Activating c-KIT mutations confer oncogenic cooperativity and rescue RUNX1-ETO-induced DNA damage in human cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE58041 | GEO | 2014/09/30
SECONDARY ACCESSION(S): PRJNA248751
REPOSITORIES: GEO
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Project description:REtr causes genomic instability in human CD34+ cells. Activated forms of c-KIT, like c-KIT(N822K), rescue the REtr-induced genomic instability by increasing the rate of DNA repair by homologous recombination. Human CD34+ hematopoietic progenitor cells were transduced with retroviral vectors encoding c-KIT(N822K) or mock control vectors, expanded in culture and FACS selected prior to molecular analysis.
2014-09-30 | E-GEOD-58041 | biostudies-arrayexpress
Project description:REtr causes genomic instability in U937 cells. Activated forms of c-KIT, like c-KIT(N822K), rescues the Retr induced genomic instability by increasing the rate of DNA repair by homologous recombination
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Project description:REtr causes genomic instability in U937 cells. Activated forms of c-KIT, like c-KIT(N822K), rescues the Retr induced genomic instability by increasing the rate of DNA repair by homologous recombination Human U937 cells were transduced with retroviral vectors encoding REtr or c-KIT(N822K), expanded in culture and FACS selected prior to molecular analysis
2014-07-29 | E-GEOD-57658 | biostudies-arrayexpress
Project description:The RUNX1/ETO (RE) fusion protein, which originates from the t(8;21) chromosomal rearrangement, is one of the most frequent translocation products found in de novo acute myeloid leukemia (AML). In RE leukemias, activated forms of the c-KIT tyrosine kinase receptor are frequently found, thereby suggesting oncogenic cooperativity between these oncoproteins in the development and maintenance of t(8;21) malignancies. In this report, we show that activated c-KIT cooperates with a C-terminal truncated variant of RE, REtr, to expand human CD34+ hematopoietic progenitors ex vivo. CD34+ cells expressing both oncogenes resemble the AML-M2 myeloblastic cell phenotype, in contrast to REtr-expressing cells which largely undergo granulocytic differentiation. Oncogenic c-KIT amplifies REtr-depended clonogenic growth and protects cells from exhaustion. Activated c-KIT reverts REtr-induced DNA damage and apoptosis. In the presence of activated c-KIT, REtr-downregulated DNA-repair genes are re-expressed leading to an enhancement of DNA-repair efficiency via homologous recombination. Together, our results provide new mechanistic insight into REtr and c-KIT oncogenic cooperativity and suggest that augmented DNA repair accounts for the increased chemoresistance observed in t(8;21)-positive AML patients with activated c-KIT mutations. This cell-protective mechanism might represent a new therapeutic target, as REtr cells with activated c-KIT are highly sensitive to pharmacological inhibitors of DNA repair.
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Project description:Acute myeloid leukemia development occurs in a step-wise fashion whereby an original driver mutation is followed by additional mutations. The first type of mutations tends to be in genes encoding members of the epigenetic/transcription regulatory machinery (i.e. RUNX1, DNMT3, TET2), while the secondary mutations often involve genes encoding members of signalling pathways that cause uncontrolled growth of such cells such as the growth factor receptors c-KIT of FLT3. Patients usually present with both types of mutations, but it is currently unclear how both mutational events shape the epigenome in developing AML cells. To this end we generated an in vitro model of t(8;21) AML by expressing its driver oncoprotein RUNX1-ETO with or without a mutated (N822K) KIT protein. The expression of N822K–c-KIT strongly increases the self-renewal capacity of RUNX1-ETO expressing cells. Global analysis of gene expression changes and alterations in the epigenome show that N822K–c-KIT expression profoundly influences the open chromatin landscape and transcription factor binding. However, our experiments also show that double mutant cells still differ from their patient derived counterparts, highlighting the importance of studying patient cells to obtain a true picture of how gene regulatory networks have been reprogrammed during tumourigenesis.
2021-04-01 | GSE152334 | GEO
Project description:Acute myeloid leukemia development occurs in a step-wise fashion whereby an original driver mutation is followed by additional mutations. The first type of mutations tends to be in genes encoding members of the epigenetic/transcription regulatory machinery (i.e. RUNX1, DNMT3, TET2), while the secondary mutations often involve genes encoding members of signalling pathways that cause uncontrolled growth of such cells such as the growth factor receptors c-KIT of FLT3. Patients usually present with both types of mutations, but it is currently unclear how both mutational events shape the epigenome in developing AML cells. To this end we generated an in vitro model of t(8;21) AML by expressing its driver oncoprotein RUNX1-ETO with or without a mutated (N822K) KIT protein. The expression of N822K–c-KIT strongly increases the self-renewal capacity of RUNX1-ETO expressing cells. Global analysis of gene expression changes and alterations in the epigenome show that N822K–c-KIT expression profoundly influences the open chromatin landscape and transcription factor binding. However, our experiments also show that double mutant cells still differ from their patient derived counterparts, highlighting the importance of studying patient cells to obtain a true picture of how gene regulatory networks have been reprogrammed during tumourigenesis.
