Abstract
The reciprocal translocation t(12;21)(p13;q22), the most common structural genomic alteration in B-cell precursor acute lymphoblastic leukaemia in children, results in a chimeric transcription factor TEL-AML1 (ETV6-RUNX1). We identified directly and indirectly regulated target genes utilizing an inducible TEL-AML1 system derived from the murine pro B-cell line BA/F3 and a monoclonal antibody directed against TEL-AML1. By integration of promoter binding identified with chromatin immunoprecipitation (ChIP)-on-chip, gene expression and protein output through microarray technology and stable labelling of amino acids in cell culture, we identified 217 directly and 118 indirectly regulated targets of the TEL-AML1 fusion protein. Directly, but not indirectly, regulated promoters were enriched in AML1-binding sites. The majority of promoter regions were specific for the fusion protein and not bound by native AML1 or TEL. Comparison with gene expression profiles from TEL-AML1-positive patients identified 56 concordantly misregulated genes with negative effects on proliferation and cellular transport mechanisms and positive effects on cellular migration, and stress responses including immunological responses. In summary, this work for the first time gives a comprehensive insight into how TEL-AML1 expression may directly and indirectly contribute to alter cells to become prone for leukemic transformation.
Highlights
We investigated whether the regulation of early direct TEL-AML1 targets identified in our mouse precursor B-cell model persist up to leukaemia-onset in paediatric patients
Several mRNA profiling studies using microarray technology have been used to identify genes differentially regulated in TEL-AML1positive patients in comparison to other subgroups
Our study revealed promoter regions corresponding to 217 genes binding to TEL-AML1 and being differentially expressed on mRNA level after TEL-AML1 induction
Summary
The reciprocal translocation t(12;21)(p13;q22) is the most frequent chromosomal rearrangement in childhood B-cell precursor acute lymphoblastic leukaemia (ALL) with an incidence of B25%.1,2 The resulting TEL-AML1 (syn.: ETV6-RUNX1) expression leads to expansion of B-cell precursors with enhanced self-renewal capacity and impaired differentiation to more mature B-cell stages. The translocation already occurs in utero in an early. B-cell progenitor cell and leads to establishment of a pre-leukemic clone persisting in the bone marrow for several years, insufficient to generate an overt leukaemia.3,9,10 It is unclear if the translocation inevitably leads to the disease or if only a small portion progress, as conflicting reports about the incidence of this translocation in healthy newborns exist.. The global binding pattern of the TEL-AML1 fusion protein on promoter regions in precursor B-cells is not known and several mechanisms of action have been proposed so far. We wanted to differentiate between direct and indirect regulatory effects of the TEL-AML1 fusion protein in a cell system void of secondary aberrations as seen in patients and patient-derived cell lines by using chromatin immunoprecipitation (ChIP)-on-chip to identify promoter-binding sites in combination with mRNA microarray analysis to assess the gene regulatory effect of TEL-AML1. Correspondence: Professor A Borkhardt, Heinrich-Heine University of Dusseldorf, Medical Faculty, Clinic for Pediatric Oncology, Hematology and Clinical Immunology, Moorenstreet 5, 40225 Dusseldorf, Germany
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