The Genetics of Caenorthabditis Elegans Brenner 1974 Lecture

The Genetics of Caenorthabditis Elegans Brenner, 1974 Lecture by Assaf Tal

Talk Outline l l l 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 - ) Ø Discoverer of frameshift mutations (1962) Ø Believed “classical” molecular biology was “solved” ØLetter to Max Perutz (1963): “. . . Attempt to define the unitary steps of development in terms of genetic analysis … “ Ø 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 Homologous Chromosomes Gene Alleles (recessive/dominant) Gene: Ø Set of bases in DNA Genotype: Aa Ø Paradigm: 1 Gene per Protein Phenotype: “what we see” ØAllele: A particular Gene Realization Wild Type: “common” phenotype Mutant: “non-natural” phenotype

How to Make a Genetic Map Step I are Find out how many genes 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 Protein A Gene B Protein B Blue Eyes Same Complementary Group How to Find Complementary Groups? Induce Mutations in Population Some disrupt Gene A Some disrupt Gene B No blue eyes Breed & Observe Phenotypes

Complementation Analysis Recessive Mutations Case I: Allelic Mutations Mutation II Mutant Phenotype Case II: Non-Allelic Mutations Mutation II Wild Type

Mapping “Linkage Groups” “Unlinked” Reproduction A a a B b a b 25% AB 25% a. B 25% Ab 25% ab b

Mapping “Linkage Groups” “Linked” Reproduction with Recombination (w/ probability p) Recombination Cis w/ A a a a B b b b Prob. p A a a a b B b b AB a. B p/2 ab Ab p/2

Mapping “Linkage Groups” Placing Genes on Chromosomes (to 0 th order): Recombination Frequency (p) ~ Distance of Genes Part of X-chromosome of Drosophila

C. Elegans – “Nature’s Gift” ØSmall ( ~ 1 mm). ØFast life cycle ~ adulthood in less than 24 hrs, live 3 weeks Ø 959 somatic cells. ØMost adults are hermaphrodites. ØGenetic composition: 5 AA + XX ØRare males: 5 AA + XO ØEasy to handle in large quantities.

Popularity of C. Elegans l l 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 Sex 460, 000

Isolating Mutants For recessive mutations: The F 2 generation of an infected parent will be 25% homozygote mutants Continue for another generation to ensure mutants can reproduce

Classifying Mutants l l Recessive vs. Dominant Need to maintain wild-type male population l l l Male population 0. 02% in nature Keep male population by crossing w/ hermaphrodites In real life, we need to take into account codominant mutations, sex-linked mutations, etc …

Some Mutants. . .

Gene Mapping in C. Elegans Example: Complementation test for Recessive Mutations Allelic Mutations Mutant 1 Mutant 2 Mutant Non-Allelic Mutations Mutant 1 Mutant 2 Allelic Mutations Non- Allelic Mutations Mutant WT Hermaphrodite Male WT Male Mut. Herm, 50% Male WT Mutants WT Male Mut. Herm, Wild Type Phenotype 100% Male WT

Brenner’s Map

Brenner’s Map

Conclusions (cont. ) l l Map 258 Autosomal Mutations into 77 Complementation Groups Accompanying paper: ~ 6. 7 x 107 base pairs l l l Naïve reasoning: ~ 6. 7 x 104 proteins (genes) EMS induces mutations at rate 5 x 10 -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 John Sulston Bob Horvitz Mapping the Nervous system Tracing cell lineage, apoptosis 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” l l 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 movementrelated 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” l l l 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 l H. R. Horvitz et al. , Neurosci. Comment. 1982, 1, 56 -65. l H. M. Ellis, H. R. Horvitz, Cell 1986, 44, 817 -829.

Visual Evidence of Cell Death Programmed cell death does not occur in a ced-3 mutant. Taken from Horvitz (1986) (bar = 10 micron)

Mapping the Genetic Pathway

From Worms to Humans

Biological Universality l l 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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