A zinc finger truncation of murine WT1 results in the characteristic urogenital abnormalities of Denys–Drash syndrome

Abstract

The Wilms tumor-suppressor gene, WT1, plays a key role in urogenital development, and WT1 dysfunction is implicated in both neoplastic (Wilms tumor, mesothelioma, leukemias, and breast cancer) and nonneoplastic (glomerulosclerosis) disease. The analysis of diseases linked specifically with WT1 mutations, such as Denys-Drash syndrome (DDS), can provide valuable insight concerning the role of WT1 in development and disease. DDS is a rare childhood disease characterized by a nephropathy involving mesangial sclerosis, XY pseudohermaphroditism, and͞or Wilms tumor (WT). DDS patients are constitutionally heterozygous for exonic point mutations in WT1, which include mutations predicted to truncate the protein within the Cterminal zinc finger (ZF) region. We report that heterozygosity for a targeted murine Wt1 allele, Wt1 tmT396 , which truncates ZF3 at codon 396, induces mesangial sclerosis characteristic of DDS in adult heterozygous and chimeric mice. Male genital defects also were evident and there was a single case of Wilms tumor in which the transcript of the nontargeted allele showed an exon 9 skipping event, implying a causal link between Wt1 dysfunction and Wilms tumorigenesis in mice. However, the mutant WT1 tmT396 protein accounted for only 5% of WT1 in both heterozygous embryonic stem cells and the WT. This has implications regarding the mechanism by which the mutant allele exerts its effect.WT1 is expressed at high levels in those mesodermally derived tissues that experience mesenchymal-epithelial transition during development, including the genital ridge and developing mesothelium, kidney, and gonads (1, 2). WT1 expression is linked with podocyte differentiation during nephrogenesis, but continues to be expressed in adult podocytes, gonads (Sertoli and granulosa cells), uterus (myometrium), spleen (stromal cells and capsule), and mesothelial cells that line the body cavities and visceral organs. Thus, WT1 has a role in both homeostasis and development. The critical role for WT1 in urogenital development is underlined by the failure of kidney and gonadal development in mice homozygous for a Wt1 null mutation (3). WT1 dysfunction also is implicated in the etiology of certain Wilms tumors (WTs) that derive from the metanephric blastemal cells that should differentiate normally into the epithelial and mesenchymal components of the kidney (4).WT1 encodes a nuclear protein with structural motifs characteristic of transcription factors, including an N-terminal glutamine͞proline-rich transregulatory domain and a Cterminal domain with four Kruppel-type Cys 2 -His 2 zinc fingers (ZFs) (ZF1-4) that bind DNA and RNA and are involved in nuclear localization (5-7). WT1 potentially encodes 16 protein isoforms as a result of RNA editing as well as alternative splicing and translation initiation (8). There are two alternatively spliced exons: ASI and ASII. The first is 17 aa long and constitutes exon 5. ASII results from variation in the splice donor site used at the end of exon 9 and inserts 3 aa (lysine-threonine-s...

Wnt-4 regulation by the Wilms' tumour suppressor gene, WT1

“…It was not possible to determine the e ciency with which the mutant transcript was translated as only a single amino acid has been altered. However, our previous experience with the generation of a mouse model of DDS using a truncation mutation would suggest that the mutant protein is produced (Patek et al, 1999). In addition, Western blotting and immuno¯uorescence were performed on C2A and C5B, revealing an increase in protein production of approximately twofold (Figure 1f).…”

Clinical and Molecular Characterization of Patients with Heterozygous Mutations in Wilms Tumor Suppressor Gene 1

“…Heterozygous germline mutations in WT1 were described in two rare human conditions: Denys-Drash syndrome (DDS) with exon mutations in the zinc-finger region (7,8) and Frasier syndrome (FS) with mutations affecting the canonic donor KTS splice site of intron 9 (9,10). DDS includes steroid-resistant nephrotic syndrome rapidly progressing to ESRD, 46 XY disorder in sex development (DSD) with sex reversal and a high risk for WT (11,12).…”

Transcriptional activity of testis-determining factor SRY is modulated by the Wilms’ tumor 1 gene product, WT1

“…Based on the observation of WT1-knockout mice, WT1 is required for the development of several organs, including kidney, gonad, spleen, adrenal gland, epicardium, and retina (Kreidberg et al, 1993;Herzer et al, 1999;Moore et al, 1999;Patek et al, 1999;Wagner et al, 2002a,b). For testis development, WT1 is expressed in the somatic cells of developing gonads and is restricted to the Sertoli cells of the testis (Pelletier et al, 1991;Armstrong et al, 1993).…”