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Mendel forever

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The dataExpt. 1. Form of seed. -- From 253 hybrids 7324 seeds were

obtained in the second trial year. Among them were 5474 round or roundish ones and 1850 angular wrinkled ones. Therefrom the ratio 2.96:1 is deduced.

Expt. 2. Color of albumen. -- 258 plants yielded 8023 seeds, 6022 yellow, and 2001 green; their ratio, therefore, is as 3.01:1.

Expt. 3. Color of the seed-coats. -- Among 929 plants, 705 bore violet-red flowers and gray-brown seed-coats; 224 had white flowers and white seed-coats, giving the proportion 3.15:1.

Expt. 4. Form of pods. -- Of 1181 plants, 882 had them simply inflated, and in 299 they were constricted. Resulting ratio, 2.95:1.

Expt. 5. Color of the unripe pods. -- The number of trial plants was 580, of which 428 had green pods and 152 yellow ones. Consequently these stand in the ratio of 2.82:1.

Expt. 6. Position of flowers. -- Among 858 cases 651 had inflorescences axial and 207 terminal. Ratio, 3.14:1.

Expt. 7. Length of stem. -- Out of 1064 plants, in 787 cases the stem was long, and in 277 short. Hence a mutual ratio of 2.84:1. In this experiment the dwarfed plants were carefully lifted and transferred to a special bed. This precaution was necessary, as otherwise they would have perished through being overgrown by their tall relatives. Even in their quite young state they can be easily picked out by their compact growth and thick dark-green foliage.

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Sweet

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Mendel's dwarfing gene: cDNAs from the Le alleles and function of the expressed proteinsMartin et al. Proc. Natl. Acad. Sci. USA Vol. 94, pp. 8907-8911

Four alleles of the Le locus known: Le > le-3 > le > led.

“gibberellins” The stature of Le mutants can be restored

by application of GA1, but not by GA20

Herr Mendel, meet Drs. Beadle and Tatum

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Mendel's green cotyledon gene encodes a positive regulator of the chlorophyll-degrading pathway

Sato et al. PNAS August 28, 2007 vol. 104 14169-14174

“Mendel's Green Cotyledon Mutation Is a Stay-Green Mutation. The pea strain JI2775 has a defect on the I locus and presents green cotyledons in mature seeds …. And also retains greenness of leaves during leaf senescence, suggesting that JI2775 is a stay-green mutant and that the I gene functions in Chl degradation both in cotyledons during seed maturation and in true leaves during senescence

Also see Armstead, et al. (2007) Science 315, 73

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1866 – 2007 – ∞

Solely on the basis of statistical analysis of plant crosses, and a full century before the discovery of the structure of DNA, the chemical nature of mutations, the “central dogma (fact) of molecular biology,” the notion that genes code for enzymes (or enzyme subunits) – in essense, before the discovery of anything at all relevant to what is being studied here – Mendel correctly determined that differenced between plants in traits such as seed shape, plant height, and seed color are determined by discrete entities that behave in a distinct, specific way during meiosis, and during ontogeny.

Nearly 150 years later, molecular genetic and biochemical analysis has shown that Mendelian inheritance of the traits Mendel studied is, in precise agreement with his model (take THAT, Karl Nägeli and your stupid apomixing Hieracium; grrrrrrr), due to specific lesions (mutations) in single genes coding for enzymes directly involved in the development of the traits that Mendel studied. Furthermore, the nature of the lesions conclusively explains the recessive nature of the alleles Mendel studied.

These examples offer wonderful case studies in the effect of genotype on phenotype.

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“It’s All in the Genes”New York Times, 5/2/04

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Gene phenotype

Other genes epistasis variable expressivity (sickle-cell anemia)

The environment “norm of reaction” variable penetrance (BRCA1-induced breast cancer)

Epigenetic effects

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Plant epigenetics 1:mitotically, but not meiotically

resistant inheritance

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Persephone, the Greek goddess of Spring

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A bit of plant biology – the seeming paradox of vernalization

“Vernalization is the process by which prolonged exposure to cold temperatures promotes flowering.

Monocarpic species senesce after flowering and setting seed. … Plants that require vernalization to flower thus typically require two seasons to complete the life cycle and are usually classified as biennials or winter annuals.

Many winter annuals and biennials become established in the fall, taking advantage of the cool and moist conditions optimal for their growth. The vernalization requirement of such plants prevents flowering until spring has actually arrived. ”

R. Amasino The Plant Cell 16:2553-2559 (2004)

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Vernalization

A) A biennial cabbage (Brassica oleracea) variety with an obligate vernalization requirement that had been growing for five years without cold exposure. The small plant in my daughter's hands is a summer-annual variety of B. oleracea that flowers rapidly without vernalization. (B) and (C) Summer annual and vernalization-requiring types of henbane (B) and Arabidopsis (C). In both examples, a single-dominant gene is responsible for the vernalization-requiring habit. All plants were grown in long days (inductive photoperiods) without vernalization. The rapid-flowering summer annuals (which have initiated flowering) are at left and the winter-annual types at right.

