Genotype-phenotype correlations among patients with 46,XY disorders of sex development carrying SF1 mutations (#28)
Steroidogenic Factor 1 (SF1) is an orphan nuclear receptor involved in reproduction and steroidogenesis. SF1 mutations can lead to adrenal failure and/or 46,XY disorders of sex development (DSD), where phenotypes range from mild forms (hypospadias) to moderate forms (ambiguous genitalia) to severe forms (complete gonadal dysgenesis and female genitalia). While its steroidogenic roles are well-studied, the function of SF1 in human sex determination is poorly understood. In mice, SF1 initiates expression of SOX9 [SRY (sex determining region Y)-box 9] via a testis-specific enhancer (TESCO). SOX9 expression is then upregulated and maintained by synergistic interactions between SF1-SRY and SF1-SOX9 on TESCO respectively, enabling normal development of the testis. We hypothesised that SF1 mutations in 46,XY DSD patients affect SOX9 expression via TESCO. We aimed to elucidate the sex determining function of SF1 by assessing whether SF1mutants found in 20 46,XY DSD patients can activate TESCO. By performing in vitro reporter assays with a TESCO-luciferase construct and wild-type or mutant SF1, either alone or in combination with SRY or SOX9, we found that SF1 mutants showed defective activation of TESCO in 15 out of 20 cases. Synergistic activation of TESCO by SF1-SRY and SF1-SOX9 was also impaired. Phenotype-genotype correlation was observed as TESCO activity was proportional to phenotype severity. SF1 protein structure analysis revealed that the mutations altered amino acids critical for DNA binding and ligand and co-factor interactions. Immunofluorescence of cells transfected with mutant SF1 revealed abnormal sub-cellular localisation for 10 mutants. Our data suggest that DSD in patients with SF1 mutations may be caused by dysregulation of SOX9 expression via TESCO, possibly due to impaired DNA binding, defective ligand or co-factor interactions or abnormal nuclear localisation. In conclusion, this study has enabled us to understand the consequences of SF1 mutations in DSDs and how they relate to phenotype severity.