The Wilms tumor suppressor WT1 directs stage-specific quiescence and differentiation of human hematopoietic progenitor cells

The WT1 gene is a tumor-suppressor gene that was isolated as a gene responsible for Wilms' tumor, a childhood kidney neoplasm. We have previously reported that the WT1 gene is strongly expressed in leukemia cells with an increase in its expression levels at relapse and an inverse correlation between its expression levels and prognosis, thus making it a novel tumor marker for leukemic blast cells. Furthermore, WT1 antisense oligomers have been found to inhibit the growth of leukemic cells. These results strongly suggested the involvement of the WT1 gene in human leukemogenesis. The present study was performed to prove our hypothesis that the WT1 gene plays a key role in leukemogenesis and performs an oncogenic function in hematopoietic progenitor cells, rather than a tumor-suppressor gene function. 32D cl3, an interleukin-3–dependent myeloid progenitor cell line, differentiates into mature neutrophils in response to granulocyte colony-stimulating factor (G-CSF). However, when transfected wild-type WT1 gene was constitutively expressed in 32D cl3, the cells stopped differentiating and continued to proliferate in response to G-CSF. As for signal transduction mediated by G-CSF receptor (G-CSFR), Stat3α was constitutively activated in wild-type WT1-infected 32D cl3 in response to G-CSF, whereas, in WT1-uninfected 32D cl3, activation of Stat3α was only transient. However, most interesting was the fact that G-CSF stimulation resulted in constitutive activation of Stat3β only in wild-type WT1-infected 32D cl3, but not in WT1-uninfected 32D cl3. Thus, WT1 expression constitutively activated both Stat3α and Stat3β. A transient activation of Stat1 was detected in both wild-type WT1-infected and uninfected 32D cl3 after G-CSF stimulation, but no difference in its activation was found. No activation of MAP kinase was detected in both wild-type WT1-infected and uninfected 32D cl3 after G-CSF stimulation. These results demonstrated that WT1 expression competed with the differentiation-inducing signal mediated by G-CSFR and constitutively activated Stat3, resulting in the blocking of differentiation and subsequent proliferation. Therefore, the data presented here support our hypothesis that the WT1 gene plays an essential role in leukemogenesis and performs an oncogenic function in hematopoietic progenitor cells and represent the first demonstration of an important role of the WT1 gene in signal transduction in hematopoietic progenitor cells.

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The Wilms' tumor suppressor Wt1 activates transcription of the erythropoietin receptor in hematopoietic progenitor cells

The FASEB Journal ◽

10.1096/fj.07-097576 ◽

2008 ◽

Vol 22 (8) ◽

pp. 2690-2701 ◽

Cited By ~ 17

Author(s):

Karin M. Kirschner ◽

Patricia Hagen ◽

Christiane S. Hussels ◽

Matthias Ballmaier ◽

Holger Scholz ◽

...

Keyword(s):

Tumor Suppressor ◽

Progenitor Cells ◽

Wilms Tumor ◽

Erythropoietin Receptor ◽

Hematopoietic Progenitor Cells ◽

Hematopoietic Progenitor

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The Wilms’ Tumor Suppressor Wt1 Activates Transcription of the Erythropoietin-Receptor in Hematopoietic Progenitor Cells.

Blood ◽

10.1182/blood.v108.11.1188.1188 ◽

2006 ◽

Vol 108 (11) ◽

pp. 1188-1188

Author(s):

Christof Dame ◽

Karin M. Kirschner ◽

Patricia Hagen ◽

Christiane S. Hussels ◽

Holger Scholz

Keyword(s):

