Functional characterization of Wilms tumor-suppressor WTX and tumor-associated mutants

Abstract

The WTX, Wilms tumor-associated tumor-suppressor gene, is present on the X chromosome and a single WTX mutation may be sufficient to promote carcinogenesis. Unlike the WT1 tumor suppressor, a transcription factor, WTX lacks conserved functional protein domains. To study the function of WTX, we constructed inducible cell lines expressing WTX and tumor-associated WTX mutants. Induction of WTX inhibited cell growth and caused G 1 /G 0 arrest. In contrast, a short, tumorassociated truncation mutant of WTX358 only slightly inhibited cell growth without a significant cell-cycle arrest, although expression of a longer truncation mutant WTX565 led to the growth inhibition and cell-cycle arrest to a similar extent as wild-type WTX. Like WT1, WTX slowed growth and caused cell-cycle arrest through p21 induction. Gene expression profiling showed that these two tumorsuppressors regulated genes in similar pathways, including those implicated in control of the cellular growth, cell cycle, cell death, cancer and cardiovascular system development. When gene expression changes mediated by wild-type WTX were compared with those affected by mutant forms, WTX565 showed a 55% overlap (228 genes) in differentially regulated genes, whereas WTX358 regulated only two genes affected by wild-type WTX, implying that amino-acid residues 358-561 are critical for WTX function.

Wilms Tumor Gene on X Chromosome (WTX) Inhibits Degradation of NRF2 Protein through Competitive Binding to KEAP1 Protein

“…We hypothesize that WTX enhances NRF2 steady-state levels by disrupting the conformation of the E3 ubiquitin ligase complex, resulting in lower ubiquitination of NRF2. In support of our findings, expression of the NRF2 target genes NQO1 and HMOX1 was significantly enhanced in HEK293T cells overexpressing a fragment of WTX containing the KEAP1 interaction domain (37). …”

Different incidences of epigenetic but not genetic abnormalities between Wilms tumors in Japanese and Caucasian children

“…A lack of WT1 induces WT precursor cells to undergo apoptosis and mutant b-catenin might rescue the cells by the activation of Wnt signaling. (24) The present study showed the comparable frequencies of WTX abnormalities in sporadic WTs with and without WT1 abnormalities; the findings were previously reported by Ruteshouser et al (Table 2). (18) In addition to the inhibitory role in Wnt signaling pathway, WTX has another role in nuclear pathways implicated in the transcriptional regulation of cellular differentiation and cell death programs, (24,25) and the damage of this pathway by WTX alterations might involve in the tumorigenic process and explain why the alterations were found in both WT1-wild-type and WT1-mutant type tumors.…”

The Na+/H+ Exchanger NHE6 Modulates Endosomal pH to Control Processing of Amyloid Precursor Protein in a Cell Culture Model of Alzheimer Disease

“…A 10-l sample was added to a disposable cell counting chamber slide and inserted into a Countess automated cell counter (Invitrogen). The cell viability and total cell numbers were recorded from six independent experiments, as described earlier (29).…”