Evolution of multiple sex-chromosomes associated with dynamic genome reshuffling in Leptidea wood-white butterflies

Sex-chromosome systems tend to be highly conserved and knowledge about their evolution typically comes from macroevolutionary inference. Rapidly evolving complex sex-chromosome systems represent a rare opportunity to study the mechanisms of sex-chromosome evolution at unprecedented resolution. Three cryptic species of wood-white butterflies—Leptidea juvernica, L. sinapis and L. reali—have each a unique set of multiple sex-chromosomes with 3–4 W and 3–4 Z chromosomes. Using a transcriptome-based microarray for comparative genomic hybridisation (CGH) and a library of bacterial artificial chromosome (BAC) clones, both developed in L. juvernica, we identified Z-linked Leptidea orthologs of Bombyx mori genes and mapped them by fluorescence in situ hybridisation (FISH) with BAC probes on multiple Z chromosomes. In all three species, we determined synteny blocks of autosomal origin and reconstructed the evolution of multiple sex-chromosomes. In addition, we identified W homologues of Z-linked orthologs and characterised their molecular differentiation. Our results suggest that the multiple sex-chromosome system evolved in a common ancestor as a result of dynamic genome reshuffling through repeated rearrangements between the sex chromosomes and autosomes, including translocations, fusions and fissions. Thus, the initial formation of neo-sex chromosomes could not have played a role in reproductive isolation between these Leptidea species. However, the subsequent species-specific fissions of several neo-sex chromosomes could have contributed to their reproductive isolation. Then, significantly increased numbers of Z-linked genes and independent neo-W chromosome degeneration could accelerate the accumulation of genetic incompatibilities between populations and promote their divergence resulting in speciation.This research was funded by grant 14-22765S and follow-up grant 17-13713S of the Czech Science Foundation (CSF). Publishing was supported by CSF grant 20-13784S given to FM. PN was supported by CSF grants 17-17211S and 20-20650Y. RV acknowledges support from project CGL2016-76322-P (AEI/FEDER, UE).Peer reviewe

Dynamic karyotype evolution and unique sex determination systems in Leptidea wood white butterflies

Background. Chromosomal rearrangements have the potential to limit the rate and pattern of gene flow within and between species and thus play a direct role in promoting and maintaining speciation. Wood white butterflies of the genus Leptidea are excellent models to study the role of chromosome rearrangements in speciation because they show karyotype variability not only among but also within species. In this work, we investigated genome architecture of three cryptic Leptidea species (L. juvernica, L. sinapis and L. reali) by standard and molecular cytogenetic techniques in order to reveal causes of the karyotype variability./nResults. Chromosome numbers ranged from 2n = 85 to 91 in L. juvernica and 2n = 69 to 73 in L. sinapis (both from Czech populations) to 2n = 51 to 55 in L. reali (Spanish population). We observed significant differences in chromosome numbers and localization of cytogenetic markers (rDNA and H3 histone genes) within the offspring of individual females. Using FISH with the (TTAGG)n telomeric probe we also documented the presence of multiple chromosome fusions and/or fissions and other complex rearrangements. Thus, the intraspecific karyotype variability is likely due to irregular chromosome segregation of multivalent meiotic configurations. The analysis of female meiotic chromosomes by GISH and CGH revealed multiple sex chromosomes: W1W2W3Z1Z2Z3Z4 in L. juvernica, W1W2W3Z1Z2Z3 in L. sinapis and W1W2W3W4Z1Z2Z3Z4 in L. reali./nConclusions. Our results suggest a dynamic karyotype evolution and point to the role of chromosomal rearrangements in the speciation of Leptidea butterflies. Moreover, our study revealed a curious sex determination system with 3–4 W and 3–4 Z chromosomes, which is unique in the Lepidoptera and which could also have played a role in the speciation process of the three Leptidea species.This research was funded by Grant 14-22765S of the Czech Science Foundation, Grant IAA600960925 of the Grant Agency of The Czech Academy of Sciences, Grant 063/2012/P of the Grant Agency of the University of South Bohemia and Grant CGL2013-48277-P from the Spanish Ministerio de Economía y Competitividad. JŠ and AV were supported by Grant 052/2013/P of the Grant Agency of the University of South Bohemia, VD by a Marie Curie International Outgoing Fellowship within the 7th European Community Framework Programme (project no. 625997), PN by Grant 14-35819P of the Czech Science Foundation and KS by JSPS 23380030 grant and JSPS Excellent Young Researchers Overseas Visit Program (21–7147