Tumor Suppressor ◽

Progenitor Cells ◽

Wilms Tumor ◽

Regulation Of Gene Expression ◽

Erythropoietin Receptor ◽

Hematopoietic Progenitor Cells ◽

Luciferase Reporter ◽

Recombinant Erythropoietin ◽

Wild Type ◽

Hematopoietic Progenitor

Abstract The Wilms’ tumor suppressor, Wt1, has been implicated in the stage-specific quiescence and differentiation of hematopoietic progenitor cells. Since Wt1 deficiency compromises the proliferation and differentiation of erythroid blast-forming units (BFU-E) and colony-forming units (CFU-E), we analyzed the potential role of the transcriptionally active Wt1 isoform, Wt1(-KTS), in regulating the expression of the erythropoietin receptor (EpoR). CD117-positive hematopoietic progenitor cells isolated from the liver of Wt1-deficient murine embryos (Wt1−/−) at 11.5 dpc exhibited a 10-fold lower proliferative responsiveness to recombinant erythropoietin than CD117-positive cells from wild-type embryos (Wt1+/+), or embryos with heterozygous Wt1-deletion (Wt1+/−). Benzidine staining revealed a reduced fraction of hemoglobin-containing cells in Wt1-deficient cells in comparison to Wt1+/+ cells after 48 hrs of culture of CD117-positive cells in the presence of 6 U rhEpo/ml medium. Consistently, expression of the EpoR was significantly reduced in hematopoietic progenitor cells that lack Wt1 compared to Wt1+/+ cells. The decrease of EpoR mRNA was not due to a general down-regulation of gene expression in Wt1−/− CD117-positive cells, since levels of c-kit and c-mpl expression were normal. In the heart of wild-type and Wt1−/− embryos, which was analyzed as a non-hematopoietic tissue expressing both EpoR und Wt1, no difference in Epo-R expression was found. Transient transfection of Wt1(-KTS) into human hepatoma-derived HepG2 cells increased EpoR transcripts approximately 2-fold. A luciferase reporter plasmid carrying 309 bp of the proximal human EpoR promoter was stimulated 8-fold by co-transfection with Wt1(-KTS). The responsible cis-element in the EpoR promoter was identified by mutation analysis, electrophoretic mobility shift assay, and chromatin immunoprecipitation assay. In summary, we identified the EpoR gene to be the first downstream target of Wt1 within the hematopoietic cytokine receptor superfamily. Our data indicate that activation of the EpoR gene by Wt1 is a critical mechanism in normal murine hematopoiesis. However, our data do not support a role for Wt1 in EpoR expression in non-erythoid tissues, such as the heart.

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JARID2 Functions as a Tumor Suppressor in Myeloid Neoplasms by Repressing Self-Renewal in Hematopoietic Progenitor Cells

Cancer Cell ◽

10.1016/j.ccell.2018.10.008 ◽

2018 ◽

Vol 34 (5) ◽

pp. 741-756.e8 ◽

Cited By ~ 9

Author(s):

Hamza Celik ◽

Won Kyun Koh ◽

Ashley C. Kramer ◽

Elizabeth L. Ostrander ◽

Cates Mallaney ◽

...

Keyword(s):

Tumor Suppressor ◽

Progenitor Cells ◽

Hematopoietic Progenitor Cells ◽

Hematopoietic Progenitor ◽

Self Renewal ◽

Myeloid Neoplasms

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The Transcriptional Program of Polycythemia Vera Revelas Altered Expression of Multiple Putative Tumor Suppressor Genes.

Blood ◽

10.1182/blood.v104.11.796.796 ◽

2004 ◽

Vol 104 (11) ◽

pp. 796-796 ◽

Cited By ~ 1

Author(s):

Windy D. Berkofsk-Fessler ◽

Jonathan D. Licht ◽

Melanie-Jane McConnell ◽

Donna S. Neuberg ◽

Timothy S. Bowler ◽

...

Keyword(s):

