The existence of males and females, which are often strikingly different in morphology, reproductive strategies and behavior, is one of the most widespread phenomena in biology. However, the genetic mechanisms that generate this ubiquitous pattern are surprisingly diverse and do not follow a phylogenetic pattern. Sex-determination mechanisms can differ between even closely related species and arise frequently and independently. Fish provide a paradigmatic example, as their sex-determination mechanisms range from environmental to different modes of genetic determination. The evolutionary meaning of this remarkable plasticity is unknown. For genetic sex determination, where the trigger for female or male development comes from the genetic constitution of the individual, the evolution of sex-determination mechanisms is connected to a very peculiar genomic process, namely the formation of sex chromosomes [1][2][3][4] .To improve understanding of the function and evolution of sex chromosomes, their genetic organization must be deciphered.However, owing to their degenerate nature and high repetitive DNA content, sex chromosomes pose almost insurmountable problems in deciphering their gene content and organization. So far, only the human 5 , chimpanzee 6 and rhesus macaque Y chromosomes 7 and the male-specific region on the Y chromosome of one fish, the medaka 8 , have been sequenced. These analyses have nevertheless provided important insights into the evolution of Y chromosomes, their genomic organization and their degeneration processes, as well as predictions as to their likely evolutionary fate 9-12 .Much less genomic information exists on W chromosomes because, as with Y chromosomes, they are predominantly highly repetitive in nature. The prevailing theory of the evolution of sex chromosomes predicts that degeneration of the heterogametic sex chromosome is a stepwise process that occurs over an extended period of time. We therefore reasoned that an evolutionarily young W chromosome Whole-genome sequence of a flatfish provides insights into ZW sex chromosome evolution and adaptation to a benthic lifestyle Genetic sex determination by W and Z chromosomes has developed independently in different groups of organisms. To better understand the evolution of sex chromosomes and the plasticity of sex-determination mechanisms, we sequenced the whole genomes of a male (ZZ) and a female (ZW) half-smooth tongue sole (Cynoglossus semilaevis). In addition to insights into adaptation to a benthic lifestyle, we find that the sex chromosomes of these fish are derived from the same ancestral vertebrate protochromosome as the avian W and Z chromosomes. Notably, the same gene on the Z chromosome, dmrt1, which is the male-determining gene in birds, showed convergent evolution of features that are compatible with a similar function in tongue sole. Comparison of the relatively young tongue sole sex chromosomes with those of mammals and birds identified events that occurred during the early phase of sex-chromosome evolution. Pertinent to...

Environmental sex determination (ESD) occurs in divergent, phylogenetically unrelated taxa, and in some species, cooccurs with genetic sex determination (GSD) mechanisms. Although epigenetic regulation in response to environmental effects has long been proposed to be associated with ESD, a systemic analysis on epigenetic regulation of ESD is still lacking. Using half-smooth tongue sole (Cynoglossus semilaevis) as a model-a marine fish that has both ZW chromosomal GSD and temperature-dependent ESD-we investigated the role of DNA methylation in transition from GSD to ESD. Comparative analysis of the gonadal DNA methylomes of pseudomale, female, and normal male fish revealed that genes in the sex determination pathways are the major targets of substantial methylation modification during sexual reversal. Methylation modification in pseudomales is globally inherited in their ZW offspring, which can naturally develop into pseudomales without temperature incubation. Transcriptome analysis revealed that dosage compensation occurs in a restricted, methylated cytosine enriched Z chromosomal region in pseudomale testes, achieving equal expression level in normal male testes. In contrast, female-specific W chromosomal genes are suppressed in pseudomales by methylation regulation. We conclude that epigenetic regulation plays multiple crucial roles in sexual reversal of tongue sole fish. We also offer the first clues on the mechanisms behind gene dosage balancing in an organism that undergoes sexual reversal. Finally, we suggest a causal link between the bias sex chromosome assortment in the offspring of a pseudomale family and the transgenerational epigenetic inheritance of sexual reversal in tongue sole fish.

9 l e t t e r sFlatfish have the most extreme asymmetric body morphology of vertebrates. During metamorphosis, one eye migrates to the contralateral side of the skull, and this migration is accompanied by extensive craniofacial transformations and simultaneous development of lopsided body pigmentation 1-5 . The evolution of this developmental and physiological innovation remains enigmatic. Comparative genomics of two flatfish and transcriptomic analyses during metamorphosis point to a role for thyroid hormone and retinoic acid signaling, as well as phototransduction pathways. We demonstrate that retinoic acid is critical in establishing asymmetric pigmentation and, via cross-talk with thyroid hormones, in modulating eye migration. The unexpected expression of the visual opsins from the phototransduction pathway in the skin translates illumination differences and generates retinoic acid gradients that underlie the generation of asymmetry. Identifying the genetic underpinning of this unique developmental process answers long-standing questions about the evolutionary origin of asymmetry, but it also provides insight into the mechanisms that control body shape in vertebrates.

