1. Ecological speciation of a hybrid sunflower, Helianthus anomalus Yuval Sapir, Loren H. Rieseberg Dept. of Biology, Indiana University, Bloomington, IN 47405, USA

2. Pre-/post-zygotic isolating barriers - reduce gene-flow. Niche shift promotes genomic differentiation (selection). Selection against unfit hybrid progeny (reinforcement). Ecological speciation = Speciation facilitated by divergent selection in contrasting environments

3. Hybridization: Increases genetic variation and creates new combinations Complementary gene action Simultaneous change at multiple traits/genes Role of hybridization in ecological speciation If hybrid colonize different ecological habitat Habitat-specific ecological selection promote hybrid speciation

5. Helianthus annuus Helianthus petiolaris Helianthus anomalus

6. Distribution of hybrids & parental spp in the US

7. Methods 96 individuals from each parental species (3 populations each) 192 individuals from 4 populations of Helianthus anomalus DNA extraction Microsatellites markers from EST library Preliminary survey  49 loci amplified in all 3 spp were chosen

8. Indices for detecting selection footprint Schlötterer C., 2002. Genetics 160:753-763 Schlötterer C. & Dieringer, D., 2005. in: Nurminsky D. (ed) Selective Sweep. Landes Bioscience, Georgetown, TX, pp 55-64 Genetic diversity - Genetic variance - H =gene diversity = expected heterozigosity

9. Reduction in genetic variance or genetic diversity Selective sweeps = hitch-hiking effect Footprint of selection Example from: Harr et al., 2002. PNAS 99(20):12949-12954 non-significant effect (95% interval) significant reduction (P<0.05)

10. lnRHlnRV 1822ano/ann 2022ano/pet 2326ano/PSP PSP = pooled data from Parental Species; ano = H. anomalus ; ann = H. annuus ; pet = H. petiolaris 12 loci showed significant effect in both allele variance and genetic diversity Results

11. lnRV and lnRH distributions lnRV=difference in allele variance; lnRH=difference in gene diversity

12. lnRV and lnRH distributions lnRV=difference in allele variance; lnRH=difference in gene diversity

13. Reduced gene diversity and shared alleles

14. Gene function – locus HT424 as an example lnRH = -2.47; 4 alleles in H. anomalus; 10 in parents Proportion of shared alleles: 0.85 Sequence: AACTTATACAATGTAGAGATATGGTTGTTCCAAAGAAGATTCAACGTAGTAATCCTTATGAAGAGCGTGTTCCTGTCGGA TTGGCTATTGTTGCAGCAGTCATAGTTCGGTTACAGCTTGCAAATACTGCTGCTGGTGGAATTGAAACACATGTTGGTGG TTCTATGTTAGTTGATGTGGTTAACAGCTCGTGGCTGCAAGTTATTCTAGCCGGTGTTACTTGGTATCTCATTGGTCTGGC TGCGGTTGAGATTATAGCAGTCCTTCGAATAAAATCGAACGATATTGAAGAATGAATAAAATAGTATATATATACAGGTAT GTAGTATGTATAGGTTAAGCTGTTATAGGAATCATATATTCATTATTATATACACTCTCGAAAGAAAGCTCAGTCAAGCGTA TGTTCATTACCCCAAAAAAAATCTTTAAAGTCGGAACACAAAAAATGTCGGGGGAAAAAAAA Putative translation: LIQCRDMVVPKKIQRSNPYEERVPVGLAIVAAVIVRLQLANTAAGGIETHVGGSMLVDVVNSSWLQVILAGVTWYLIGLAA VEIIAVLRIKSNDIEE* BLAST results: DNAJ heat shock N- terminal domain- containing protein [Arabidopsis thaliana] Interact with heat-shock proteins Can function as chaperones on their own Role in targeting and folding of polypeptides Regulation of gravitropism

15. Summary and conclusion: Twelve of 49 loci exhibited significant reduction in variation compared to both parental species. The proportion of shared alleles is correlated with reduced variation. Genes exhibit footprint of selection may contribute to drought tolerance, heat tolerance, or deepening the root. Selection on ecologically important genes allowed H. anomalus to colonize desert sand dunes.

16. Future directions Finer-scale hitch-hiking mapping to determine size of genomic regions affected by selective sweep Sequence analyses to confirm history of selection Functional analyses (correlations with QTLs, gene expression studies, transformations)

17. University of Georgia Beau Brouillette Lisa Donovan Thanks Rieseberg lab, IU Amanda Posto Briana Gross Nolan Kane Rebecca Randel Yoko Kakugawa Zhao Lai $upported by: BARD-Vaadia post-doctoral fellowship University of Veterinary Medicine, Vienna Christian Schlötterer Daniel Dieringer

19. Kruskal-Wallis Test:  2 2 =45.95; P<0.001; Paired tests - Wilcoxon Sign Ranked Proportion of shared alleles with parental species