“for difference determined by one gene…”

MCB140 02-12-07 1

“For difference determined by one gene…”

MCB140 02-12-07 2

MCB140 02-12-07 3

MCB140 02-12-07 4http://seer.cancer.gov/publications/raterisk/rates28.html

Cancer Site 1974-76 1977-79 1980-82 1983-90Brain & Other Nervous 22.3 24.4 25 27.3Breast (females) 74.3 74.5 76.2 80.4Cervix Uteri 68.5 67.7 66.9 67.4Colon & Rectum 49.5 51.7 54.2 59.2Esophagus 4.7 5.1 6.7 9.2Hodgkin's Disease 71.1 73 74.3 78.9Kidney & Renal Pelvis 51.3 50.8 51.4 56.3Larynx 65.4 66.8 68 67Leukemias 34.2 36.6 37.4 38.3Liver & Intrahep 3.8 3.7 3.4 6Lung & Bronchus 12.3 13.3 13.3 13.4Melanoma of Skin 79.7 81.5 82.1 85.1Multiple Myeloma 24.4 26.1 28 27.7Ovary 36.5 38.1 38.9 41.8Pancreas 2.6 2.5 3.1 3.2Prostate 66.7 70.9 73.1 79.6Stomach 15.1 16.7 17.5 18.5Testis 78.6 87.2 91.7 93.3Thyroid 91.9 92.5 94.2 94.6Urinary Bladder 72.4 74.8 77.9 79.8All Sites 49.3 49.8 50.6 53.9

Five year survival rates, in %.

MCB140 02-12-07 5Hanahan and Weinberg (2000) Cell 100: 57–70.

MCB140 02-12-07 6Hanahan and Weinberg (2000) Cell 100: 57–70.

MCB140 02-12-07 7

MCB140 02-12-07 8

A Russian proverb that aptly describes most current cancer treatment modalities

«Лучшее средство от кровотечения из носа – жгут на шею».

“The best cure for a nosebleed is a torniquet on the neck.”

MCB140 02-12-07 9

“Chemo” drugs

Intrastrand DNA crosslinking

Apoptosis (programmed cell death)

Depolymerization of microtubules

Cell cycle arrest

MCB140 02-12-07 10

“Malignancy of somatic cell hybrids” B. Ephrussi et al., Nature (1969) 224:1314

The studies of Ephrussi et al. and Harris provided compelling evidence that the ability of cells to form a tumor is a recessive trait. They observed that the growth of murine tumor cells in syngeneic animals could be suppressed when the malignant cells were fused to nonmalignant cells, although reversion to tumorigenicity often occurred when the hybrids were propagated for extended periods in culture. The reappearance of malignancy was found to be associated with chromosome losses. Stanbridge and his colleagues studied hybrids made by fusing human tumor cell lines to normal, diploid human fibroblasts. Their analysis confirmed that hybrids retaining both sets of parental chromosomes were suppressed, with tumorigenic variants arising only rarely after chromosome losses in the hybrids. Moreover, it was demonstrated that the loss of specific chromosomes, and not simply chromosome loss in general, correlated with the reversion to tumorigenicity.

The observation that the loss of specific chromosomes was associated with the reversion to malignancy suggested that a single chromosome (and perhaps even a single gene) might be sufficient to suppress tumorigenicity. To directly test this hypothesis, single chromosomes were transferred from normal cells to tumor cells, using the technique of microcell-mediated chromosome transfer. It was found that the transfer of a single chromosome 11 into the HeLa cervical carcinoma cell line suppressed the tumorigenic phenotype of the cells. Many studies have now demonstrated that transfer of even very small chromosome fragments will specifically suppress the tumorigenic properties of certain cancer cell lines.

MCB140 02-12-07 11Ventura et al Nature 445: 661

MCB140 02-12-07 12Xue et al Nature 445: 656

MCB140 02-12-07 13Martins et al Cell 127: 1323

MCB140 02-12-07 14

Why does restoration of p53 function lead to tumor

regression?

MCB140 02-12-07 15

MCB140 02-12-07 16Hanahan and Weinberg (2000) Cell 100: 57–70.

MCB140 02-12-07 17

Epistasis(or: an epistatic interaction between two loci)

1. Pick a trait.2. Find a mutant phenotype #13. Find a different mutant different phenotype

(#2)4. Cross the two mutants: get not a mix of

phenotypes, but instead, either phenotype #1 or #2.

The term “epistasis” refers to a phenomenon in which an allele of one gene masks (“stops”) the effects on the phenotype of an allele of a different gene.

MCB140 02-12-07 18

How can one tell, if two organisms under study that

exhibit mutant phenotypes for a particular trait have a mutation in

different genesor in the same gene?

MCB140 02-12-07 19

Complementation test

“Complementation is the production of a wild-type phenotype when two haploid genomes bearing different recessive mutations are united in the same cell.”

MCB140 02-12-07 20

The cis-trans test(aka complementation test)

Edward Lewis (NP 1995)

Are two different recessive mutations that appear to affect the same trait

in the SAME gene or in DIFFERENT genes?

MCB140 02-12-07 21

Orgo

trans-2-butene cis-2-butene

MCB140 02-12-07 22

The cis-trans test, 1949:lozenge (M. Greene)

Two different recessive mutants, both with the same phenotype (small eyes and fused facets).

Are they mutations in the same gene?

Make two different fly lines and compare their phenotypes.

Cis: Trans:

lz(BS) lz(g)

wt wt

lz(BS)

lz(g)wt

wt

MCB140 02-12-07 23

lz(BS) lz(g)

wt wt

lz(BS)

lz(g)wt

wt

If flies are normal, thenmutations are in different genes.

If the phenotype is still mutant,then BS and g must be in the

same gene!!!

