Genome Size Variation: Consequences and Evolution

Ilia Leitch

D N VALUE D N VALUE

Genome size variation: consequences and evolution

(i) How genome size varies across plants (ii) What are the consequence of this variation (iii) How did such variation evolve Early genome size studies in plants

First genome size of a plant: Lilium longiflorum

Ogur M et al. 1951. Exp. Cell Res. 2: 73-89.

Concept of C-value: D N DNA amount in unreplicated VALUE gametic nucleus

Swift H. 1950. Proc. Natl. Acad. Sci. USA 36: 643-654. Plant DNA C-values database

www.kew.org/genomesize/homepage.html

5150 species

Land plants Algae 4427 angiosperms 91 Chlorophyta 207 gymnosperms 44 Phaeophyta 87 pteridophytes 118 Rhodophyta 176 bryophytes C-values in angiosperms range nearly 2000-fold

Genlisea UtriculariaTrillium Fritillaria margaretae rhombifoliumgibba assyriaca 1C = 0.065 pg 1C1C = 0.090 = 111.5 pg pg 1C = 127.4 pg

Greilhuber et al. 2006. GreilhuberGrif etet alal.. 2006.1980. Bennett. 1972. Plant Biology PlantTsitologiya Biology Proc. Roy. Soc. Lond. B 8: 770-777 8: 22:770-777 1331-1338 181: 109-135. Range of DNA amounts in land plants

Angiosperms (1.4%) 0.065 – 127.4

Gymnosperms (25%) 2.3 – 36.0 Seed plants Seed

Monilophytes (0.6%) 0.8 – 72.7 Vascular plants

Lycophytes (0.4%) 0.16 – 12.0

Bryophytes (1%) 0.09 – 6.4

1C DNA amount (pg) DNA amount in algae DNA amount in algae

Phaeophyta Rhodophyta Chlorophyta 44 species 118 species 153 species

Kapraun DF. 2005. Kapraun DF. 2007. Annals of Botany 95: 7-44. Annals of Botany 99: 677-701 Range of DNA amounts in algae 0.065 – 127.4 Angiosperms

Gymnosperms 2.3 – 36.0

Monilophytes 0.8 – 72.7

Lycophytes 0.16 – 12.0

Bryophytes 0.09 – 6.4

Phaeophyta (2.9%) 0.1 – 0.9

Rhodophyta (1.8%) 0.017 – 1.4

Chlorophyta (1.3%) 0.013 – 23.4 Ostreococcus tauri (Prasinophyta)

Chloroplast & starch grain Mitochondrion Golgi body Nucleus 0.1 μm

1C = 0.013 pg (12.56 Mb)

Derelle et al. 2006. Genome analysis of the smallest free-living eukaryote Ostreococcus tauri unveils many unique features. Proc Natl Acad Sci 103: 11647-11652. Genome size variation

Fritillaria assyriaca Genlisea margaretae Ostreococcus tauri 1C = 127.4 pg 1C = 0.065 pg 1C = 0.01 pg C-value paradox

Thomas CA. 1971. The genetic organization of chromosomes. Annual Review of Genetics 5: 237-256.

‘why the lowly liverwort has 18 times as much DNA as we have, and the slimy, dull salamander known as Amphiuma has 26 times our complement of DNA’.

Comings DE. 1972. Advances in Human Genetics 3: 237-431. C-value enigma Gregory TR. 2001. Coincidence, co-evolution, or causation? DNA content, cell size, and the C-value enigma. Biological Reviews 76: 65-101.

What types of non-coding DNA sequences predominate in genomes of different sizes? What are the mechanisms involved in generating genome size variation? What are the evolutionary forces driving genome size changes? What are the consequences of genome size variation? How has genome size evolved? Variation of genome size: Consequences at nuclear level ) 3 400 ) 3

m 8 m μ μ 300 6

200 4

100 2

0 Centromere ( volume Chromosome volume ( Chromosome volume 0 5 10 15 20 5 10 15 20

2 1C DNA amount (pg) 1C DNA amount (pg) 0 1 Bennett et al. 1983. Bennett et al. 1981. J. Cell Sci. 63: 173-179. J. Cell Sci. 47: 91-115. ) 35 3 3.5 m m) x μ μ 30 25 3.0 2 20 15 2.5 μm)x 10 ( 10

5 2.0 Mean total SC length length SC total Mean

0 ( volume nuclear Log Mean total SC length ( 0 10 20 30 40 1.0 1.5 2.0 1C DNA amount (pg) Log DNA per cell (pg)

