Identification of genomic, epigenomic and proteomic targets of the CALM/AF10 fusion protein

Coordinator:    Prof. Dr. med Stefan K. Bohlander
Institution: Universität München, Abt. Medizin III, Krankenhaus Grosshadern
The analysis of chromosomal translocations has contributed immensely to our understanding of the pathogenesis of acute leukemias. The translocation t(10;11)(p12;q14) results in the formation of the CALM/AF10 fusion gene. The CALM/AF10 fusion gene is found in acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL) as well as in some cases of malignant lymphoma. While CALM is a clathrin assembly protein, AF10 is a putative transcription factor of the polycomb group of genes. The CALM/AF10 fusion protein contains the complete CALM and almost the complete AF10 protein with the exeption of the N terminal Plant homeo Domain (PHD) of AF10. The retroviral expression of CALM/FA10 in murine bone marrow cells leads to the development of an aggressive acute leukemia. It is our aim to understand the mechanisms that contribute to CALM/AF10 mediated leukemogenesis. To this end we want to identify the genomic, epignomic and proteomic targets of CALM/AF10. For the identification of the genomic tragets of CALM/AF10 we will inducibly express CALM/AF10 in a cell line system, cross-link to DNA and analyze the DNA bound to CALM/AF10 after immunoprecipitation with promoter arrays or by direct sequencing. Subsequently we will analyze the epigenetic marks of these direct CALM/AF10 target genes (eg. DNA methylation, histone acetylation or methylation). In addition we want to analyze the significance of changes in the expression levels of CALM/AF10 target genes as well as of CALM/AF10 protein interaction partners, which we already identified, for the prognosis of AML patients. Recently we could show that CALM/AF10 expression leads to a global reduction of histone H3 lysine 79 methylation in CALM/AF10 positive patients through its interaction with the histone methyltransferase DOT1L. This was the first description of a global change in histone modification patterns caused by a leukemogenic fusion protein. Since H3K79 methylation is not only involved in transcriptional elongation but also in DNA repair processes, we will ask the qustion whether the expression of CALM/AF10 might lead to an increased genomic instability. It could be that increased genomic instability which leads to secondary oncogenic mutations plays a critical role in the development of leukemia.
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