Genomic Conflict and DNA Sequence Variation Marcy K. Uyenoyama Department of Biology Duke University

Overview Population genetics Self-incompatibility systems in plants Historically model-rich Present need: model-based interpretation of observed patterns of genomic variation What are hallmarks of each model? Self-incompatibility systems in plants Recognizing genomic conflict due to sexual antagonism

Canonical models Neutral evolution Selection Pure neutrality: distribution of offspring number is independent of any trait in parent Demographic history: deme founding, gene flow Purifying selection: maintain functioning state against random deleterious mutations Selection Balancing selection: maintenance of different forms Selective sweeps: substitution of most fit for less fit

Hallmarks of evolution How do we know it when we see it? Patterns evident in genome variation Model selection Choosing among a small number of canonical models for any particular system

A random sample of genes Observed Sample Ancestral sequence

Allele and mutation spectra Site frequency spectrum Number of mutations Multiplicity a = {a1 = 6, a3 = 1, a5 = 1, a6 = 1}, for ai the number of alleles with multiplicity i

The neutral coalescent Sample root from stationary distribution of P, mutation transition matrix and bifurcate After an interval choose a lineage at random Replace it by two identical copies with probability Mutate it according to P with probability

Evolutionary rates Events on level k Bifurcation at rate Mutation at rate Population parameters: ratios of rates Next event is a bifurcation/coalescence with probability

Allele and mutation spectra Site frequency spectrum Number of mutations Multiplicity a = {a1 = 6, a3 = 1, a5 = 1, a6 = 1}, for ai the number of alleles with multiplicity i

Infinite-alleles model Mutation Novel allelic types formed at rate u per gene per generation Reproduction Frequency of allele i in the parental population: pi Multinomial sampling of N genes to form the offspring To find: probability of the sample of n genes (n1, n2, …, nk) or (a1, a2, …, an) for k the number of distinct haplotypes (alleles) ni the number of replicates of allele i ai the number of alleles with i replicates

Ewens sampling formula a = (a1, a2, …, an), for ai the number of alleles represented by i replicates in a sample of size n θ = 2Nu, for N the effective number of genes and u the per-locus, per-generation rate of mutation Ewens (1972, Theoretical Population Biology)

Allele and mutation spectra Site frequency spectrum Number of mutations Multiplicity a = {a1 = 6, a3 = 1, a5 = 1, a6 = 1}, for ai the number of alleles with multiplicity i

Population genomics About 750 accessions isolated from natural populations worldwide Summary statistics for sample of 19 entire genomes http://www.arabidopsis.org

Arabidopsis SNP spectra Minor allele counts 2 3 4 5 6 7 8 Site frequency spectra differ among functional classes Kim et al. (2008 Nature Genetics. 39: 1151)

ESF conditioned on two alleles Biallelic sample of size m Multiplicities i and (m – i ) independent of θ!

Ewens sampling formula a = (a1, a2, …, an), for ai the number of alleles represented by i replicates in a sample of size n θ = 2Nu, for N the effective number of genes and u the per-locus, per-generation rate of mutation Ewens (1972, Theoretical Population Biology)

Actual site frequency spectra Excess of rare and common types, deficiency of intermediate types Data from NIEHS Environmental Genome Project Direct resequencing of loci considered environmentally-sensitive Global representation of ethnicities Hernandez, Williamson, and Bustamante (2007)

Spectrum shape Signature of expansion? Signature of selective sweep? Expansions maintain more rare mutations Signature of selective sweep? Neutral variants experience selection as a population bottleneck Braverman et al. (1995) Black: constant population size Grey: recent expansion from small population size

Arabidopsis SNP spectra Minor allele counts 2 3 4 5 6 7 8 Site frequency spectra differ among functional classes Kim et al. (2008 Nature Genetics. 39: 1151)

Modelling a SNP data set Nordborg (2001 Handbook of Statistical Genetics) Single segregating mutation in the sample genealogy Conditional on exactly one segregating site, determine the distribution of the size (number of descendants) of the branch on which the mutation occurs Exactly two alleles in the sample Conditional on two haplotypes, bearing any number of segregating sites, determine the distribution of numbers of the two alleles

Conditioning Two alleles One segregating site

Multiplicity conditioned on a SNP Single segregating site in a sample of size m Multiplicity i dependent on θ ! Ganapathy and Uyenoyama (2009 Theoretical Population Biology)

Arabidopsis SNP spectra Minor allele counts 2 3 4 5 6 7 8 Site frequency spectra differ among functional classes Kim et al. (2008 Nature Genetics. 39: 1151)

Overview Population genetics Self-incompatibility systems in plants Historically model-rich Present need: model-based interpretation of observed patterns of genomic variation What are hallmarks of each model? Self-incompatibility systems in plants Recognizing genomic conflict due to sexual antagonism

