Expression of MLL-AF4 or AF4-MLL fusions does not impact the efficiency of DNA damage repair
| Autor: | Norma C. Gutiérrez, Aldeheid Bursen, Federico González, Julio Castaño, Pablo Menendez, Rolf Marschalek, Ana B. Herrero |
|---|---|
| Přispěvatelé: | European Commission, Generalitat de Catalunya, Fundación 'la Caixa', Josep Carreras Leukemia Foundation, Asociación Española Contra el Cáncer, Ministerio de Economía y Competitividad (España), Instituto de Salud Carlos III, European Research Council |
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
DNA End-Joining Repair Oncogene Proteins Fusion DNA damage Gene Expression AF4.MLL Biology DSB Histones 03 medical and health sciences 0302 clinical medicine PARP1 Radiation Ionizing hemic and lymphatic diseases medicine Humans Secondary Acute Myeloid Leukemia DNA Breaks Double-Stranded t(4 11) Homologous Recombination neoplasms Etoposide Genetics Ku70 MLL.AF4 Infant Cell Cycle Checkpoints Precursor Cell Lymphoblastic Leukemia-Lymphoma medicine.disease Molecular biology 3. Good health Comet assay Leukemia HEK293 Cells 030104 developmental biology Oncology 030220 oncology & carcinogenesis Comet Assay Infant leukemia Homologous recombination Myeloid-Lymphoid Leukemia Protein Research Paper DNA Damage medicine.drug |
| Zdroj: | Digital.CSIC. Repositorio Institucional del CSIC instname Oncotarget |
| Popis: | The most frequent rearrangement of the human MLL gene fuses MLL to AF4 resulting in high-risk infant B-cell acute lymphoblastic leukemia (B-ALL). MLL fusions are also hallmark oncogenic events in secondary acute myeloid leukemia. They are a direct consequence of mis-repaired DNA double strand breaks (DNA-DSBs) due to defects in the DNA damage response associated with exposure to topoisomerase- II poisons such as etoposide. It has been suggested that MLL fusions render cells susceptible to additional chromosomal damage upon exposure to etoposide. Conversely, the genome-wide mutational landscape in MLL-rearranged infant B-ALL has been reported silent. Thus, whether MLL fusions compromise the recognition and/or repair of DNA damage remains unanswered. Here, the fusion proteins MLL-AF4 (MA4) and AF4-MLL (A4M) were CRISPR/Cas9-genome edited in the AAVS1 locus of HEK293 cells as a model to study MLL fusion-mediated DNA-DSB formation/repair. Repair kinetics of etoposide- and ionizing radiation-induced DSBs was identical in WT, MA4- and A4M-expressing cells, as revealed by flow cytometry, by immunoblot for γH2AX and by comet assay. Accordingly, no differences were observed between WT, MA4- and A4M-expressing cells in the presence of master proteins involved in non-homologous end-joining (NHEJ; i.e.KU86, KU70), alternative-NHEJ (Alt-NHEJ; i.e.LigIIIa, WRN and PARP1), and homologous recombination (HR, i.e.RAD51). Moreover, functional assays revealed identical NHEJ and HR efficiency irrespective of the genotype. Treatment with etoposide consistently induced cell cycle arrest in S/G2/M independent of MA4/A4M expression, revealing a proper activation of the DNA damage checkpoints. Collectively, expression of MA4 or A4M does neither influence DNA signaling nor DNA-DSB repair. This work was supported by the European Research Council to P.M (ERC-2014-CoG-646903), MINECO (SAF2013-43065 to P.M), The Foundation Inocente Inocente and the Spanish Association of Cancer Research (AECC) to P.M and the Deutsche José Carreras Leukämie Stiftung to R.M/P.M. P.M also acknowledges the financial support from The Obra Social La Caixa-Fundaciò Josep Carreras and The Generalitat de Catalunya (SGR330). F.G. is supported by a Ramón y Cajal Grant (RYC-2014-16751) from the Ministry of Economy and Competitiveness (MINECO), Spain. N.C.G and A.B.H acknowledge financial support from The Cooperative Research Thematic Networks (RTICC) (RD12/0036/0058) and the INNOCAMPUS Program (CEI10-1-0010). |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|