Y chromosome divergence despite ancient origin of dioecy in poplars ( Populus ) 1 2

a 5 # Equal contribution 6 Departments of a Botany, 19 Supplementary information: All supplementary files will be made available upon request. 20 Data Accessibility: All sequence data will be deposited on Genbank and SRA and all 21 other data files on Dryad. 22 23 peer-reviewed) is the author/funder. All rights reserved. No reuse allowed without permission. Abstract 24 All species of the genus Populus (poplar, aspen) are dioecious, suggesting an 25 ancient origin of this trait. Theory suggests that non-recombining sex-linked regions 26 should quickly spread, eventually becoming heteromorphic chromosomes. In contrast, we 27 show using whole genome scans that the sex-associated region in P. trichocarpa is small 28 and much younger than the age of the genus. This indicates that sex-determination is 29 highly labile in poplar, consistent with recent evidence of "turnover" of sex determination 30 regions in animals. We performed whole genome resequencing of 52 Populus 31 trichocarpa (black cottonwood) and 34 P. balsamifera (balsam poplar) individuals of 32 known sex. Genome-wide association studies (GWAS) in these unstructured populations 33 identified 650 SNPs significantly associated with sex. We estimate the size of the sex-34 linked region to be ~100 Kbp. All significant SNPs were in strong linkage disequilibrium 35 despite the fact that they were mapped to six different chromosomes (plus 3 unmapped 36 scaffolds) in version 2.2 of the reference genome. We show that this is likely due to 37 genome misassembly. The segregation pattern of sex associated SNPs revealed this to be 38 an XY sex determining system. Estimated divergence times of X and Y haplotype 39 sequences (6-7 MYA) are much more recent than the divergence of P. trichocarpa 40 (poplar) and P. tremuloides (aspen). Consistent with this, in P. tremuloides we found no 41 XY haplotype divergence within the P. trichocarpa sex-determining region. These two 42 species therefore have a different genomic architecture of sex, suggestive of at least one 43 turnover event in the recent past. 44 45 peer-reviewed) is the author/funder. All rights reserved. No reuse allowed without permission.