89 MALE-TO-FEMALE SEX REVERSAL IN WOLF DERIVED FROM SOMATIC CELL NUCLEAR TRANSFER WITH 78,XY KARYOTYPE

Abstract

Normal sexual differentiation relies on a complex cascade of numerous genes, many of which have yet to be identified. Gene defects can cause disorders of sexual development with varying severity. Among them, SRY (sex-determining region on Y chromosome) genes are involved in the early steps of maleness determination. In human, mutations of the SRY gene have been diagnosed in up to 20% cases of females with XY karyotype. There have been a few instances of intersexuality such as true hermaphrodites or pseudo hermaphrodites, but no reports yet in canid of XY male-to-female sex reversal. Recently, we reported the cloning of endangered male gray wolves (Canis lupus) from somatic cells collected postmortem (Oh HJ et al. 2008 Theriogenology). In this study, the 3 wolves were delivered with normal karyotype (78,XY), but 1 wolf had female external genitalia such as a vulva and clitoris. For further diagnosis, we conducted macroscopic, radiologic, surgical, histological, and cytogenetic study. On gonadotropin-releasing hormone stimulation test, the blood serum testosterone concentration was low (<1 ng mL–1). Imaging diagnosis by radiography in the form of uterogram and by ultrasonography revealed a normal uterus and bilateral ovary-like structures located caudal to the kidney. During exploratory laparotomy, we found a normal uterus and bilateral ovary-like mass. Histopathological analysis showed the uterus and ovary to be normal, but no evidence of the presence of testicular tissue. The molecular analysis of the SRY genes with a direct DNA sequencing technique detected nucleotide changes more than one position in this affected animal compared with that of the control groups (cell donor and other female litter mates) although at a low rate. However, we did not yet analyze the nature of this mutation such as amino acid change, location within gene, protein function, and so on, so it will be needed further studies. These results indicate that this abnormal sex development of the studied wolf was probably caused by SRY gene mutations. Herein, we reported the case of a cloned wolf with normal ovaries and uterus having a 78,XY karyotype. Further studies are required to elucidate whether this mutation in the SRY gene could impair the normal function of the SRY protein. This study was financially supported by the Korean MEST, through KOSEF (grant # M10625030005-08N250300510) and the BK21 program for Veterinary Science, and SNU foundation(Benefactor; RNL BIO)/Hanwha L&C.