SOX9 AND DMRT1 EXPRESSION IN MALE AND FEMALE GONADS OF Apareiodon sp. (CHARACIFORMES, PARODONTIDAE)

Phenotypic sex is manipulated easily by hormonal treatment in aquaculture; however the molecular mechanism of sex determination is poorly understood in fishes. Studies have demonstrated that several genes (sox9a, dmrt1, amh, wt1, ftz-f1, gata) have previously been associated to this process in zebrafish, medaka, groupers, tilapia and rainbow trout. Sox9 is transcription factor and is critical for testis determination and chondrogenesis in vertebrates. However, the expression of Sox9 in gonad fishes seem to be somewhat diversified. DMRT (doublessex and mab-3 related transcription factor) family gene is widely conserved from invertebrates to humans. Vertebrate Dmrt1 (DM related transcription factor 1) gene expression occurs predominantly in testis and is a good candidate for male sex-determining gene studies. Parodontidae Neotropical fishes are a good model for investigating sex differentiation because in this group there are species in different stages of sex chromosome differentiation (without sex chromosome, sex proto-chromosome, ZZ/ZW and ZZ/ZW₁W₂). This study was conducted to determine the difference Sox9 and Dmrt1 expression between adult male and female of Apareiodon sp. (ZZ/ZW - Parodontidae group). In this respect, the understanding of function of Sox9 and Dmrt1 in sex determination is important for use in aquaculture.

A total of four adult males and four adult females of Apareiodon sp. were obtained from Laranjinha River, Cinzas Basin, PR, Brazil. Total RNA was isolated from the gonads using Trizol according to the manufacturer's instructions. Gene expression was quantified by qRT-PCR. The 18S rDNA was used as an internal control. The threshold cycle (Ct) was measured and the relative change in the expression level in male and female comparative of Sox9 and Dmrt1 genes was presented as 2^-ΔΔCt. Statistical analysis was performed using Student's t-test (P<0.05) using Action software.

The expression of Sox9 in Apareiodon sp. was observed in adult testis and ovary but the levels were not different between sexes (Figure 1a). However Dmrt1 expression was 23-fold significantly (P<0.05) bigger in male gonad compared to the female gonad (Figure 1b). Thus, Dmrt1 exhibited a male specific expression suggesting an important role for this gene in spermatogenesis.