Anderson et al. 1985. Baetcke et al. 1967. Exp. Cell Res. 156: 367-378. Proc. Natl. Acad. Sci. USA 58: 533-540. Variation of genome size: Consequences of timing

Mitosis Meiosis 30 300

20 200

10 100 Duration of meiosis (h) Duration of mitosis (h)

0 30 60 90 100 0 15 30 45 DNA amount per cell (pg) 1C1C DNA DNA amount amount (pg) (pg)

Van't Hof & Sparrow AH. 1963. Bennett MD. 1977. Proc. Natl. Acad. Sci. USA 49: 897-902. Phil. Trans. Roy. Soc. B 277: 201-277. Variation of genome size: Consequences at cell and tissue level

0.8 160

0.6 )

3 120

x 10 0.4 3 80 μm ( 40 0.2 Mass of 100 seeds (g)

Pollen volume dehiscenceat 0 0 0 102030 0102030 1C DNA amount (pg) 1C DNA amount (pg)

Relationship between pollen Relationship between seed volume and DNA amount weight and DNA amount in in 16 grass species. 12 Allium species. Bennett et al. 1972 Bennett et al. 1972 Consequences of variation in DNA amount

Whole plant level a) Life cycle options b) Life strategy options c) Ecology options d) Coping with environmental change Consequences of variation in DNA amount

Whole plant level a) Life cycle options Bennett MD. 1972. Nuclear DNA content and minimum generation time in herbaceous plants. Proceedings of the Royal Society of London Series B-Biological Sciences 181: 109-135. Consequences: life cycle options

400

300

Fritillaria meleagris 1C = 70.7 pg Arabidopsis 200 thaliana 1C = 0.16 pg 100 Duration of meiosis (h) meiosis of Duration

0 0 50 100 150 200 250 3C DNA amount (pg)

Bennett MD. 1977. Phil. Trans. Roy. Soc. B 277: 201-277. Consequences: life cycle options

No species in this triangle

Obligate Max. limiting perennials DNA amount

for annuals

A V

- Annuals and C

perennials

N D

Max. limiting DNA amount for ephemerals Annuals and perennials

Ephemerals

7 52 weeks weeks MINIMUM GENERATION TIME

Bennett MD. 1972. Proc. Roy. Soc. Lond. B 181: 109-135. Consequences of variation in DNA amount

Life cycle options: Conclusions

• DNA amount can impose limits on the type of life cycle a species can display • Species with small genomes may be ephemerals, annuals or perennials • Species with large genomes are restricted to being obligate perennials Consequences of variation in DNA amount

Whole plant level a) Life cycle options b) Life strategy options c) Ecology options d) Coping with environmental change Consequences of variation in DNA amount

Whole plant level b) Life strategy options: Potential to become a weed Bennett, Leitch & Hanson. 1998. DNA amounts in two samples of angiosperm weeds. Annals of Botany 82: 121-134. Consequences: option to be a weed

Method DNA amounts for 156 angiosperms recognised as weeds compared with 2685 non-weed species Consequences: option to be a weed

Non-weed species Weeds (c. 0.1 – 127.4 pg) (0.16 – 25.1 pg)

250 25 200 Mean 1C 20 Mean 1C 150 7.0 pg 15 2.9 pg 100 10 Max = Max = 50 127.4 pg 5 25.1 pg No. of species No. of species 0 Number of species 0 Number of species 0 100 200 300 400 500 0 25 50 75 100 125 00 25 100 20050 75 300 100 400 125 500 1C1C DNA DNA amount amount (pg)(pg) 1C1C DNA DNA amountamount (pg) (pg)

Bennett, Leitch & Hanson. 1998. DNA amounts in two samples of angiosperm weeds. Annals of Botany 82: 121-134. Success of an invasive weed

● Rapid establishment or completion of reproductive development ● Short generation time ● Rapid production of many small seeds Consequences: option to be a weed

12 10 8 6 4 % of all species which are weeds 2 0 0123456 Group (increasing DNA amount) Consequences of variation in DNA amount

Whole plant level a) Life cycle options b) Life style options c) Ecology options d) Coping with environmental change Genome size and latitude Pop. Several Picea sitchensis Miksche 1967, 1971

Sp. Tropical vs. temperate grasses Avdulov 1931 Sp. 329 tropical vs. 527 temperate plants Levin and Funderburg 1979 + correlation Sp. 17 Poaceae and 15 Fabaceae crops Bennett 1976

Pop. 24 Berberis in Patagonia Bottini et al. 2000

Sp. 20 Arachis duranensis in Argentina/ Temsch & Greilhuber 2001 Bolivia Pop. Several Festuca arundinacea Ceccarelli et al. 1992

