|Abstract:||Sex chromosomes evolved independently and repeatedly animals and plants. Although considerable differences existed among diverse lineages in terms of sexual dimorphism and genetic basis of sex determination, the underlining principles and evolutionary forces that lead to the formation and progression of sex chromosomes are the same in plants and animals. The well-established theories and observations indicate that sex chromosomes in birds and mammals are ancient, but sex chromosomes in fish, amphibians, and plants are more recent. Studying those early-stage sex chromosomes could shed light on the inception and forces that shape sex chromosome evolution. We selected garden spinach, an annual leafy dioecious vegetable with a pair of nascent sex chromosomes to study sex chromosome evolution.
Based on a theory of sex chromosome evolution, sex chromosomes evolved from autosomes by genetic mutations relate to the female and male reproductive organ development. Suppression of recombination at the sex determination locus or loci is the pivotal point of sex chromosome evolution. As consequences of ceased recombination at the sex-determining region, accumulation of repetitive sequences, chromosomal rearrangements, and gene mutations occurred, accompanied with a strong selection due to a small effective population size of genes on the Y chromosome compared with autosomes. Genomic localization of the non-recombining region is critical for studying spinach sex chromosome evolution.
To identify the region associated with sex determination in spinach, I developed DNA markers to test 40 spinach accession from different countries. I found that a small 1.78 Mb region is featured by strong linkage with sex type and it is conserved across 40 different spinach accessions by genotyping plants with 12 DNA markers. However, a large-scale test by genome-wide polymorphic markers is needed to map the position of the region and estimate the size and numbers of genes within this sex co-segregation genomic region.
We re-sequenced the genomes of progenies from a segregating pseudo-test cross mapping population and used genome-wide DNA markers to construct two genetic maps, one for each parent, to identify this region. The two high-density linkage maps anchored a total of 40,324 SNPs for female map and 256,636 SNPs for male map with highly consensus orders with each other. The sex determination region on the Y chromosome was mapped to the largest linkage group at 45.18cM locus by male map, comprised 4,567 sex co-segregating SNPs. The two maps substantially improve resolution of the sex determination region, which also enabled a higher quality female genome assembly by anchoring contigs to a published reference genome. However, we need to generate sequences for both the complete genomic region of the Y chromosome and its X corresponded part to further understand the evolutionary forces related to sex chromosome evolution.
The viable YY spinach overcomes the difficulties of assembling heterozygous XY haplotypes. De-novo assembly facilitated by Hi-C and linkage maps of two new genomes (XX and YY) enabled us to define the male-specific region (MSY) and its X counterpart. Surprisingly, the comparison revealed a 39.26 Mb MSY and 38.92 Mb X counterpart, features by two inversions within the region along with gene loss and structural variations between paired genes from two sex types. A 6 Mb small region with a continuous pattern of gene divergence between X and Y chromosome further divide MSY into three evolutionary strata dated from 0.1-0.2 million years ago.
Together, these results expanded our knowledge on sex chromosome evolution in dioecious spinach. The knowledge and genomic resources generated can be applied to identify sex-determining genes to decipher sex determination and differentiation gene networks and improve efficiency and shorten cycles of spinach breeding.