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Ebf1 heterozygosity results in increased DNA damage in pro-B cells and their synergistic transformation by Pax5 haploinsufficiency.

ABSTRACT: Early B-cell factor 1 (Ebf1) is a transcription factor with documented dose-dependent functions in normal and malignant B-lymphocyte development. To understand more about the roles of Ebf1 in malignant transformation, we investigated the impact of reduced functional Ebf1 dosage on mouse B-cell progenitors. Gene expression analysis suggested that Ebf1 was involved in the regulation of genes important for DNA repair and cell survival. Investigation of the DNA damage in steady state, as well as after induction of DNA damage by UV light, confirmed that pro-B cells lacking 1 functional allele of Ebf1 display signs of increased DNA damage. This correlated to reduced expression of DNA repair genes including Rad51, and chromatin immunoprecipitation data suggested that Rad51 is a direct target for Ebf1. Although reduced dosage of Ebf1 did not significantly increase tumor formation in mice, a dramatic increase in the frequency of pro-B cell leukemia was observed in mice with combined heterozygous mutations in the Ebf1 and Pax5 genes, revealing a synergistic effect of combined dose reduction of these proteins. Our data suggest that Ebf1 controls DNA repair in a dose-dependent manner providing a possible explanation to the frequent involvement of EBF1 gene loss in human leukemia.

SUBMITTER: Prasad MA

PROVIDER: S-EPMC4481594 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Ebf1 heterozygosity results in increased DNA damage in pro-B cells and their synergistic transformation by Pax5 haploinsufficiency.

Prasad Mahadesh A J MA Ungerbäck Jonas J Åhsberg Josefine J Somasundaram Rajesh R Strid Tobias T Larsson Malin M Månsson Robert R De Paepe Ayla A Lilljebjörn Henrik H Fioretos Thoas T Hagman James J Sigvardsson Mikael M

Blood 20150402 26

Early B-cell factor 1 (Ebf1) is a transcription factor with documented dose-dependent functions in normal and malignant B-lymphocyte development. To understand more about the roles of Ebf1 in malignant transformation, we investigated the impact of reduced functional Ebf1 dosage on mouse B-cell progenitors. Gene expression analysis suggested that Ebf1 was involved in the regulation of genes important for DNA repair and cell survival. Investigation of the DNA damage in steady state, as well as aft ...[more]

PMID: 25838350

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Project description:Ebf1 is a transcription factor with documented, and dose dependent, functions in both normal and malignant B-lymphocyte development. In order to understand more about the role of Ebf1 in malignant transformation, we have investigated the impact of reduced functional Ebf1 dose on early B-cell progenitors. Gene expression analysis in loss and gain of function analysis suggested that Ebf1 was involved in the regulation of genes of importance for DNA repair as well as cell survival. Investigation of the level of DNA damage in steady state as well as after induction of DNA damage by UV light supported that pro-B cells lacking one functional allele of Ebf1 display a reduced ability to repair DNA damage. This was correlated to a reduction in expression of Rad51 and combined analysis of published 4C and chromatin Immuno precipitation data suggested that this gene is a direct target for Ebf1. Even though the lack of one allele of Ebf1 did not result in any dramatic increase of tumor formation, we noted a dramatic increase in the formation of pro-B cell leukemia in mice carrying a combined heterozygote mutation in the Ebf1 and Pax5 genes. Even though the tumors were phenotypically similar and stable, we noted a large degree of molecular heterogeneity well in line with a mechanism involving impaired DNA repair. Our data support the idea that Ebf1 controls homologous DNA repair in a dose dependent manner and that this may explain the frequent involvement of Ebf1 in human leukemia 230238: Briefly, 230238 preB-cell line was infected with pMIG-Ebf1-engrailed or pMIG-control (empty vector). Cells were GFP sorted and 500.000 cells were resuspended in RLT for subsequent RNA-extraction. Analysis were perfomred in triplicate. FL Ebf1-KO: E14.5 fetal liver (FL) Ebf1-/- were isolated and expanded expanded in vitro on OP9 stroma cells on conditions permissive for B-cell development. Expanded cells were infected with pMIG-Ebf1-ER and sorted on GFP. Cells were reseeded on OP9 stroma cells and tamoxifen was added to a final concentration of 0.1 uM. 500.000 cells were harvested and RNA extracted 0, 6, 12 and 24h. Gene expression levels were compared to 0h and analysis were performed in duplicates. Tumor LNs: Enlarged lymphnodes from approximately 30 weeks old C57BL/6 Ebf1+/-Pax5+/- were dissected, homogenized and live frozen. After thawing, 1 million lymphnodes cells were live sorted and resuspended in RLT prior to RNA extraction. 8 samples were analyzed in duplicates. 1 million proB cells from wt C57BL/6 were sorted and used as controls.

Dataset Information

Ebf1 heterozygosity results in increased DNA damage in pro-B cells and their synergistic transformation by Pax5 haploinsufficiency.

Publications

Ebf1 heterozygosity results in increased DNA damage in pro-B cells and their synergistic transformation by Pax5 haploinsufficiency.

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