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Complications to Mendel: Gene Interactions Lecture starts on next page

The Complicated Relationship between Genotype and Phenotype

This is a single gene trait. The coloration of the flowers depends on the: • production of the appropriate anthocyanin pigments • presence of metal ions and co-pigments • the vacuolar pH in epidermal cells

These wild-type (b) and mutant (a) morning glories produce exactly the same anthocyanin pigment in their petals. The phenotypic difference in the two plants is due to a single gene difference: the flower on the left has a recessive, loss-of-function mutation in a gene that codes for a Na+/H+ exchanger. This mutation results in a decrease in the vacuolar pH of epidermal cells causing the petals to appear purple rather than blue. Using genetical conventions, we would name the gene defined by this phenotypic variation the purple gene and assign the following allele symbols: p+ = blue p = purple

a. vaculolar pH = 6.6 b. vacuolar pH = 7.7

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The art and genetics of color in plants and animals

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Are these examples of color variation single gene traits? In other words, can a single gene (perhaps with multiple alleles and complications to dominance)

explain color variation in budgie parakeets or in bell peppers?

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The table below shows the results of crossing true-breeding lines of green, blue, yellow and white budgies. parents Green Blue Yellow White

Green green green green green

Blue green blue green blue

Yellow green green yellow yellow

White green blue yellow white Possibilities: • One gene: four alleles; dominance complete • One gene: three alleles; some incomplete dom • Two genes: two alleles; dominance complete

• Three or more genes

Number of genes? Number of alleles? Dominance?

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P true-breeding yellow X true-breeding blue

â

F1 green X F1 green â

F2 9/16 green 3/16 blue 3/16 yellow 1/16 white

How many genes involved?

Speculate about genotypes using elementary principles of

combining colors

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Additive Gene Effects • independent, additive contribution to phenotype: effects of the alleles of the

two loci are essentially the sum of the independent gene actions • unmodified Mendelian ratio: AaBb X AaBb à 9:3:3:1 (two genes; each gene

has two alleles, complete dominance) • essentially there is no gene interaction – the genotype at one gene locus

doesn’t affect the expression/function of the alleles at a second gene locus

Yellow and blue pigments are synthesized in independent biochemical pathways. A (mutational) disruption in one pathway does not affect the other pathway

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See also this beautiful example of additive gene effects in corn snakes (Chapter 6 of text) Two genes independently controlling the synthesis of two different pigments; each gene has two alleles showing complete dominance

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Drug metabolizing enzymes, DMEs (Phase I enzymes/Cytochrome P450 enzymes, e.g. CYP2D6; Phase II enzymes, e.g. N-acetyl transferases) • Drug transporters (Solute Carrier (SLC)- and ATP Binding Cassette (ABC)-transporters, e.g. organic cation transporters, OCTs, as members of the SLC family) • Drug receptors (ligand controlled ion channels or class 1 receptors, e.g. glutamate receptor; G-protein coupled receptors (GPCRs) or class 2 receptors, e.g. ß-receptor; enzymatic receptors, e.g. insulin receptor; receptors regulating gene expression, e.g. steroid hormone receptor) • G-proteins, e.g. GNAS1 or GNB3

Polymorphic Determinants of Drug effects

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Pharmacogenomics: Translating Functional Genomics into Rational Therapeutics SCIENCE VOL 286 15 OCTOBER 1999 • In this hypothetical example (next page) there are two genes that

influence the therapeutic effect of a particular drug. • Each gene has two alleles that show incomplete dominance

Gene Functions

1. One gene is involved in metabolizing the drug (for eventual excretion) – see previous lecture notes on the CYP genes

2. The second gene codes for a receptor protein via which the drug exerts its therapeutic effect

NOTE: not addressed in the following example is the potential role of genetic polymorphisms in drug transporters . Although passive diffusion accounts for cellular uptake of some drugs and metabolites, increased emphasis is being

placed on the role of membrane transporters in absorption of oral medications across the gastrointestinal tract; excretion into the bile and urine; distribution into “therapeutic sanctuaries,” such as the brain and testes; and

transport into sites of action, such as cardiovascular tissue, tumor cells, and infectious microorganisms.

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If the therapeutic effects of the two genes are strictly additive, we should see nine phenotype categories. Note, though, the effects of the mm receptor genotype. It determines the % therapeutic effect independent of the metabolism genotype

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Additive gene effects: RECAP • independent, additive contribution to phenotype: effects of the alleles

of the two loci are essentially the sum of the independent gene actions • unmodified Mendelian ratio: AaBb X AaBb à 9:3:3:1 • no interaction of the alleles – the genotype at one gene locus doesn’t

affect the expression/function of the alleles at a second gene locus Gene Interactions: Specific alleles of one gene mask or modify (enhance, suppress or in some way alter) the expression of alleles of a second gene • complementary gene action • epistatic gene interaction • modifiers & suppressors

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The complicated relationship between genes and phenotypes

Coat Variation in the Domestic Dog Is Governed by Variants in Three Genes 2 OCTOBER 2009 VOL 326 SCIENCE

Coat color and type are essential characteristics of domestic dog breeds. Although the genetic basis of coat color has been well characterized, relatively little is known about the genes influencing coat growth pattern, length, and curl. We performed genome-wide association studies of more than 1000 dogs from 80 domestic breeds to identify genes associated with canine fur phenotypes. Taking advantage of both inter- and intrabreed variability, we identified distinct mutations in three genes, RSPO2, FGF5, and KRT71 (encoding R-spondin–2, fibroblast growth factor–5, and keratin-71, respectively), that together account for most coat phenotypes in purebred dogs in the United States. Thus, an array of varied and seemingly complex phenotypes can be reduced to the combinatorial effects of only a few genes. See Figure on the next page:

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- represents ancestral allele (found in wolves) + = variant allele 3 genes, 2 alleles (complete dominance) 7 out of 8 combos are shown here Which is missing?