Chinese tongue sole is a marine fish with ZW sex determination. Genome sequencing suggested that the Z-linked dmrt1 is a putative male determination gene, but direct genetic evidence is still lacking. Here we show that TALEN of dmrt1 efficiently induced mutations of this gene. The ZZ dmrt1 mutant fish developed ovary-like testis, and the spermatogenesis was disrupted. The female-related genes foxl2 and cyp19a1a were significantly increased in the gonad of the ZZ dmrt1 mutant. Conversely, the male-related genes Sox9a and Amh were significantly decreased. The dmrt1 deficient ZZ fish grew much faster than ZZ male control. Notably, we obtained an intersex ZW fish with a testis on one side and an ovary on the other side. This fish was chimeric for a dmrt1 mutation in the ovary, and wild-type dmrt1 in the testis. Our data provide the first functional evidence that dmrt1 is a male determining gene in tongue sole.

High-density genetic linkage maps of half-smooth tongue sole were developed with 1007 microsatellite markers, two SCAR markers and an F1 family containing 94. The female map was composed of 828 markers in 21 linkage groups, covering a total of 1447.3 cM, with an average interval 1.83 cM between markers. The male map consisted of 794 markers in 21 linkage groups, spanning 1497.5 cM, with an average interval of 1.96 cM. The female and male maps had 812 and 785 unique positions, respectively. The genome length of half-smooth tongue sole was estimated to be 1527.7 cM for the females and 1582.1 cM for the males. Based on estimations of the map lengths, the female and male maps covered 94.74 and 94.65% of the genome, respectively. The consensus map was composed of 1007 microsatellite markers and two SCAR markers in 21 linkage groups, covering a total of 1624 cM with an average interval of 1.67 cM. Furthermore, 159 sex-linked SSR markers were identified. Five sex-linked microsatellite markers were confirmed in their association with sex in a large number of individuals selected from different families. These sex-linked markers were mapped on the female map LG1f with zero recombination. Two QTLs that were identified for body weight, designated as We-1 and We-2, accounted for 26.39% and 10.60% of the phenotypic variation. Two QTLs for body width, designated Wi-1 and Wi-2, were mapped in LG4f and accounted for 14.33% and 12.83% of the phenotypic variation, respectively. Seven sex-related loci were mapped in LG1f, LG14f and LG1m by CIM, accounting for 12.5–25.2% of the trait variation. The results should prove to be very useful for improving growth traits using molecular MAS.

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Background: Edwardsiella tarda causes acute symptoms with ascites in Japanese flounder (Paralichthys olivaceus) and is a major problem for China's aquaculture sector. Genomic selection (GS) has been widely adopted in breeding industries because it shortens generation intervals and results in the selection of individuals that have great breeding potential with high accuracy. Based on an artificial challenge test and re-sequenced data of 1099 flounders, the aims of this study were to estimate the genetic parameters of resistance to E. tarda in Japanese flounder and to evaluate the accuracy of single-step GBLUP (ssGBLUP), weighted ssGBLUP (WssGBLUP), and BayesB for improving resistance to E. tarda by using three subsets of pre-selected single nucleotide polymorphisms (SNPs). In addition, SNPs that are associated with this trait were identified using a single-SNP genome-wide association study (GWAS) and WssGBLUP. Results: We estimated a heritability of 0.13 ± 0.02 for resistance to E. tarda in Japanese flounder. One million SNPs at fixed intervals were selected from 4,978,724 SNPs that passed quality controls. GWAS identified significant SNPs on chromosomes 14 and 24. WssGBLUP revealed that the putative quantitative trait loci on chromosomes 1 and 14 contained SNPs that explained more than 1% of the genetic variance. Three 50 k-SNP subsets were pre-selected based on different criteria. Compared with pedigree-based prediction (ABLUP), the three genomic methods evaluated resulted in at least 7.7% greater accuracy of predictions. The accuracy of these genomic prediction methods was almost unchanged when pre-selected trait-related SNPs were used for prediction. Conclusions: Resistance to E. tarda in Japanese flounder has a low heritability. GWAS and WssGBLUP revealed that the genetic architecture of this trait is polygenic. Genomic prediction of breeding values performed better than ABLUP. It is feasible to implement genomic selection to increase resistance to E. tarda in Japanese flounder with 50 k SNPs. Based on the criteria used here, pre-selection of SNPs was not beneficial and other criteria for pre-selection should be considered.

The Japanese flounder is one of the most widely farmed economic flatfish species throughout eastern Asia including China, Korea, and Japan. Edwardsiella tarda is a major species of pathogenic bacteria that causes ascites disease and, consequently, a huge economy loss for Japanese flounder farming. After generation selection, traditional breeding methods can hardly improve the E. tarda resistance effectively. Genomic selection is an effective way to predict the breeding potential of parents and has rarely been used in aquatic breeding. In this study, we chose 931 individuals from 90 families, challenged by E. tarda from 2013 to 2015 as a reference population and 71 parents of these families as selection candidates. 1,934,475 markers were detected via genome sequencing and applied in this study. Two different methods, BayesCπ and GBLUP, were used for genomic prediction. In the reference population, two methods led to the same accuracy (0.946) and Pearson's correlation results between phenotype and genomic estimated breeding value (GEBV) of BayesCπ and GBLUP were 0.912 and 0.761, respectively. In selection candidates, GEBVs from two methods were highly similar (0.980). A comparison of GEBV with the survival rate of families that were structured by selection candidates showed correlations of 0.662 and 0.665, respectively. This study established a genomic selection method for the Japanese flounder and for the first time applied this to E. tarda resistance breeding.