Pop. North American cultivars of Zea mays Rayburn et al. 1985 Sp. 162 British plants Grime and Mowforth 1982 - correlation Sp. 23 Arctic plants Bennett et al. 1982

Pop. 22 N American Zea mays Rayburn et al. 1985

Pop. 11 N American Zea mays Laurie & Bennett 1985

Sp. 18 pines Joyner et al. 2001

Pop. 6 Allium cepa cultivars Bennett et al. 2000

Pop. Several Picea glauca Teoh & Rees 1976

Pop. 10 Dactylis glomerata Creber et al. 1994 no correlation

Sp. 11 Tropical vs. Temperate Pines Hall et al. 2000

Sp. 19 Helianthus Sims & Price 1985 Consequences: ecology options

401 species in the state of California

Knight & Ackerly. 2002. Variation in nuclear DNA content across environmental gradients: a quantile regression analysis. Ecology Letters 5: 66-76. Consequences: ecology options

Species with large genomes excluded from extreme environments

Species with small genomes can occupy all

DNA amount DNA amount environments

Ecological parameter e.g. temperature

Knight & Ackerly. 2002. Ecology Letters 5: 66-76. Consequences: ecology options

annual annual 25 perennial perennial

5 1C DNA amount (pg) 0 0.8 18 22 26 30 34 38 42 1.0 July maximum temperature (°C) 1.2 1.4 1.6

Knight & Ackerly. 2002. Variation in nuclear DNA content across environmental gradients: a quantile regression analysis. Ecology Letters 5: 66-76. Consequences: ecology options

annual 25 perennial 0.05 0.00 20 -0.05 15 -0.10

10 -0.15 -0.20 5 1C DNA amount (pg) Quadratic coefficient -0.25 0 -0.30 0 18 22 26 30 34 38 42 015 2040608010030 45 60 75 90 July maximum temperature (°C) Conditional quantile

Knight & Ackerly. 2002. Variation in nuclear DNA content across environmental gradients: a quantile regression analysis. Ecology Letters 5: 66-76. Consequences: ecology options

Summary

• The relationship between genome size and environmental factors is not uniform but appears to be stronger for species with large genomes • Species with large genomes are excluded from extreme environments Consequences of variation in DNA amount

Whole plant level a) Life cycle options b) Life style options c) Ecology options d) Coping with environmental change (i) Pollution (ii) Threat of extinction Consequences: Coping with environmental change

Pollution: The question

What effect does genome size have on the survival of plants in lead polluted soils?

B. Vilhar, T. Vidic and J. Greilhuber Consequences: Genome size and pollution

Lead smelter

Smelter chimney

Study plots

Dolina smrti in Slovenia Consequences: Genome size and pollution

Conc. of Reference lead in soil plot Conc. lead in 670 m 0.7% soil = 0.1% 520 m 1.5%

420 m 2.0% 330 m 3.3%

Smelter chimney

Lead smelter Consequences: Genome size and pollution

Range of 1C values (pg)

670 m

520 m

420 m 330 m 0 5101520 1C DNA amount (pg)

Smelter chimney

Lead smelter Consequences: Genome size and pollution

Percentage of species with large genomes in individual plots

30 Reference plot r = -0.986 P = 0.0021 20

10 genomes

Site closest Percentage of species

% of species with large to smelter 0 0102030 Concentration of Pb (g kg-1) Consequences: Genome size and pollution

Conclusions

Species with large genomes are at selective disadvantage in extreme environmental conditions induced by pollution. Consequences of variation in DNA amount

Whole plant level a) Life cycle options b) Life style options c) Ecology options d) Coping with environmental change (i) Pollution (ii) Threat of extinction Consequences: Genome size and threat of extinction

Pollution Invasive species

Habitat loss

Is genome size important? Vinogradov AE. 2003. Selfish DNA is maladaptive: evidence from the plant Red List. Trends in Genetics 19: 609-614. Consequences: Genome size and threat of extinction

Data and analysis

Plant DNA C-values Global concern = 305 database 3036 species Local concern = 1329 No concern = 1402

Vinogradov AE. 2003. Selfish DNA is maladaptive: evidence from the plant Red List. Trends in Genetics 19: 609-614. Consequences: Genome size and threat of extinction Results

1.7 42.5 pg 1.5

1.3

1.1 12.5 pg 0.9 7.4 pg

Log genome size (pg) 0.7 NoNo LocalLocal GlobalGlobal concern concernconcern concern Conservation status

Vinogradov AE. 2003. Trends in Genetics 19: 609-614. Consequences: Genome size and threat of extinction Analysis within families Genome size for species Genome size contrast = Mean genome size of family