This is a control experiment.The flies will be wild-type

regardless of whetherBS and g are in the same gene or not.

Cis: Trans:

MCB140 02-12-07 24

lz(BS)

lz(g)wt

wt

Gene BGene A

Aa Bb

MCB140 02-12-07 25

Baur et al. Nature 444: 337.

Lagouge et al. Cell 127: 1109.

MCB140 02-12-07 26Jasper Rine and Ira Herskowitz (1987) Genetics 116: 9-22.

Rine schematic

mate to a cells

Fig. 17.14

MCB140 02-12-07 27

The data

• Colonies screened: 675,000

• Colonies that mated to a: 295

• Major complementation groups: 4

silent information regulators:

SIR1, SIR2, SIR3, SIR4

Jasper Rine and Ira Herskowitz (1987) Genetics 116: 9-22.

MCB140 02-12-07 28

MCB140 02-12-07 29

Xeroderma pigmentosum

MCB140 02-12-07 30

What was actually done

1. By linkage analysis, it was discovered that the same disease (XP) can be caused by mutations in 7 distinct loci.

2. The cDNA from each gene was cloned.3. An assay was developed to measure, how sensitive to

UV light cells are.4. Experiment: take cells from patient type A, and

introduce each of the 7 cDNAs, one after another.5. Whichever cDNA restores the wild-type phenotype

corresponds to the gene that is mutated in that cell. Bin XP mutations into “complementation groups”!

MCB140 02-12-07 31

Wait a minute

Ahem.

Fine. You take a cell that’s mutant, stick in a gene, the cell is now wild-type, and you tell us this means the gene you stuck in is the gene that is mutated in the cell.

What if the cell has a mutation in a completely different gene, and the gene you stuck in is just epistatic to the first one?!

Good question.

MCB140 02-12-07 32

Auf Wiedersehen (with respect to phenotype), Herr Mendel

Gene interactions in the establishment of phenotype

MCB140 02-12-07 33

For complete clarity

1. All genotypes – except in cases of nondisjunction – follow Mendel’s first law, and – except in cases of linked genes < 50 cM away from each other – Mendel’s second law.

2. With the exception of human genetic disease, which is, let’s face it, very rare, and things like blood group inheritance, which belongs mostly on the MCAT (note – its inheritance, not blood groups themselves), the inheritance of phenotype seldom follows Mendel’s laws.

MCB140 02-12-07 34

http://www.gmi.oeaw.ac.at/

MCB140 02-12-07 35

“… the stadium capacity is now officially listed as 75,662”

MCB140 02-12-07 36

MCB140 02-12-07 37Steinberg Curr Opin Hematol 13: 131

MCB140 02-12-07 38

“An SCN9A channelopathy causes congenital inability to experience pain”

Nature Dec. 14, 2006“The index case for the present study was a ten-year-old child, well known to the medical service after regularly performing 'street theatre'. He placed knives through his arms and walked on burning coals, but experienced no pain. He died before being seen on his fourteenth birthday, after jumping off a house roof.”

MCB140 02-12-07 39

So – let’s think about this

The small fraction of African-Americans who are relatively pain-free …

… could they be heterozygous for a loss-of-function mutation in SCN9A?

In other words, could this be recessive epistasis?

If yes, could this suggest that a small-molecule inhibitor of that specific pain receptor could be a more effective analgesic for SCA patients than God-awful parenteral morphine!

MCB140 02-12-07 40

Strict definition

Epistasis is revealed in modified Mendelian ratios in dihybrid crosses.

MCB140 02-12-07 41

MCB140 02-12-07 42

MCB140 02-12-07 43

MCB140 02-12-07 44

Recessive epistasis

MCB140 02-12-07 45

Dominant epistasis

MCB140 02-12-07 46

MCB140 02-12-07 47

MCB140 02-12-07 48

Future e-mail

Professor,

Thanks for nothing, BUSTER.

First you tell us that modified Mendelian ratios can occur in monohybrid crosses (e.g., in a dominance series), then you tell us they occur in dihybrid crosses and are, in fact, a hallmark of epistasis.

How can one tell the difference?

MCB140 02-12-07 49

MCB140 02-12-07 50

Genetics of continuous variation

MCB140 02-12-07 51Muggsy Bogues, 5’3’’

Yao Ming, 7’6’’

Michael Jordan, 6’6’’

MCB140 02-12-07 52

In a population, phenotypes of individuals for a quantitative trait tend to be normally distributed

MCB140 02-12-07 53

The Great Schism (1901-1935)

“Naturalists”• the origin and

meaning of diversity• populations, groups,

higher taxa• gradualism• ultimate causation

“The Mendelians”• transformation of

genes• individual genes/loci• saltationism• no “why” questions

“Only the experimental method would permit an objective discussion of the theory of evolution, in striking contrast to the older speculative method of treating evolution as a problem of history.” (T.H. Morgan)

MCB140 02-12-07 54

Synthesis: Population Genetics

• H. Nilsson-Ehle, R.A. Fisher, J.B.S. Haldane, S. Wright: continuous phenotypic variation is not at odds with particulate inheritance:

multiple loci + epistasis

• S.S. Chetverikov: naturally occurring recessives as food for natural selection.

MCB140 02-12-07 55

Morgan, ch. 8

MCB140 02-12-07 56

Hermann Nilsson-Ehle

MCB140 02-12-07 57

Central limit theorem Carl Friedrich Gauss

If a variable is the sum of many independent variables, then its distribution will be normal:

e x 2

MCB140 02-12-07 58

MCB140 02-12-07 59

“Additive effects of genes”

MCB140 02-12-07 60

Next time:why are all calico cats female? –

and a related question:why is metastatic prostate cancer almost always incurably lethal?