The Genetics of Caenorthabditis Elegans
Brenner, 1974
Lecture by Assaf Tal
Talk Outline
Background for Paper Concept of a Genetic Map Present Experimental System Present & Discuss Results Further Work
From Genes to Structure
“How do genes specify the structure of an organism?”
- Brenner, 1974
Sydney Brenner (1927 - )
Discoverer of frameshift mutations (1962)
Believed “classical” molecular biology was “solved”
Letter to Max Perutz (1963): “ ... Attempt to define the unitary steps of development in terms of genetic analysis … “
1963 Research Proposal: “The New Major problem in molecular biology is the genetics … of control mechanisms”
The Goal:
To study the connection between micro (genes) and macro (development) on a multi-cellular “simple” organism
A Genetic Map
Genetics Refresher
Gene:
Set of bases in DNA
Paradigm: 1 Gene per Protein
Allele: A particular Gene Realization
Homologous Chromosomes
Gene
Alleles(recessive/dominant)
Genotype: Aa
Phenotype: “what we see”
Wild Type: “common” phenotype
Mutant: “non-natural” phenotype
How to Make a Genetic Map
Step I Find out how many genes are responsible for each phenotype
( “Complementary Analysis”)
Step II Place them on Chromosomes( “Linkage Mapping”)
Step III Determine their distances & refine (“Linkage, 3-point Mapping”)
Complementary Groups
Gene A
Blue Eyes
Induce Mutations in Population
No blue eyes
Gene B
Protein A Protein B
How to Find Complementary
Groups?
Same Complementary
Group
Some disrupt Gene A
Some disrupt Gene B
No blue eyes
Breed & Observe Phenotypes
Complementation Analysis
Mutation I Mutation II
Case I: Allelic Mutations Case II: Non-Allelic Mutations
Mutation I Mutation II
Recessive Mutations
Mutant
Phenotype
Wild Type
Mapping “Linkage Groups”“Unlinked” Reproduction
A B a b
a b a bA a
B b bb
a a
25% aB
25% ab
25% AB
25% Ab
Mapping “Linkage Groups”“Linked” Reproduction with Recombination (w/ probability p)
A a
B b bb
a a
ab
AB
A a
Bb bb
a a
aB
p/2
Cis
1
2
p
Recombination
w/
Prob. p
1
2
p
Ab
p/2
Mapping “Linkage Groups”
Placing Genes on Chromosomes (to 0th order):
Recombination Frequency (p) ~ Distance of Genes
Part of X-chromosome of Drosophila
C. Elegans – “Nature’s Gift”Small ( ~ 1 mm).
Fast life cycle ~ adulthood in less than 24 hrs, live 3 weeks
959 somatic cells.
Most adults are hermaphrodites.
Genetic composition:
5AA + XX
Rare males:
5AA + XO
Easy to handle in large quantities.
Popularity of C. Elegans
First organism to have its DNA sequenced (1998).
Only organism to survive Columbia shuttle crash (2003)
Exhibit same symptoms as humans when quit smoking.
www.wormbase.org
Popularity of C. Elegans
Search Term Google Entries
C. Elegans 2,550,000
Drosophila 1,040,000
Depeche Mode 4,120,000
God 421,000,000
Sex 460,000,000
Isolating Mutants
For recessive mutations:
The F2 generation of an infected parent will be 25% homozygote mutants
Continue for another generation to ensure mutants can reproduce
Classifying Mutants
Recessive vs. Dominant Need to maintain wild-type male population
Male population 0.02% in nature Keep male population by crossing w/
hermaphrodites In real life, we need to take into account co-
dominant mutations, sex-linked mutations, etc …
Some Mutants . . .
Gene Mapping in C. Elegans
Mutant 1 Mutant 2
Allelic Mutations Non-Allelic Mutations
Example: Complementation test for Recessive Mutations
Mutant
Phenotype
Wild Type
Mutant 1 Mutant 2
Allelic Mutations
Mutant
Hermaphrodite
WT
Male
WT Male
Mutant
Hermaphrodite
WT
Male
WT MaleMut. Herm, Mut. Herm,
Non- Allelic Mutations
50% Male
WT
50% Male
Mutants
100% Male WT
Brenner’s Map
Brenner’s Map
Conclusions (cont.)
Map 258 Autosomal Mutations into 77 Complementation Groups
Accompanying paper: ~ 6.7x107 base pairs Naïve reasoning: ~ 6.7x104 proteins (genes) EMS induces mutations at rate 5x10-4/gene Mutations per worm: ~ 34
In practice: induced lethal frequency is 0.15 per X chromosome
Conclusion: 300 “essential” X-Chrom. Genes Scaled estimate: 2000 “essential” genes
Further Work
John White
Mapping the Nervous system
John Sulston
Tracing cell lineage, apoptosis
Bob Horvitz
Connection between worms & humans
Tracing the Cell Lineage
• Somatic cells vs. Germ cells
• John Sulston: the first to observe cell differentiation in a multicellular organism in real time
• Always the same
• Cells die of themselves
• C. Elegans is ideal:
• Simple (<1000 somatic cells)
• Transparent
Complete Lineage Map:
Cell Differentiation is “Rigid”
Nature or nurture? Development of
Reproduction Organs
It seems that nature!
Do genes really specify the development?
Genes dictate development
• 302 Neurons for adult hermaphrodite (382 for male)• To map: 20,000 slices, 0.05μm thick• Lineage of neurons can be traced• Re-enter: Brenner
Induce movement-
related Mutation
Examine Nervous System
Check if it is
Hereditary
Genes indeed dictate development!!!
Cell Death (Apoptosis)
• Cell death:• Injury, infection, . . . • By their own volition:
• By external signals• By internal “pre-programming”
• Purposes: structure formation, removing aged cells, …• In C. Elegans, 131 cells are “programmed to die”• Internal preprogramming ↔ genetic origin?
Evidence of Cell “Fate” J. E. Sulston, Philos. Trans. R. Soc. London Ser.B 1976, 275, 287-298: The
nuc-1, controlling the activity of DNA endonuclease E.M.Hedgecock,J.E.Sulston,J.N.Thomson, Science 1983, 220, 1277-1279:
ced-1, ced-2 genes responsible for cell engulfment (phagocytosis) “Jackpot”: ced-3
H. R. Horvitz et al. ,Neurosci. Comment. 1982, 1,56-65. H. M. Ellis, H. R. Horvitz, Cell 1986, 44, 817-829.
Visual Evidence of Cell Death
Taken from Horvitz (1986)
(bar = 10 micron)
Programmed cell death does not occur in a ced-3 mutant.
Mapping the Genetic Pathway
From Worms to Humans
Biological Universality
C. Elegans shares about 35% of its genome with humans
Horvitz Nobel lecture, 2002:
“One point that emerges . . . is the striking similarity of genes . . .
among organisms . . . I like to refer to this theme as “the principle of
biological universality” . . . and it underlies my conviction strong
conviction that the . . . study of the biology of any organism is likely
to lead to findings of importance in the understanding of other
organisms, including ourselves.”
The End
Thank you!
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