Analysis of WT1 target gene expression in stably transfected cell lines

Thäte, Claudia; Englert, Christoph; Gessler, Manfred

doi:10.1038/sj.onc.1202055

Cited by 30 publications

(20 citation statements)

References 22 publications

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“…In contrast, M15 cells were transfected with a WT1 antisense construct to abolish endogenous expression of all WT1 isoforms. Like previous expression profiling analyses of WT1 function (Thate et al, 1998;Lee and Pelletier, 2001;Palmer et al, 2001), we were unable to confirm changes in the expression of the majority of previously proposed WT1 target genes, with the exception of c-myc and ODC. However, by examining the complementary overlap in expression change elicited by WT1 induction versus repression in these two cell lines, we have demonstrated that the most pronounced changes in mRNA steady-state levels was seen for genes involved in the synthesis of cholesterol and fatty acids.…”

Section: Discussioncontrasting

confidence: 40%

“…Expression profiling has been applied to WT1 biology both to investigate the validity of previously proposed WT1 targets, and to identify additional physiological targets. Generally, this has involved the generation of stable cell lines (including Saos2, U2OS, G401, HEK293) expressing one of the common WT1 isoforms often under an inducible promoter (Thate et al, 1998;Lee et al, 1999;Lee and Pelletier, 2001;Sim et al, 2002). The universal conclusion drawn from expression profiling analyses of the effect of WT1 expression is that few, if any, of the previously proposed targets of WT1 are indeed physiologically relevant.…”

Section: Introductionmentioning

confidence: 99%

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Anlaysis of complementary expression profiles following WT1 induction versus repression reveals the cholesterol/fatty acid synthetic pathways as a possible major target of WT1

et al. 2004

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Section: Discussioncontrasting

confidence: 40%

Section: Introductionmentioning

confidence: 99%

Anlaysis of complementary expression profiles following WT1 induction versus repression reveals the cholesterol/fatty acid synthetic pathways as a possible major target of WT1

et al. 2004

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“…WT1 was previously thought to inhibit cell proliferation by repression of growth-promoting genes. However, a survey using several cellular models failed to confirm regulation of 16 genes proposed to be repressed by WT1 (50). This information suggested that the initial characterization of WT1 as a transcriptional repressor needed to be re-evaluated.…”

Section: Spry1 a Wt1 Target Gene During Kidney Developmentmentioning

confidence: 99%

The Receptor Tyrosine Kinase Regulator Sprouty1 Is a Target of the Tumor Suppressor WT1 and Important for Kidney Development

Groß

Morrison

Hyink

et al. 2003

Journal of Biological Chemistry

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WT1 encodes a transcription factor involved in kidney development and tumorigenesis. Using representational difference analysis, we identified a new set of WT1 targets, including a homologue of the Drosophila receptor tyrosine kinase regulator, sprouty. Sprouty1 was up-regulated in cell lines expressing wild-type but not mutant WT1. WT1 bound to the endogenous sprouty1 promoter in vivo and directly regulated sprouty1 through an early growth response gene-1 binding site. Expression of Sprouty1 and WT1 overlapped in the developing metanephric mesenchyme, and Sprouty1, like WT1, plays a key role in the early steps of glomerulus formation. Disruption of Sprouty1 expression in embryonic kidney explants by antisense oligonucleotides reduced condensation of the metanephric mesenchyme, leading to a decreased number of glomeruli. In addition, sprouty1 was expressed in the ureteric tree and antisense-treated ureteric trees had cystic lumens. Therefore, sprouty1 represents a physiologically relevant target gene of WT1 during kidney development.The development of the mammalian metanephric kidney is a model for the study of cellular and molecular mechanisms of organogenesis (1). Wilms tumor, a pediatric kidney malignancy, is characterized by a triphasic histopathology (blastemal, stromal, epithelial) signifying an abnormal differentiation program. In accordance with this notion, the Wilms Tumor suppressor gene 1 (WT1), inactivated in a subset of Wilms tumors, plays an essential role in normal development of the kidney and the genitourinary system (2-4). Targeted disruption of WT1 in mice leads to a complete agenesis of the kidneys and gonads (5). The specific temporal and spatial pattern of WT1 expression suggests multiple roles for WT1 during nephrogenesis. In particular, WT1 expression peaks during the mesenchymal-to-epithelial transition (MET), 1 suggesting an instructive role in the formation of the renal glomerulus. In the mature kidney, WT1 expression becomes restricted to the podocytes, perhaps being involved in maintaining a differentiated phenotype.The WT1 gene encodes a C 2 -H 2 zinc finger transcription factor that binds to both GC-rich and TC repeat elements. Alternative splicing at two sites yields four major isoforms, each containing or lacking 17 amino acids between the transactivation domain and the zinc finger region and/or a 3-amino acid insertion (KTS) between the third and fourth zinc fingers. The KTS insertion disrupts the spacing between the zinc fingers, altering DNA binding (2, 4). The A isoform lacks both these motifs and binds strongly to DNA. The C isoform contains the KTS insertion, displays a weaker DNA binding to alternative sequences, associates with splicing factors, and therefore, may be involved in RNA processing. WT1 binding sites were identified in multiple promoters that respond to WT1 in transient transfection assays (2, 6, 7); however, most are not regulated by WT1 in vivo (8). Thus, identification of bona fide target genes would provide insight into WT1 function in development and tumo...

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“…293T cells were chosen for these experiments, since it has been well established that these cells do not express detectable amounts of WT1 RNA or proteins (Thäte et al 1998;Lee and Pelletier 2001;Vajjhala et al 2003;Rae et al 2004). Thus, these cells provide an excellent "complementation" system for analysis of WT1 effects.…”

Section: The Wt1(+kts) Protein Enhances Expression From Unspliced Rnamentioning

confidence: 99%

The Wilms’ tumor 1 (WT1) gene (+KTS isoform) functions with a CTE to enhance translation from an unspliced RNA with a retained intron

Bor¹,

Swartz²,

Morrison³

et al. 2006

Genes Dev.

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The Wilms' tumor 1 (WT1) gene plays an important role in mammalian urogenital development, and dysregulation of this gene is observed in many human cancers. Alternative splicing of WT1 RNA leads to the expression of two major protein isoforms, WT1(+KTS) and WT1(−KTS). Whereas WT1(−KTS) acts as a transcriptional regulator, no clear function has been ascribed to WT1(+KTS), despite the fact that this protein is crucial for normal development. Here we show that WT1(+KTS) functions to enhance expression from RNA possessing a retained intron and containing either a cellular or viral constitutive transport element (CTE). WT1(+KTS) expression increases the levels of unspliced RNA containing a CTE and specifically promotes the association of this RNA with polyribosomes. These studies provide further support for links between different steps in RNA metabolism and for the existence of post-transcriptional operons.

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Analysis of WT1 target gene expression in stably transfected cell lines

Cited by 30 publications

References 22 publications

Anlaysis of complementary expression profiles following WT1 induction versus repression reveals the cholesterol/fatty acid synthetic pathways as a possible major target of WT1

Anlaysis of complementary expression profiles following WT1 induction versus repression reveals the cholesterol/fatty acid synthetic pathways as a possible major target of WT1

The Receptor Tyrosine Kinase Regulator Sprouty1 Is a Target of the Tumor Suppressor WT1 and Important for Kidney Development

The Wilms’ tumor 1 (WT1) gene (+KTS isoform) functions with a CTE to enhance translation from an unspliced RNA with a retained intron

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