Polymorphisms within promoter of Japanese flounder (Paralichthys olivaceus) ovary cytochrome P450-c19 (CYP19a) gene associated with reproductive traits.

Cytochrome P450 aromatase, which is encoded by the CYP19a gene, converts androgens to estradiol. Considerable evidence suggests that estrogens play an important role in fish reproductive process. Therefore CYP19a is an excellent candidate gene for reproductive traits. Variants in the promoter of the CYP19a gene might also be involved in the control of aromatase expression and affect regulatory mechanism linking cholesterol metabolism to the synthesis of sex steroids. In this study, nine single-nucleotide polymorphisms (SNPs) were detected with polymerase chain reaction-single stranded conformational polymorphism (PCR-SSCP), namely A-680G, G-672A, AGTAGT-649 inserting or deleting, T-623C, C-410A, T7-454A, T-402C, TTTCCAGACTGA-345 inserting or deleting, and G-297C. Nine SNPs within the promoter of the CYP19a gene were tested for association with four reproductive traits [serum testosterone (T), serum 17beta-estradiol (E(2)), hepatosomatic index (HSI), and gonadosomatic index (GSI)] in a population of 50 female Japanese flounder individuals. A locus, P3 (TTTCCAGACTGA-345 inserting or deleting, G-297C), was significantly associated with 17beta-estradiol (E(2)) level (P < 0.05) in female Japanese flounder. In addition, there was significant association between one diplotype based on nine SNPs and reproductive trait. The genetic effect for E(2) level of diplotype D3 was significantly higher than those of other diplotypes (P < 0.05). Results indicate that these genetic effects of those variants on E(2) level may help to explain CYP19a gene status in the reproductive endocrinology of Japanese flounder.

Single nucleotide polymorphisms within the estrogen receptor beta gene are linked with reproductive indices in Japanese flounder, Paralichthys olivaceus.

The objectives of this study were to characterize polymorphisms within the coding region of estrogen receptor beta (ERbeta) gene in a population of 57 female Japanese flounder (Paralichthys olivaceus) and to analyze the association of ERbeta polymorphisms with reproductive indices by polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP). Two single nucleotide polymorphisms (SNPs), SNP1 (c.577delC) and SNP2 [c.A891T (p.Gln114Leu)], were identified in the ERbeta gene. A one-way ANOVA revealed that SNP1 was significantly associated with the gonadosomatic index (GSI) in female Japanese flounder (P < 0.05). And SNP2 was significantly associated with the serum 17beta-estradiol (E2) level and GSI (P < 0.05). Individuals with genotype AB of SNP2 had significantly higher serum E2 level and GSI than those of genotype AA (P < 0.05). Moreover, the hepatosomatic index (HSI), a marker for genetic effects, was significantly higher for diplotype D2 compared with the other three diplotypes (P < 0.05). These results obtained in this study suggested that SNP2 could influence reproductive endocrinology of female Japanese flounder and be useful as a potential candidate genetic marker for the selection of reproductive indices in female Japanese flounder.

Identification of estrogen receptor alpha gene polymorphisms by SSCP and its effect on reproductive traits in Japanese flounder (Paralichthys olivaceus).

Estrogen receptor (ERalpha) modifies the expression of genes involved in cell growth, proliferation and differentiation through binding to estrogen response elements (EREs) located in a number of gene promoters, so the ERalpha gene is considered as an important factor affecting reproductive endocrinology in Japanese flounder (Paralichthys olivaceus). In this study, twelve single nucleotide polymorphisms (SNPs) within eight CDS exons and 1 kb of 3'-UTR of the ERalpha gene were tested to association with four reproductive traits in a population of 119 Japanese flounder individuals with polymerase chain reaction-single stranded conformational polymorphism (PCR-SSCP). The association analysis of SNPs within Japanese flounder ERalpha gene with the reproductive traits was carried out using General Linear Model (GLM) estimation. Results indicated that two SNPs in the exon4 of ERalpha gene, P1 (A803G and C864T), were significantly associated with hepatosomatic index (HSI) (P<0.05) in female Japanese flounder. Other ten SNPs in 3'-UTR associated to serum 17beta-estradiol (E(2)) and HSI showed that P2 (A1982T) was significantly associated with E(2) (P<0.01) and P3 (A2149G, 2181TTACAAG2182 insertion or deletion, T2324G, A2359G and G2391A) was significantly associated with HSI (P<0.05) in female Japanese flounder. However, P2 (A1982T) and P4 (G2256T, T2294C, T2309G and A2333T) had significant effects on E(2) (P<0.05 and P<0.01, respectively) in male Japanese flounder. In addition, there were significant associations between diplotype D1 based on fourteen SNPs and reproductive traits. The genetic effects for HSI (female) or E(2) (male) of diplotype D1 were significantly higher than those of other eight diplotypes (P<0.05), respectively. Our findings implied that P1 of ERalpha gene affecting the reproductive traits could be a potential QTN (quantitative trait nucleotide) which would be useful genetic marker in the selection of some reproductive traits for its in Japanese flounder.