ROBO2 gene variants in children with primary nonsyndromic vesicoureteral reflux with or without renal hypoplasia/dysplasia

BACKGROUND: Primary nonsyndromic vesicoureteral reflux (VUR) and VUR with renal hypoplasia/dysplasia (VUR-RHD) are common congenital anomalies of the kidney and urinary tract (CAKUT). Sequence variations of the ROBO2 gene were investigated in children with nonsyndromic VUR or VUR-RHD. METHODS: Single-strand conformation polymorphism (SSCP) electrophoresis or multiple restriction fragment SSCP (MRF-SSCP), followed occasionally by direct sequencing, was used to screen 103 patients and 200 controls for nucleotide changes. Gene polymorphisms and transposable elements were investigated using bioinformatics. RESULTS: Two single-nucleotide polymorphisms were detected: IVS1-53 and IVS5-31. The frequency of A allele of IVS1-53G>A did not differ significantly between patients and controls. IVS1-53 does not affect mRNA splicing according to in silico analysis. IVS5-31A>G substitution was found in one patient, reported here for the first time in VUR. In silico results demonstrated alteration in two serine/arginine-rich (SR) protein-binding sites and two additional acceptor sites. The ROBO2 gene sequence was found to contain 25.9% transposable elements. CONCLUSION: ROBO2 variants were not found to be associated with nonsyndromic VUR or VUR-RHD, providing further evidence for genetic heterogeneity. The role of transposable elements in ROBO2 gene expression in CAKUT needs further investigation since they are generally considered to be mutagens.