32.4 Genome size contrast = = 4.2 7.7

1.2 Genome size contrast = = 0.3 4.3 Consequences: Genome size and threat of extinction Results

0.091.7

0.061.5

0.031.3

1.10

-0.030.9

-0.060.7 No Local Global

Genome size contrast (log) No Local Global concernconcern concernconcern concernconcern Conservation status

Vinogradov AE. 2003. Trends in Genetics 19: 609-614. Consequences: Genome size and threat of extinction Conclusions

Species with large genomes are at greater risk of extinction than those with small genomes. - Independent of life cycle type (at least partially) - Independent of polyploidy Consequences: Genome size and threat of extinction

Slow growing Restricted ecological obligate perennial distribution Threat of extinction of species with large More sensitive genomes to pollution Reduced rates of diversification (?)

Restricted trait variation (e.g. only large seeds) DNA amount variation and consequences

Summary

Huge variation in DNA amount in plants Consequences of this variation visible at: Cellular level Tissue level Whole organism level Possession of large genomes appear to impose constraints which operate at: Functional level Ecological level Evolutionary level DNA amount variation in angiosperms

Mode Mode = 0.6 pg 250 0.6 pg 200 150 100 Max. 50 127.4 pg 0 Number of species 0 25 50 75 100 125 0 100 200 300 400 500 1C DNA amount (pg) Range of DNA amounts

0.065 – 127.4 Angiosperms

Gymnosperms 2.3 – 36.0

Monilophytes 0.8 – 72.7

Lycophytes 0.16 – 12.0

Bryophytes 0.09 – 6.2

Phaeophyta 0.1 – 0.9

Rhodophyta 0.02 – 1.4

Chlorophyta 0.01 – 23.4

1C DNA amount (pg) Variation of genome size

1. Consequences of genome size variation in plants

2. Evolution of genome size variation Phylogenetic tree of angiosperms Santalales

and core C-value data eudicots Eudicots eudicots Early diverging Early

Monocots

Modified from:

Leitch, Chase, Basal angiosperms Bennett MD. 1998. Annals of Botany 82: 85-94. Angiosperms with ‘very large’ genomes (≥ 35 pg)

MONOCOTS CORE EUDICOTS • Liliales • Santalales – Liliaceae - Santalaceae – Melanthiaceae Viscum • Asparagales – Alliaceae – Alstroemeriaceae – Orchidaceae – Xanthorrhoeaceae • Commelinids – Commelinaceae Large scale analysis of genome size evolution in angiosperms

Data: Genome sizes for 4,119 species Method: The ‘All most parsimonious states’ resolving option of MacClade

C-value range Description ≤ 1.4 pg Very small ≤ 3.5 pg Small 3.51 – 13.99 pg Intermediate ≥ 14 pg Large ≥ 35 pg Very large

Soltis, Soltis, Bennett, Leitch. 2003. Am J Bot 90: 1596-1603. Large scale analysis of genome size evolution in angiosperms

Out- ‘BASAL’ ANGIOSPERMS EUDICOTS early group diverging monocots magnoliids eudicots core eudicots

asterids

Caryophyllales Proteales Ranunculales magnoliids eudicots monocots All angiosperms Reconstruction of C-value diversification across angiosperms

Out- BASAL ANGIOSPERMS EUDICOTS early group diverging monocots magnoliids eudicots core eudicots Reconstruction of C-value diversification across angiosperms

Out- BASAL ANGIOSPERMS EUDICOTS early group diverging monocots magnoliids eudicots core eudicots Land plant phylogeny

Angiosperms

Gymnosperms plants Seed

Monilophytes Vascular plants

Lycophytes

Bryophytes SEED PLANTS

Angiosperms Gymnosperms

25 1200 1000 20

800 15 600 10 400 Max. species of No. Max.

No. of species of No. 127.4 pg 5 200 36.0 pg 0 0 0 25 50 75 100 125 0 25 50 75 100 125 1C DNA amount (pg) 1C DNA amount (pg)

Mode: 0.6 pg Mode: 12 pg Mean: 6.13 pg Mean: 17.0 pg Range: 0.065 – 127.4 pg Range: 2.25 – 36.0 pg Reconstruction of genome size diversification across gymnosperms

Cycads

Ginkgo

Gnetales

Pinaceae

Coniferales II Land plant phylogeny

Angiosperms

Gymnosperms Seed plants

0.8 – 72.2 pg 16 Mode = 10 pg Monilophytes 12 8 4 No. species of 0

Lycophytes 0255075100125

1C DNA amount (pg)

Bryophytes Reconstruction of genome size diversification across monilophytes

? ferns Water Reconstruction of genome size diversification across monilophytes

? ? ? ferns Water Land plant phylogeny

Angiosperms

Gymnosperms plants Seed

Monilophytes Vascular plants

Lycophytes • Data for only 4 species • C-values range 75-fold Bryophytes Land plant phylogeny

Angiosperms

Gymnosperms plants Seed

Monilophytes Vascular plants

Lycophytes

Mosses Bryophytes Hornworts Liverworts C-values in mosses

25 No. Moss species with data = 176 20 Range: c. 12-fold 15

Min.: 0.17 pg in Holomitrium arboreum 10 No. of species of No.

