Berthold HEINZE Federal Research Centre for Forests, Department of Genetics, Vienna, Austria

1. Genetic variability of grey poplar hybrids (Populus x canescens = P. alba x P. tremula) and the significance of hybrids for adaptation Berthold HEINZE Federal Research Centre for Forests, Department of Genetics, Vienna, Austria TBX Partner No. 2

2. Natural poplar hybrids • often occur where „compatible“ parent species overlap • show a high diversity of characters • Mendelian segragation beyond F1 • but the parent species do not „collapse“ • they stay distinct species • could natural hybrids contribute to adaptation?

3. Our starting point: phylogeography of 2 hybridizing poplar species in Central Europe • P. tremula and P. alba • contrasting habitats • upland – boreal vs. riverine • hybridizing freely where they meet • backcrosses and introgression • natural clones may persist • our hypothesis: individual genes have a different tendency for introgression – making functional analysis possible

4. Populus alba Populus tremula Populus x canescens

5. flowering phenolgy of both species overlaps in Vienna depending on weather conditions in each year in-situ and ex-situ observations Populus tremula: female and male flowers Populus alba: female flowers and fruits

6. Phylogeography:genetic maps showing phylogenetic relationships different scales of geographic areas require different genetic markers chloroplast DNA molecule often useful for identifying range-wide groupings evolves slowly (many copies in a plant) clonal evolution (no recombination) DNA sequence changes in length (insertions/deletions) or restriction sites detectable and easily analysed

7. A „simple“ genome: the Populus chloroplast functions of ~ 15% of genes still unknown some highly variable regions

8. Technique: PCR-RFLP-electrophoresis • universal primers • database at: http://bfw.ac.at/rz/bfwcms.web?dok=977 • preliminary results from years of experience • restriction analysis • agarose gels • fragment analysis and sequencing on a capillary electrophoresis automat for one fragment

9. Tandem repeat structure in chloroplast gene rpl16 intron • stepwise variation of 16bp • P. alba und P. tremula • visible on agarose gels, but analysed on sequencer

10. tandem repeats in Populus rpl16 intron

11. In Europe, chloroplast lineages often tell something about range-shifts during the glaciations tree populations restricted to southern penninsulas during ice maxima isolated evolution secondary contact after ice retreated today‘s genetic patterns contain information on possible re-colonization routes

12. P. tremula and P. albachloroplast variants in Central Europe • high number of variants (haplotypes), but a few main ones dominate • two groups of haplotypes separate the species • but hybridization and introgression „moves“ some variants into hybrids and into the other species • Italian and Balkan Penninsulas as main sources for P. alba • no clear structure in P. tremula

13. High diversity of chloroplast types typical P. tremula ... but hybrids can have haplotypes from either side (arrows) ... typical P. alba

14. High diversity in Central Europe

15. Two Populus species show different patterns of geographical differentiation in Central and South-Eastern Europe practically no chloroplast DNA differentiation in P. tremula P. alba shows patterns of re-immigration different glacial refugia and secondary contact after / during re-immigration

16. P. tremula types are little differentiated P. albatypesarequitedifferentiated

17. STRUCTURE suggests clear species differentiation,and some population differentiation K=2 K=24 K=24 order by Q

19. is the genetic shape of hybrids a product of these phylogeographic differences only, or do cyto-nuclear interactions interfere? flowering phenology suggest that P. alba can capture chloroplasts from P. tremula more easily are all chromosomes equally affected by cyto-nuclear disequilibrium?

20. Most P x canescens are F2 (NewHybrids) extensive cyto-nuclear interactions as the cyto-nuclear interactions do not affect all chromosomes equally, migration and drift alone are insufficient reasons for this phenomenon * * * * * * * * * * * * * * * * * * * * * * * * * * *

21. Postglacial admixture of P. tremula lineages in Scandinavia • P. tremula in Scandinavia • de Carvalho/Lexer/Ingvarsson, manuscript in press Molec. Ecol. • using 70+ nuclear microsatellites Christian Lexer and Pär Ingvasson

22. Postglacial admixture of P. tremula lineages in Scandinavia Christian Lexer and Pär Ingvasson

24. Clinal variation, but ... • Increased variance for bud set in the centre of the cline • Stronger selection in the centre of the cline • the cline is rather a step than continuous

25. great variation in marker ancestry along e.g. chromosome 6, known to exhibit normal levels of recombination • Yin et al. 2004 TAG • Low variation in marker ancestry along chromosome 19, consistent with reduced recombination • Yin et al. 2008 Genome Res.

26. Conclusions – P. tremula Scandinavia • Adaptive population divergence dectable at the European scale, indicating that recombination is lower or selection greater than previously thought • Postglacial contact zone of P. tremula in N-Europe coincides with those postulated for many other species • Admixture facilitates adaptation from standing variation, as visible from cline shapes, variances, and selection differentials • Great deal of variation for marker ancestry across the genome = great potential of admixture mapping

27. General conclusions • Admixture – most often of post-glacial lineages in Europe – contributes to the adaptation potential of tree species • Admixture of hybridizing species may involve subtle interactions • e.g. cyto-nuclear ones • Populus sp. are an excellent model to study admixture and adaptation

28. Acknowledgements • grants by Austria Academy of Science – Jubiläumsfonds der Stadt Wien, DOC fFORTE (to Barbara Fussi) • sampling support: • Cvrčková H., Máchová P. (FGMRI, CZ), Bartha D. (Sopron, HU) Benke A. (ERTI Sarvar, HU), Bogdan S. (Zagreb, HR), Gonzales-Martinez S. (Madrid, E), Nica M.S., Ipati A., ICAS (RO), and Castiglione S. (Salerno, IT) • lab support: Bianca Widmar, Renate Slunksy, summer students (B. Sokcevic, K. Duran) • thank you for your kind attention!