R. Amasino The Plant Cell 16:2553-2559 (2004)

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“I think it is reasonable to refer to the vernalization-induced, mitotically stable acquisition of the competence to flower as an epigenetic switch because it is a change that can be propagated through cell divisions in the absence of the inducing signal.”

R. Amasino The Plant Cell 16:2553-2559 (2004)

15MCB 140, 10-3-07R. Amasino The Plant Cell 16:2553-2559 (2004)

“The vernalization-mediated repression of FLC is epigenetic in the sense discussed above: The repressed state of FLC is maintained after vernalized plants are returned to warm growing conditions. Thus, in Arabidopsis, vernalization provides competence to flower by repressing the expression of a flowering repressor. As expected, FLC expression is on again in the next generation. This resetting of the epigenetic switch during passage to the next generation is reminiscent of genomic imprinting in animals. But the unique aspect of this switch is that the on-to-off direction of the switch is set by perception of the environment, whereas the off-to-on direction is set by passage to the next generation.”

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Tragic memories“… some might argue that the term epigenetic

should be used only for changes that persist from one generation to the next. This of course does not happen in the case of vernalization; if it did, a biennial would only be a biennial for one generation.

One of Lysenko's false claims was that the vernalized state was heritable; that is, a vernalized plant would transmit the rapid-flowering trait to the next generation. This fit the Marxist ideology that the environment of the members of a Marxist society could produce heritable changes in attitude, and, thus, if the proper environment was provided, future generations would consist of improved citizens. Lysenko's efforts to obtain or fabricate results that supported a political ideology and, with the assistance of Stalin's regime, to force others to accept his views had disastrous consequences for Russian genetics.”

R. Amasino The Plant Cell 16:2553-2559 (2004)

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Plant epigenetics 2:mitotically and meiotically

resistant inheritance

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Maize (corn) – Zea mays

Maize is monoecious – distinct male (tassel) and female (“corn silk”) flowers on the same plant.

19MCB 140, 10-3-07Courtesy of Prof. Jay Hollick, MCB Department

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?!!!

R. Alexander Brink, 1950Vicky Chandler, Jay Hollick

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What is going on?1. The green maize plant is not

outcrossing, it is selfing.2. The red allele is

incompletely penetrant, except when in a homozygous state.

3. This is actually two-locus inheritance, and the green locus exhibits full dominant epistasis to the red locus.

4. This is actually two-locus inheritance, and the green locus exhibits full recessive epistasis to the red locus.

5. Mmmmm … Milla …

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What IS going onThis is paramutation:a meiotically heritable change in

the phenotype of a plant that results not from the physical alteration of the primary DNA sequence of the underlying genes, but from an effect on their expression.

Recall Mendel’s “compromise that the two alleles of a gene reach in a heterozygous organism for the life of a plant.”

In this case, the compromise is not reached. The Pl’ allele somehow causes the epigenetic silencing of the Pl allele – silencing that persists through meiosis.

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Transcriptional Regulation of Anthocyanins

Courtesy of Prof. Jay Hollick, MCB Department

24MCB 140, 10-3-07Courtesy of Prof. Jay Hollick, MCB Department

Other genes involved in maize pigment synthesis can be paramutated

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A critical aspect of the paramutation process – one that is shared by many epigenetic processes

It is sensitive to the environment:

paramutation can be variably expressive (in other words, the extent of paramutation can vary, and how variable it is can be specified by the environment).

26MCB 140, 10-3-07Genetics 140 1379-1387

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32C LL 22C LL

Grandpaternal seedling conditions

Genetics 140 1379-1387

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Three processes

Vernalization:

Environment epigenetic state that is mitotically heritable

Paramutation

“Dialog” of two alleles epigenetic state that is meiotically heritable and sensitive to the environment

Transgenerational memory of stress:

environment epigenetic state that is meiotically heritable

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Transgeneration memory of stress in plants – Barbara Hohn et al. Nature 442, 1046-1049

“Owing to their sessile nature, plants are constantly exposed to a multitude of environmental stresses to which they react with a battery of responses. …Here we show that in Arabidopsis thaliana plants treated with short-wavelength radiation (ultraviolet-C) or flagellin (an elicitor of plant defences6), somatic homologous recombination of a transgenic reporter is increased in the treated population and these increased levels of homologous recombination persist in the subsequent, untreated generations. The epigenetic trait of enhanced homologous recombination could be transmitted through both the maternal and the paternal crossing partner, and proved to be dominant. The increase of the hyper-recombination state in generations subsequent to the treated generation was independent of the presence of the transgenic allele (the recombination substrate under consideration) in the treated plant. We conclude that environmental factors lead to increased genomic flexibility even in successive, untreated generations, and may increase the potential for adaptation.”