2021-04-01 | GSE152333 | GEO
Project description:Acute myeloid leukemia development occurs in a step-wise fashion whereby an original driver mutation is followed by additional mutations. The first type of mutations tends to be in genes encoding members of the epigenetic/transcription regulatory machinery (i.e. RUNX1, DNMT3, TET2), while the secondary mutations often involve genes encoding members of signalling pathways that cause uncontrolled growth of such cells such as the growth factor receptors c-KIT of FLT3. Patients usually present with both types of mutations, but it is currently unclear how both mutational events shape the epigenome in developing AML cells. To this end we generated an in vitro model of t(8;21) AML by expressing its driver oncoprotein RUNX1-ETO with or without a mutated (N822K) KIT protein. The expression of N822K–c-KIT strongly increases the self-renewal capacity of RUNX1-ETO expressing cells. Global analysis of gene expression changes and alterations in the epigenome show that N822K–c-KIT expression profoundly influences the open chromatin landscape and transcription factor binding. However, our experiments also show that double mutant cells still differ from their patient derived counterparts, highlighting the importance of studying patient cells to obtain a true picture of how gene regulatory networks have been reprogrammed during tumourigenesis.
2021-04-01 | GSE152332 | GEO
Project description:The t(8;21) RUNX1-ETO translocation is one of the most frequent cytogenetic abnormalities in acute myeloid leukemia (AML). In RUNX1-ETO(+) patient samples, differing classes of activating c-KIT receptor tyrosine kinase mutations have been observed. The most common (12%-48%) involves mutations, such as D816V, which occur in the tyrosine kinase domain, whereas another involves mutations within exon 8 in a region mediating receptor dimerization (2%-13% of cases). To test whether distinct subtypes of activating c-KIT mutations differ in their leukemogenic potential in association with RUNX1-ETO, we used a retroviral transduction/transplantation model to coexpress RUNX1-ETO with either c-Kit(D814V) or c-Kit(T417I?418-419) in murine hematopoietic stem/progenitor cells used to reconstitute lethally irradiated mice. Analysis of reconstituted animals showed that RUNX1-ETO;c-Kit(D814V) coexpression resulted in 3 nonoverlapping phenotypes. In 45% of animals, a transplantable AML of relatively short latency and frequent granulocytic sarcoma was noted. Other mice exhibited a rapidly fatal myeloproliferative phenotype (35%) or a lethal, short-latency pre-B-cell leukemia (20%). In contrast, RUNX1-ETO;c-Kit(T417I?418-419) coexpression promoted exclusively AML in a fraction (51%) of reconstituted mice. These observations indicate that c-Kit(D814V) promotes a more varied and aggressive leukemic phenotype than c-Kit(T417I?418-419), which may be the result of differing potencies of the activating c-Kit alleles.
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Project description:A network of lineage-specific transcription factors and microRNAs tightly regulates differentiation of hematopoietic stem cells along the distinct lineages. Deregulation of this regulatory network contributes to impaired lineage fidelity and leukemogenesis. We found that the hematopoietic master regulator RUNX1 controls the expression of certain microRNAs, of importance during erythroid/megakaryocytic differentiation. In particular, we show that the erythorid miR144/451 cluster is epigenetically repressed by RUNX1 during megakaryopoiesis. Furthermore, the leukemogenic RUNX1/ETO fusion protein transcriptionally represses the miR144/451 pre-microRNA. Thus RUNX1/ETO contributes to increased expression of miR451 target genes and interferes with normal gene expression during differentiation. Furthermore, we observed that inhibition of RUNX1/ETO in Kasumi1 cells and in RUNX1/ETO positive primary acute myeloid leukemia patient samples leads to up-regulation of miR144/451. RUNX1 thus emerges as a key regulator of a microRNA network, driving differentiation at the megakaryocytic/erythroid branching point. The network is disturbed by the leukemogenic RUNX1/ETO fusion product.
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Project description:The fusion protein RUNX1-ETO is an oncogenic transcription factor generated by t(8;21) chromosome translocation, which is found in FAB-M2-type acute myeloid leukemia (AML). RUNX1-ETO is known to dysregulate the normal RUNX1 transcriptional network, which should involve essential factors for the onset of AML with t(8;21). In this study, we screened for possible transcriptional targets of RUNX1 by reanalysis of public data in silico, and identified C11orf21 as a novel RUNX1 target gene because its expression was down-regulated in the presence of RUNX1-ETO. The expression level of C11orf21 was low in AML patient samples with t(8;21) and in Kasumi-1 cells, which carry RUNX1-ETO. Knockdown of RUNX1-ETO in Kasumi-1 cells restored C11orf21 expression, whereas overexpression of RUNX1 up-regulated C11orf21 expression. In addition, knockdown of RUNX1 in other human leukemia cells without RUNX-ETO, such as K562, led to a decrease in C11orf21 expression. Of note, the C11orf21 promoter sequence contains a consensus sequence for RUNX1 binding and it was activated by exogenously expressed RUNX1 based on our luciferase reporter assay. This luciferase signal was trans-dominantly suppressed by RUNX1-ETO and site-directed mutagenesis of the consensus site abrogated the reporter activity. This study demonstrated that C11orf21 is a novel transcriptional target of RUNX1 and RUNX1-ETO suppressed C11orf21 transcription in t(8;21) AML. Thus, through this in silico approach, we identified a novel transcriptional target of RUNX1, and the depletion of C11orf21, the target gene, may be associated with the onset of t(8;21) AML.
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