Tumor Suppressor ◽

Polycythemia Vera ◽

Progenitor Cells ◽

Tumor Suppressor Genes ◽

Hematopoietic Progenitor Cells ◽

Multiple Testing Correction ◽

Hematopoietic Progenitor ◽

Suppressor Genes ◽

Putative Tumor Suppressor ◽

Cd34 Cells

Abstract Polycythemia vera (PV) is a myeloproliferative disease characterized by accumulation of erythrocytes and cells of the myeloid and megakaryocyte lineages. Although genes like PRV-1 and PTP-MEG2 have been implicated in the pathology of PV, there is no consensus on their importance in the disease process. Progenitor cells from PV patients can grow in the absence of erythropoietin, and are hypersensitive to a variety of other growth factors. This suggests that polycythemic hematopoietic progenitor cells possess a significantly different genetic program. We tested this idea by molecular profiling hematopoietic progenitor cells (CD34+) from PV specimens and normal donors. We purified CD34+ cells from the marrow of 10 PV patients and harvested total RNA. Biotinylated cRNA was made through two rounds of linear amplification, and hybridized to Affymetrix HGU133A genechips. CD34+ cells from marrow mononuclear cells of 5 normal controls were processed similarly. The resulting datasets were normalized to the median across chips and across genes. Unsupervised hierarchical clustering showed that PV samples had a distinct gene expression profile from the controls. We then performed supervised clustering using a non-parametric t-test (Wilcoxon rank sum test) using the Benjamini and Hochberg multiple testing correction held to a p-value of 0.01 to determine genes that were significantly different between disease and normal samples. Using these stringent criteria, there were 331 genes that reached significance. Strikingly most of these were decreased in expression compared with control CD34+ cells and only 34 genes were upregulated in PV. A 35 gene predictor set was discovered through the use of a k-nearest neighbor metric. This set was 100% accurate for the prediction of PV in a leave one out cross-validation approach. Among these genes are EVI1, a known oncogene and one of only two genes upregulated in PV on this list, and the putative tumor suppressor genes TUSC4 (NPR2), NDRG1 and KLF4. Also among the predictor genes is BAALC, a gene expressed in normal CD34+ cells and known to be a prognostic indicator gene for acute myeloid leukemia.

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Leukemia associated mutant Wilms’ tumor gene 1 protein promotes expansion of human hematopoietic progenitor cells

Leukemia Research ◽

10.1016/j.leukres.2013.06.018 ◽

2013 ◽

Vol 37 (10) ◽

pp. 1341-1349 ◽

Cited By ~ 4

Author(s):

Karina Vidovic ◽

Tove Ullmark ◽

Bodil Rosberg ◽

Andreas Lennartsson ◽

Tor Olofsson ◽

...

Keyword(s):

Progenitor Cells ◽

Wilms Tumor ◽

Hematopoietic Progenitor Cells ◽

Hematopoietic Progenitor ◽

Wilms Tumor Gene ◽

Tumor Gene

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Aberrant Overexpression of the Wilms Tumor Gene (WT1) in Human Leukemia

Blood ◽

10.1182/blood.v89.4.1405 ◽

1997 ◽

Vol 89 (4) ◽

pp. 1405-1412 ◽

Cited By ~ 258

Author(s):

Kazushi Inoue ◽

Hiroyasu Ogawa ◽

Yoshiaki Sonoda ◽

Takafumi Kimura ◽

Hideaki Sakabe ◽

...

Keyword(s):

Progenitor Cells ◽

Wilms Tumor ◽

Leukemic Cells ◽

Hematopoietic Progenitor Cells ◽

Cell Populations ◽

Human Leukemia ◽

Wt1 Gene ◽

Hematopoietic Progenitor ◽

Expression Levels ◽

Hla Dr

Abstract To clarify whether the expression of the WT1 gene in leukemic cells is aberrant or merely reflects that in normal counterparts, the expression levels of the WT1 gene were quantitated for normal hematopoietic progenitor cells. Bone marrow (BM) and umbilical cord blood (CB) cells were fluorescence-activated cell sorting (FACS)-sorted into CD34+ and CD34− cell populations, and the CD34+ cells into nine subsets (CD34+CD33−, CD34+CD33+, CD34+CD38−, CD34+CD38+, CD34+HLA-DR−, CD34+HLA-DR+, CD34+c-kithigh, CD34+c-kitlow, and CD34+c-kit−) according to the expression levels of CD34, CD33, CD38, HLA-DR, and c-kit. Moreover, acute myeloid leukemic cells were also FACS-sorted into four populations (CD34+CD33−, CD34+CD33+, CD34− CD33+, and CD34− CD33−). FACS-sorted normal hematopoietic progenitor and leukemic cells and FACS-unsorted leukemic cells were examined for the WT1 expression by quantitative reverse transcriptase-polymerase chain reaction. The WT1 expression in the CD34+ and CD34− cell populations and in the nine CD34+ subsets of BM and CB was at either very low (1.0 to 2.4 × 10−2) or undetectable (<10−2) levels (the WT1 expression level of K562 cells was defined as 1.0), whereas the average levels of WT1 expression in FACS-sorted and -unsorted leukemic cells were 2.4 to 9.3 × 10−1. Thus, the WT1 expression levels in normal hematopoietic progenitor cells were at least 10 times less than those in leukemic cells. Therefore, we could not find any normal counterparts of BM or CB that expressed the WT1 at levels comparable with those in leukemic cells. These results indicate an aberrant overexpression of the WT1 gene in leukemic cells and imply the involvement of this gene in human leukemogenesis.

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