Genomic conflict Phenotypes Conflict Multiple genes generally influence a given phenotype Conflict Target trait value differs among genes that control phenotype Sexual antagonism Male and female function collaborate in reproduction Genes influencing each function may come into conflict

Conflict and genomic variation Mating type regions as a battleground S-locus controls self-incompatibility in flowering plants How does sexual antagonism affect the pattern of molecular-level variation within the S-locus? What are hallmarks of conflict? Develop a basis for inference Model-based approach to the analysis of genetic variation

Flower development Fertilization Basic perfect flower includes both male and female components Fertilization Pollen grains deposited on stigma germinate and pollen tubes grow down style to the ovary Mariana Ruiz http://commons.wikimedia.org/wiki/File:Mature_flower_diagram.svg

Gametophytic SI (GSI) Pollen rejection Mariana Ruiz http://commons.wikimedia.org/wiki/File:Mature_flower_diagram.svg Gametophytic SI (GSI) Specificity expressed by individual pollen grain or tube determined by own S-allele Pollen rejection Growth of pollen tube arrested in style Norbert Holstein http://commons.wikimedia.org/wiki/File:Gametophytic_self-incompatibility.png

Sporophytic SI (SSI) Pollen rejection Mariana Ruiz http://commons.wikimedia.org/wiki/File:Mature_flower_diagram.svg Sporophytic SI (SSI) Specificity expressed by individual pollen grain or tube determined by the S-locus genotype of its parent Pollen rejection Germination of pollen grain may be arrested at stigma surface Norbert Holstein http://commons.wikimedia.org/wiki/File:Sporophytic_self-incompatibility.png

Sn An Bn Pistil (A) component: rejection of recognized specificities Mariana Ruiz http://commons.wikimedia.org/wiki/File:Mature_flower_diagram.svg Norbert Holstein http://commons.wikimedia.org/wiki/File:Gametophytic_self-incompatibility.png Pistil (A) component: rejection of recognized specificities Pollen (B) component: declaration of specificity An Bn Sn Norbert Holstein http://commons.wikimedia.org/wiki/File:Sporophytic_self-incompatibility.png

Mating type regions Uyenoyama (2005)

Human Y chromosome Non-recombining male-specific Y (MSY) Skaletsky et al. (2003 Nature 423: 825) Non-recombining male-specific Y (MSY) Euchromatic region ~ 23 MB Differences between two random Ys every 3 – 4 KB Mammalian sex determinant SRY Y-linked regulator of transcription of many male-specific Y-linked genes

Mating type regions Linkage between pistil (A) and pollen (B) components is essential to SI function Pollen: declaration of specificity Pistil: rejection of recognized specificities Uyenoyama (2005)

Brassica S-locus Natural populations often contain 30 – 50 S-alleles Pollen component Pistil component Natural populations often contain 30 – 50 S-alleles Nasrallah (2000 Curr. Opin. Plant Biol.)

Ubiquitin tags proteins for degradation Style: S-RNase disrupts pollen tube growth Upon entering a pollen tube, S-RNases initially sequestered in a vacuole In incompatible crosses, vacuole breaks down, releasing S-RNases into cytoplasm of pollen tube Pollen: SLF (S-locus F-box) Mediator of ubiquitinylation (attachment of ubiquitin) Disables all S-RNases except those of the same specificity Vierstra (2009, Nature Reviews Molecular Cell Biology)

Sexual antagonism Pistil: why reject fertilization? Screening of potential mates may improve offspring quality Cost under incomplete reproductive compensation: ovules may go unfertilized Pollen: why provoke rejection? Self-rejection may improve quality of own ovules Rejection by other plants reduces siring success Hide behind another S-specificity in sporophytic SI? Decline to declare S-specificity altogether?

GSI model Basic discrete time recursion Symmetries in genotype and allele frequencies Model change in frequency of focal allele i, assuming all other alleles in equal frequency Wright (1937, Genetics)

Diffusion approximation Change in allele frequency Diffusion equation coefficients holds for large population size (N) and u (rate of mutation to new S-alleles) of order 1/N Wright (1937, Genetics)

Wright’s diffusion model Diffusion with jumps Turnover rate Number of S-alleles Frequency in population

Expansion of time scale under balancing selection High rate of invasion of rare alleles Promotes invasion of new and retention of rare types Maintains high numbers of alleles Genealogical relationships Tree shape similar under symmetric balancing selection and neutrality Greatly expanded time scale Takahata (1993, Mechanisms of Molecular Evolution)

S-allele turnover Quasi-equilibrium of S-alleles Invasion of new, rare S-alleles balanced by extinction of common S-alleles Expansion of time scale Rate of divergence among S-allele classes similar to rate among neutral lineages, but in a population of size fN:

Gametophytic SI models Basic discrete time recursion Diffusion approximation Parameters: Effective population size (N) Rate of mutation to new S-specificities (u)

Simulation results Stationary distribution of allele frequency Most time spent close to deterministic equilibrium (1/n) or in boundary layer close to extinction Number of S-alleles Analytical expectation for number of common S-alleles Vallejo-Marín and Uyenoyama (2008)

Sn An Bn Pistil (A) component: rejection of recognized specificities Mariana Ruiz http://commons.wikimedia.org/wiki/File:Mature_flower_diagram.svg Norbert Holstein http://commons.wikimedia.org/wiki/File:Gametophytic_self-incompatibility.png Pistil (A) component: rejection of recognized specificities Pollen (B) component: declaration of specificity An Bn Sn Norbert Holstein http://commons.wikimedia.org/wiki/File:Sporophytic_self-incompatibility.png

Pollen specificity in GSI Each pollen expresses its own specificity Rarer specificities are incompatible with fewer plants Incompatible matings For n S-alleles in equal frequencies, a pollen type is incompatible with a proportion 2/n of all plants Norbert Holstein http://commons.wikimedia.org/wiki/File:Gametophytic_self-incompatibility.png

Sexual antagonism Pistil: why reject fertilization? Screening of potential mates may improve offspring quality Cost under incomplete reproductive compensation: ovules may go unfertilized Pollen: why provoke rejection? Self-rejection may improve quality of own ovules Rejection by other plants reduces siring success Hide behind another S-specificity in sporophytic SI? Decline to declare S-specificity altogether?

Fate of style-part mutant Self-pollen fraction (s) Relative viability of inbred offspring (s ) An+1 Bn Sa Full SC Polymorphism Full SI

Fate of pollen-part mutant Bn+1 Sb Full SC Relative viability of inbred offspring (s ) Disruption Polymorphism Full SI Self-pollen fraction (s) Uyenoyama, Zhang, and Newbigin (2001)

Sn Sb Sa Sn+1 An Bn An Bn+1 An+1 Bn An+1 Bn+1 Direction of pollen flow Uyenoyama, Zhang, and Newbigin (2001)

Sn Sb Sa Sn+1 An Bn An Bn+1 An+1 Bn An+1 Bn+1 TURN OFF Partial breakdown of SI by pollen disablement Sn Evolutionarily unlikely An Bn+1 An+1 Bn Sb Sa TURN ON Restoration of SI by stylar recognition Evolutionarily unlikely An+1 Bn+1 Sn+1 Uyenoyama, Zhang, and Newbigin (2001)

Joint genealogies Unlike S-RNase genes, SLF genes show Solanaceae and Plantaginaceae Rosaceae Unlike S-RNase genes, SLF genes show Low divergence between allelic types No trans-specific sharing of lineages Newbigin, Paape, and Kohn (2008)

Cycles of loss/restoration of SI? Family-specific genealogies Rosaceae: do highly-diverged, ancient SFB lineages reflect continuous operation or restoration of same F-box genes? Solanaceae, Plantaginaceae: Recruitment of new F-box genes? Turnover of pollen-specificity loci Expression and recognition of a paralogue of the former pollen specificity gene? Can homologues be distinguished from paralogues with new function?

Brassica S-locus Natural populations often contain 30 – 50 S-alleles Pollen component Pistil component Natural populations often contain 30 – 50 S-alleles Nasrallah (2000 Curr. Opin. Plant Biol.)

An appeal for inference methods Sexual antagonism in mating type regions Neutral variation in linked regions Rates of substitution at determinants of mating type Inference Goal: use the pattern of variation in population samples of genomic regions as a basis for inference about the evolutionary process Detection genomic conflict and other forms of selection mating systems and population structure

Pollen specificity in SSI Codominance Both specificities expressed Almost twice as many incompatible styles under SSI than GSI for same number of S-alleles Complete dominance One specificity expressed Norbert Holstein http://commons.wikimedia.org/wiki/File:Sporophytic_self-incompatibility.png

SRK genealogies Sporophytic SI Class II: pollen-recessive Diploid genotype of pollen parent determines S-specificity of each pollen grain Class I is dominant over Class II, with codominance within class Class II: pollen-recessive Lower number of segregating alleles, each with relatively higher frequency in population Greater genealogical relationship within class? Edh, Widén and Ceplitis (2009)

Is class II younger than class I? MRCA ages Class I: 25.5 ± 8.1 MY Class II: 3.1 ± 0.9 MY I/II: 41.4 ± 12.7 MY Origin of SLG/SRK system 42.1 ± 9.0 MY Uyenoyama (1995)