRAIN
MAKING AND CHARACTERIZING MUTANTS
MUTAGENESIS
SCREEN
(overview)
NUMBER OF GENES ? complementation tests
damage DNA of one gene(single gene mutation)(somatic - cancer)(germline - pass on to progeny)
identify mutants with learning test or screen(dominant mutations, recessive mutations)
FUNCTIONAL PATHWAYS ?double mutant analysis
MAPPING
a) linkage (which chromosome) ?b) distance from other genesc) linear order of genes "Genetic model" organisms
bacteria, yeast, Drosophila, C. elegans, zebrafish, Arabidopsis, mouse, human recombination
LEARNING THAT ODORMEANS ELECTRIC SHOCK
(classical conditioning)
ODOR+
SHOCK
training
association
AVOIDODORALONE
testing
Odor "symbolizes"electric shock
PsychologyShock = unconditioned stimulus (US)Odor = conditioned stimulus (CS)
DROSOPHILA LEARNING APPARATUS
Professor Mark A. Tanouye April 16, 2000
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RESULTS OF LEARNING TESTSLearning in normal flies dunce flies: no learning
(all go toward light)
MUTATIONS(radiation)
MUTATIONS(radiation)
IONIZING RADIATION Ultraviolet X-rays Gamma rays
IONIZING RADIATION Ultraviolet X-rays Gamma rays
X
xxx
***
DELETIONDELETION
TRANSLOCATIONTRANSLOCATION
MUTATIONSchemical
MUTATIONSchemical
Mutagen/Carcinogenethylmethanesulfonateethylnitrosourea(alkylating drugs)
Mutagen/Carcinogenethylmethanesulfonateethylnitrosourea(alkylating drugs)
REPLICATION
G*
A
GENE
normal
mutation
AMINO ACID SUBSTITUTION GGT AGT gly ser
STOP CODON SUBSTITUTION TGG TAG trp stop
AMINO ACID SUBSTITUTION GGT AGT gly ser
STOP CODON SUBSTITUTION TGG TAG trp stop
PATHWAYS DEFINEDBY DROSOPHILA
LEARNING GENESAVOID ODOR
INITIALLEARNING
linotte
SHORT-TERMMEMORY
duncerutabaga
MEDIUM-TERMMEMORY
amnesiac
ANESTHESIA-RESISTANTMEMORY radish
LONG-TERMMEMORYdCREB2
DROSOPHILACHROMOSOMEMAPS
1st (X) 2nd 3rd 4th
3 major chromosomes (1,2,3)1 minor chromosome (4th)
180Mb13,600 genes
1889–1938, b. Schuyler Falls, N.Y., grad.Columbia (B.S., 1912; Ph.D., 1916)discoverer of white mutant; most mutantspolytene chromosome maps; free love
Professor Mark A. Tanouye April 16, 2000
3
which chromosome is the dnc mutation on?
CHROMOSOMAL LINKAGE
Possibilities for two mutations (genes)(dnc + marker mutation)
Linkage(+ recombination)
Linkage(+ recombination)
independent assortment (common)(uncommon, "closely-linked")
No linkage(different chromosomes)
No linkage(different chromosomes)
Linkage(no recombination)
Linkage(no recombination)
dnc
1(X)
3Sb
dnc
1(X)
rb B
dnc
1(X)
which chromosome is the dnc mutation on?
CHROMOSOMAL LINKAGE
dnc
1(X)
B
FM, B TM, Sb
Sb
3
Cy
2SM, Cy
dnc/Y; 2/2; 3/3 (males) mated to FM,B/+; SM,Cy/+; TM,Sb (females)
mated to male: (dnc/Y; 2/2; 3/3)
(female)
Look for missing class of animals for linkage
dnc/dnc; Cy/+; Sb/+ (present) dnc/dnc flies w/ B (absent)
GENETIC MAPPING(Recombination mapping - distance)
dncdnc
X
(female) (male)
ww
dnc = stupid (exptl)w = white eyes (marker)
(phenotypes?)
dnc
w
+
+(female double heterozygote)
Xdnc
w
+
+Xdnc
w
+
+
dnc
+
w
+
Xdnc+
w +
dnc+
w +
RECOMBINANTSdistance (eg. 2% = 2 map units)
GENETIC MAPPING
dncdnc
cv = missing wing vein(marker)
(Recombination mapping - direction)
dnc
w
+
+
dnc
w
+
+?
2 map units 2 map units
dnc
w
+
+ cv
+
THREE FACTOR CROSS
w-dnc 2 map unitsdnc-cv 3 map unitsw-cv 5 map units(if w & cv are known, unique solution)
POSITIONAL CLONING
w dnc cvmap
position
map intervalcontaining dnc
overlappingDNA clones
(chromosome walk)
25kb
1. clone entire map interval2. identify all genes (transcription units)3. transgene rescue (final identification)
DROSOPHILALEARNINGMUTANTS
dunce cAMP phosphodiesterasenamed after John Duns Scotus, 13thCopponent of classical learning
rutabaga adenyl cyclase turnip unknownradish unknown cabbage unknown
The vegetable series: because they're as dumb as vegetables
CaMKII Ca/calmodulin-dependent protein kinase IIdCREB2 cAMP response element binding protein (transcription factor)
amnesiac neuropeptidesimilar to mammalian pituitary adenyl cyclase activating peptide (PACAP)
linotte novel productnamed fr French expression tete de linotte having the same idiomaticmeaning as "bird-brain"
latheo unknownGreek for "forgetful" related to the Lethe River of Hades, a drink and youforget your former life
Professor Mark A. Tanouye April 16, 2000
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dCREB2 duncerutabaga
PACAP (amnesiac)