Integrated gene mapping and synteny studies give insights into the evolution of a sex proto-chromosome in Solea senegalensis

Abstract

The evolution of genes related to sex and reproduction in fish shows high plasticity and, to date, the sex determination system has only been identified in a few species. Solea senegalensis has 42 chromosomes and an XX/XY chromosome system for sex determination, while related species show the ZZ/ZW system. Next-generation sequencing (NGS), multi-color fluorescence in situ hybridization (mFISH) techniques, and bioinformatics analysis have been carried out, with the objective of revealing new information about sex determination and reproduction in S. senegalensis. To that end, several bacterial artificial chromosome (BAC) clones that contain candidate genes involved in such processes (dmrt1, dmrt2, dmrt3, dmrt4, sox3, sox6, sox8, sox9, lh, cyp19a1a, amh, vasa, aqp3, and nanos3) were analyzed and compared with the same region in other related species. Synteny studies showed that the co-localization of dmrt1-dmrt2-drmt3 in the largest metacentric chromosome of S. senegalensis is coincident with that found in the Z chromosome of Cynoglossus semilaevis, which would potentially make this a sex proto-chromosome. Phylogenetic studies show the close proximity of S. senegalensis to Oryzias latipes, a species with an XX/XY system and a sex master gene. Comparative mapping provides evidence of the preferential association of these candidate genes in particular chromosome pairs. By using the NGS and mFISH techniques, it has been possible to obtain an integrated genetic map, which shows that 15 out of 21 chromosome pairs of S. senegalensis have at least one BAC clone. This result is important for distinguishing those chromosome pairs of S. senegalensis that are similar in shape and size. The mFISH analysis shows the following co-localizations in the same chromosomes: dmrt1-dmrt2-dmrt3, dmrt4-sox9-thrb, aqp3-sox8, cyp19a1a-fshb, igsf9b-sox3, and lysg-sox6.

Sex Chromosome Evolution and Genomic Divergence in the Fish Hoplias malabaricus (Characiformes, Erythrinidae)

“…21, could disable the detection of the hypothetical sex-determining region in its vicinity. Maybe a finer-scale approach such as BAC FISH mapping of specific candidate genes identified based on utilization of recent genome sequencing approaches and corresponding bioinformatic tools, togehter with other related “state-of-the-art” technologies may shed more light on this issue (for examples in fishes, see: Reichwald et al, 2015 ; Portela-Bens et al, 2017 ; Sutherland et al, 2017 ; Liu et al, 2018 ).…”

In Silico Detection and FISH Analysis to Determine Location of miRNAs in Solea Senegalensis Chromosomes Using BACs

“…Most of the groups (26 of 45) could not be assigned to a particular chromosome, but 8, 3, and 2 groups were assigned to chromosomes 1, 4, and 6, respectively. For other chromosomes (12,16,18,19, and 21) only one group was observed. Finally, one group of miRNAs corresponded to a BAC that hybridizes with multiple chromosomes.…”

“…BAC-FISH data was obtained from previous works [18,23,31] and by employing the FISH techniques described in [31]. All applicable international, national, and/or institutional guidelines for the care and use of animals were followed.…”

“…To assign miRNAs to a particular chromosome, BAC-FISH data was employed [18,23,31]. The BAC library was screened for genes of interest and then positive clones were sequenced and annotated.…”

“…Nevertheless, the existence of sexual dysfunction of males reared in captivity, high larval mortality, and diseases have hampered its production [16]. In recent years, an increasing number of studies have assessed different genetic and genomic aspects of S. senegalensis such as transcriptomics [17], integration of genetic maps [18], and cytogenomics [19]. There are also various studies assessing gene expression [20], but to date only one study has explored miRNA sequences in S. senegalensis [21].…”