ESPE Abstracts

Background: An activation cascade of specific genes sets up the initiation of sex determination leading in males to testes formation and synthesis of testicular hormones. Disruption of this gene cascade may cause a spectrum of disorders/differences of sex development (DSD) phenotypes. Here we describe for the first time two sisters suffering from 46,XY DSD, who by whole exome sequencing were shown to carry a mutation in the X-linked StAR-related lipid transfer domain protein 8 (STARD8) gene. STARD8, also known as deleted in liver cancer 3 (DLC-3) is a functional Rho-specific GAP protein, the loss of which enhances perinuclear Ras homolog gene family member A (RhoA) activity which in turn is known to be involved in SOX9 expression regulation. Additionally, STARD8 downregulation severely disturbs recruitment of β-catenin to sites of cell adhesion, this one being, moreover, a key pro-ovarian and anti-testis signaling molecule.

Objectives: To gain new insights in human sex development mechanisms, we aimed to analyze the functional consequences of STARD8 mutations. Since the STARD8 knockout NMRI mouse model we generated did not recapitulate the human clinical picture, we chose to use another in vivo model to study the mechanisms of disease. Interestingly, Crossveinless-c (Cv-c) the Drosophila homolog of DLC-3/STARD8, has similar location and function than its mammalian counterpart. We therefore chose to study the consequences of Cv-c mutations in the gonadal development of the fruit fly.

Methods: Gonad development was analyzed in cv-c7, cv-cM62 and cv-cC524 alleles using Immunohistochemistry and confocal microscopy to visualize gonad specifics markers. Cv-c expression in the male gonad was confirmed by cv-c fluorescent RNA in situ and Cv-c-GFP TRAP construct.

Results: We found defects affecting the germ cells (GCs) migration from the beginning of embryogenesis with different degrees of severity in the cv-c mutant embryos, preventing gonad coalescence in the most severe cases. We also observed a decrease in the number of GCs in male mutant gonads compared to wild type males.

Conclusions: Our results indicate that cv-c is required for gonadal development in Drosophila embryos, suggesting that the defect in STARD8 is the most likely cause of DSD in our patients. We were able to exploit the fruit fly, Drosophila melanogaster, for functional investigation of findings from human whole exome sequencing, by creating a fly model of a defect in the protein STARD8 found in two patients with 46, XY DSD.