who carried a de novo pseudodicentric duplicated chromosome 8 of maternal origin. This gave us the opportunity to study the genes contained within the duplication, which include SOX17. We have identified a p.Y259N mutation in SOX17 in this girl, and we subsequently identified the same mutation in five additional patients with congenital anomalies of the kidney and the urinary tract (CAKUT). Furthermore, two patients presenting with CAKUT carried other SOX17 mutations. Functional studies of SOX17–p.Y259N demonstrated that this protein abnormally accumulates in cultured kidney cells, resulting in inhibition of the canonical Wnt signaling pathway. Given the vital role of this pathway in multiple stages of kidney development, we propose a model by which elevated levels of mutant SOX17– p.Y259N protein inhibit Wnt signaling resulting in abnormal kidney and urinary tract development. Materials and Methods Patients Fifty-eight familial cases with vesico ureteral reflux (VUR) belonging to 10 small families were analyzed. In two of the families the same p.Y259N mutation in SOX17 was found (see the Results section). Pedigrees of the two families are shown in Figure 1A. Family 1 Patient (II,1) was born from non consanguineous parents. The pregnancy was achieved after in vitro fertilization (FIVET). A Cesarean section at the 34th week of gestation was performed because of polydramnios and a prenatal ultrasonography showing severe bilateral hydronephrosis in the fetus. At birth, she was put on intensive care because of her critical health conditions. In the first 3 years of her life, she had presented mild axial hypotonia and Figure 1. A: Pedigrees of affected families. Family 1: I.1 healthy father; I.2 mother with recurrent urinary infections, duplex ureter with dilatation, chronic constipation; II.1 46,XX,dirdup(8q)(q11.1q12.1)dic, duplex ureter dx, reflux dx (III degree), hydronephrosis sn, reflux (IV degree), chronic constipation, thinning of the posterior portion of the body of the corpus callosum. Family 2: I.1 Healthy father; I.2 vesicoureteral reflux; II.1 hydronephrosis sn, stenosis of the pyeloureteral joint sn. B: Localization of mutations along the SOX17 gene coding sequence. The location of the HGM box and the G/P-rich region are indicated. The positions of tyrosine residues ( ) predicted to undergo phosphorylation are also indicated. One of these residues is the mutated Y259. psychomotor development with a deficit in expressive language. She had chronic constipation. Echocardiography showed a normal heart with large PDA and a bidirectional shunt. Encephalic RM demonstrated abnormal morphology of the corpus callosum, characterized by evident thinning of the posterior portion of the body and of the isthmus. Her right kidney showed a duplicated pyeloureteral collecting system of about 7 cm in diameter and bilateral VUR (III). Ureteroplasty was performed to correct the patient’s urinary defects. The patient’s mother (I,2) showed VUR associated to megaureter with recurrent urinary infections, chronic constipation, and coloboma of the iris in her left eye. Her father was healthy. Family 2 Patient (II,1) is a male child with left hydronephrosis due to stenosis of the pyeloureteral joint. His mother (I,2) has vesicoureteral reflux, whereas his father is healthy. Sporadic Cases Additionally, 178 patients with proven VUR and a history of urinary tract infection were enrolled in the study. Control DNAs were obtained from 88 normal subjects, 82 cord blood samples from normal donors, and 135 patients with nephrotic syndrome. Although the former two groups were not controlled in respect to the presence of any defect of the urinary tract, all patients with nephrotic syndrome had an echosonography showing normality. The demographic and clinical data relative to all the above cohorts are given in Table 1. All patients with VUR had been studied with echosonography, voiding cystography and Tc99 mDMSA scan in order to verify the presence of malformations of the kidney and urinary tract and/or renal scars. Static renal scintigraphy was recorded 3 to 4 hr after injection of a weight-scaled dose of technetium-99 m DMSA to obtain views in the posterior and both posterior oblique projections for 300 kilocounts or more. Focal or diffuse areas of decreased uptake in the first scan, without evidence of cortical loss, indicated acute pyelonephritis. Renal scarring was defined as decreased uptake with distortion of the contours or as cortical thinning with loss of parenchymal volume. Two nuclear physicians, blind to the test results, interpreted the scans independently and resolved discrepancies by discussion. Cytogenetics, Immunofluorescence, and Array-CGH Investigations Chromosome preparations were made from peripheral blood, skin, and ureter biopsies from the proposita using standard techniques. To define the extension of the duplicated segment an array-CGH experiment was performed using the Human Genome CGH Microarray Kits 44B (Agilent Technologies, Palo Alto, CA) covering the whole genome with a resolution of 100 kb. Briefly, 1 mg of patient and sex-matched pooled reference DNAs were processed according to the manufacturer’s protocol. Fluorescence was scanned in a dual-laser scanner and the images were extracted and analyzed with Agilent Feature Extraction software (v9.5.3.1) and CGH Analytics software (v3.5.14), respectively. Changes in test DNA copy number at a specific locus are observed as the deviation of the log ratio value from a modal value of 0. To confirm the breakpoints, structure, and orientation of the duplicated region we used fluorescent in situ hybridization (FISH) HUMAN MUTATION, Vol. 31, No. 12, 1352–1359, 2010 1353