Max.: 2.01 pg in Mnium marginatum 5

Mode: 0.45 pg 0 0 0.4 0.8 1.2 1.6 2

1C DNA amount (pg)

Data from: Voglmayr H. 2000. Annals of Botany 85: 531-546. C-values in hornworts

Anthoceros agrestis

1C = 0.087 pg

Unpublished data kindly provided by Greilhuber and colleagues, University of Vienna C-values in liverworts

Pellia endiviifolia 1C = c. 3.3 & 7.1 pg

Plagiochila asplenioides 1C = c. 1.4 & 3.1 pg

Mylia taylorii 1C = 6.4 pg

Unpublished data kindly provided by Greilhuber and colleagues, University of Vienna Genome size evolution across land plants

Mosses Lyco. Monilophytes Angios. Gymnosperms

Modified from Leitch et al. 2005. Ann Bot 95: 207-217. Analysis of genome size as a continuously varying character

‘Continuous’ • Mode (punctuated or gradual) • Tempo (recent or adaptive radiation) • Ancestral genome size reconstructed Pagel M. 1999. Nature 401: 877-844.

‘Analysis of traits’ (AOT) in Phylocom • Identifies nodes in phylogenetic tree which contribute significantly to observed pattern of genome size evolution Ackerly DD. 2006. http://www.phylodiversity.net/phylocom Analysis of genome size as a continuously varying character

GENOME SIZE EVOLUTION IN LILIACEAE

Data: 32 new C-value estimates 46 estimates from the Plant DNA C-values database

Leitch IJ, Beaulieu JM, Cheung K, Hanson L, Lysak MA & Fay MF Punctuated genome size evolution in Liliaceae. J Evol Biol (online 2007) Genome size evolution in Liliaceae

Fay et al. (2006) Ailso 559-565

Tempo of evolution: Accelerated rather than slow Mode of evolution: Punctuated rather than gradual Ancestral genome size: 1C = 6.7 pg Genome size evolution in Orobanche

Weiss-Schneeweiss H, Greilhuber J, Schneeweiss GM. 2005.

Genome size evolution in holoparasitic Orobanche (Orobanchaceae) and related genera.

American Journal of Botany 93: 148-156.

Mode of evolution: Punctuated rather than gradual Tempo of evolution: Slow rather than accelerated Genome size evolution in Liliaceae

‘Analysis of traits’ module in Phylocom Genome size evolution across land plants Tracking genome size evolution in fossils

Amphibian (Rana) 2 Birds Mammals Reptiles (non-avian

Genome size (ln [pg]) 0

3 4 5 6 Osteocyte cell size (ln [µm3])

Modified from Organ et al. 2007. Nature 446: 180-184 Tracking genome size evolution in fossils

20 μm

1000

800

600

400

200 x width (mean)

Guard cells from length Guard cell Fig. 4a redrawn from 0 fossil Platanus sp. Masterson (1994) 02468101214 Picograms of DNA

From: Masterson J. 1994. Stomatal size in fossil plants - evidence for polyploidy in majority of angiosperms. Science, 264: 421-424 Tracking genome size evolution

1C = 0.59 1C = 0.64 1C = 0.66 1C = 0.91 1C = 0.95

1C = 1.06 1C = 1.28 1C = 1.96 1C = 3.92 1C = 5.91

1C = 10.34 1C = 20.22 1C = 27.93 1C = 28.37 1C = 42.48

Unpublished data from Beaulieu, Knight and Leitch Tracking genome size evolution

Unpublished data from Beaulieu, Knight and Leitch Tracking genome size over land plant evolution Tracking genome size over mass extinctions Acknowledgements

Jodrell Laboratory,Royal Botanic Gardens, Kew, UK Mike Bennett Mike Fay Lynda Hanson

Masaryk University, Brno,Czech Republic

Martin Lysak

California Polytechnic State University, USA Charley Knight Arjun Pendharkar

Biological Sciences, Yale University, USA Jeremy Beaulieu