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Gene Interactions: Specific alleles of one gene mask or modify (enhance, suppress or in some way alter) the expression of alleles of a second gene • complementary gene action • epistatic gene interaction • modifiers & suppressors

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Complementary Gene Action

NATURE|VOL 431 | 21 OCTOBER 2004 |www.nature.com/nature See article: http://fire.biol.wwu.edu/trent/trent/Naturedeafdesign.pdf

Both John and Karen

are deaf due to

genetics.

Why then could they have only

normal children?

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A mutant gibberellin-synthesis gene in rice New insight into the rice variant that helped to avert famine over thirty years ago. Nature 416: 701 April 18, 2002

Right: wild-type Left: semi-dwarf (sd1) mutant (more resistant than wild-type to damage by wind and rain and respond better to certain fertilizers) Arabidopsis dwarf mutant

The chronic food shortage that was feared after the rapid expansion of the world population in the 1960s was averted largely by the development of a high-yielding semi-dwarf variety of rice known as IR8, the so-called rice ‘green revolution’. The short stature of IR8 is due to a mutation in the plant’s sd1 gene, which encodes an oxidase enzyme involved in the biosynthesis of gibberellin, a plant growth hormone. Gibberellin is also implicated in green-revolution varieties of wheat, but the reduced height of those crops is conferred by defects in the hormone’s signaling pathway. There are various reasons for the dwarf phenotype in plants, but gibberellin (GA) is one of the most important determinants of plant height. To investigate whether the sd1 gene in semi-dwarf rice (Fig. 1a) could be associated with malfunction of gibberellin, we tested the response of this mutant to the hormone. We found that sd1 seedlings are able to respond to exogenous gibberellin, which increases their height to that of wild-type plants

In the early 1970s, a Dr. Rutger, then in Davis, Calif., fired gamma rays at rice. He and his colleagues found a semi-dwarf mutant that gave much higher yields, partly because it produced more grain. Its short size also meant it fell over less often, reducing spoilage. Known as Calrose 76, it was released publicly in 1976. Today, Dr. Rutger said, about half the rice grown in California derives from this dwarf.

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You have three different (independently isolated) dwarf strains of Arabidopsis: Mutant Strains 1, 2 & 3 • Each dwarf strain has a recessive, loss-of-function mutation • Treatment of each mutant strain with gibberellic acid restores normal height

suggesting that they have the same primary defect • You cross dwarf strain 1 X dwarf strain 3 and the untreated F1 are dwarf • But when you cross dwarf strain 1 X dwarf strain 2 the untreated F1 are

wild-type in height (non-dwarf) • How to explain: dwarf X dwarf = wild-type?

These two plants have the exactly the same genotype: they are homozygous for the same recessive, loss-of-function mutation The plant on the left has been treated with an application of gibberellin. The plant on the right is untreated.

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gibberellin biosynthetic pathway

Dwarf plants can result from a mutation affecting gibberellin production as well as mutations affecting any step in the cellular response to this signal

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These mutations show complementation: Complementation: the production of wild-type F1 progeny when crossing two parents showing the same recessive mutant phenotype M1 X M2 wild-type F1 progeny (alleles symbols: uppercase = functional allele; lowercase = loss-of-function) mutant 1: aaBBCC X mutant 2: AAbbCC ê F1 AaBbCC wild-type ê self What phenotypes will appear in the F2? Will the progeny ratios be in 1/4’s or 1/16’s or 1/64’s

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Genes are assorting independently 3/4 A- 1/4 aa 3/4 B- 1/4 bb all CC (not segregating) 9/16 A-B- CC 3/16 aaB- CC 3/16 A- bb CC 1/16 aabbCC Since a homozygous recessive mutation in either A or B results in the mutant phenotype 9/16 wild-type 7/16 dwarf = modified Mendelian ratio

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Dwarfism in plants and deafness in humans are examples of genetic heterogeneity

Genetic (or locus) heterogeneity: Mutations in any one of several genes may result in identical phenotypes (such as when the genes are required for a common biochemical pathway or cellular structure) Heterogeneous trait or genetic heterogeneity: a mutation at any one of a number of genes can give rise to the same phenotype Although a single gene difference causes the phenotypic difference between the dwarf and the wild-type plants, this does not mean that normal height is the result of the action of a single gene. It means simply that only one gene differed* between the dwarf and wild-type plants under consideration

* carried alleles with functional differences

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Optional interesting growth-hormone-related genetics FDA approved genetically modified salmo: addition of Chinook growth hormone to smaller species http://opinionator.blogs.nytimes.com/2011/03/17/frankenfish-phobia/?nl=opinion&emc=tya1

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See also optional reading on loss-of-function mutations in growth hormone receptor in humans: Equadorean villagers may hold secret to longevity http://fire.biol.wwu.edu/trent/trent/larondwarfism.pdf