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1. Plants are engineered to have a “reporter gene” – if the gene undergoes recombination, blue sectors appear.

2. Irradiation of the plants increases the number of blue sectors (not surprising).

3. Most surprisingly, the offspring of the irradiated plants show increased recombination frequency.

Molinier et al. Nature 442, 1046-1049

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Great-grand-grand-children

Molinier et al. Nature 442, 1046-1049

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Hmmmmmmmmmmmm

“Influences of the environment on the plant genome have been documented, [and] have been interpreted as 'genomic shock'. Here we demonstrated that environmental influences, specifically ultraviolet radiation and a bacterial elicitor, change the flexibility of the plant genome in somatic tissue of treated plants and in somatic tissue of their progeny. As these influences persist in the entire population of plants, the basis for the change is epigenetic rather than genetic. Plants carrying the transgene locus do not have to face the environmental challenges themselves in order to transmit the epigenetic change to the offspring; the stimulus for an increase of recombination can be imposed in trans by a single treated parent.”

Molinier et al. Nature 442, 1046-1049

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Epigenetics:from phenomenon to mechanism

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An important point

Paramutation and across-generation memory of stress are both epigenetic phenomena: the loci they affect do not change DNA sequence.

This does not mean, however, that the mediators of these epigenetic effects are fundamentally “magical” – there are genes, the products of which are required for epigenetic effects.

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From screen … To mechanism:

Hollick and Chandler Alleman et al.Genetics, Vol. 157, 369-378 Nature 442, 295-298

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Arabidopsis thaliana

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Some mutations that affect flower structure

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S. Jacobsen and E. Meyerowitz

“Superman”

Wild-type flower: 6 stamens (♂).

superman null mutations: 12 stamens.

39MCB 140, 10-3-07S. Jacobsen, UCLA

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No comment on the nomenclature

There are two ways to mutate the SUPERMAN gene and get the same phenotype (12 stamens):

1. Just delete the gene.

2. Keep the gene as it and METHYLATE it!

The methylated form is an “epi-allele” of SUPERMAN and is called clark kent.

S. Jacobsen, UCLA

41MCB 140, 10-3-07S. Jacobsen, UCLA

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Suppressor mutants:Cmt3 (the DNMT)or kyp (kryptonite)

clk-st

S. Jacobsen, UCLA

Suppressors of clark kent

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kyp (kryptonite) codes for an H3K9 histone methyltransferase!

Jackson et al. (Jacobsen) Nature 416: 556-560 (2002).

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From plants to mice

Chandler (2007) Cell 128: 641

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An evolutionary perspective

Mikula (1995) Genetics 140 1379-1387

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And now, to conclude the section on “classical” genetics – a real-

world perspective on the complexity of life

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Emery and Clayton (2005) “The Mentality of Crows: Convergent Evolution of Intelligence in

Corvids and Apes” Science 306: 1903

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“Many animals use tools, but their understanding of physical forces or causal relations is unclear. Primates are considered the most versatile and complex tool users, but observations of New Caledonian crows (Corvus moneduloides) raise the possibility that these birds may rival nonhuman primates in tool-related cognitive capabilities. We report here an experiment inspired by the observation that a captive female spontaneously bent a piece of straight wire into a hook and successfully used it to lift a bucket containing food from a vertical pipe.”

Weir et al. (2002) Science 297: 981

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No comment

“The male rarely attempted this task and never bent the wire. He observed the female bending the wire and stole the food from her in three trials.”

Weir et al. (2002) Science 297: 981.

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An actual e-mail

Hey Professor Urnov, This is X from your MCB 140 course at Berkeley. How are you doing and how are things at Sangamo? I hope things are going well for you. As for me, [after] graduat[ing] from Berkeley I have moved back home to X.

I have am also considering a career in Y and am currently applying. I was wondering if you'd be willing to provide a letter of reference for me. If possible, please let me know so that I can properly send you the materials necesary for doing so. Thank you. Sincerely,

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An actual application

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A two-step solution

1.

2. Horace Judson The Eighth Day of Creation; Steven Jay Gould The Mismeasure of Man

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Next

1. No office hours today.

2. Review session 7 pm (sharp) – 9 pm – will feature practice midterm and lots of “clicking” – answers to homework will be given – Thursday, Oct. 4, Dwinelle 155

3. Answers to previous midterm and to homeworks will be posted shortly