Mechanism and relevance of active DNA methylation during monocyte differentiation

• Applicant: Professor Dr. Michael Rehli
• Subject Area: Hematology, Oncology ; Biochemistry ; Immunology
• Term: from 2015 to 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 278631977

Experimental evidence as well as the high frequency of mutations in TET2 and IDH genes in acute myeloid leukaemia suggests that DNA methylation turnover is essential for normal myelopoiesis. Previous work from others as well as work from our lab suggests a major role for transcription factors in targeting DNA demethylation to lineage-specific enhancers. However, in the large majority of cases it is not actually clear whether the observed demethylation is 'just' the consequence of transcription factor binding, chromatin remodelling and e.g. enhancer activation, or whether it is required for the latter. To understand the relevance and function of DNA demethylation both in normal myeloid cells as well during leukaemogenesis we need to understand better and define, which sequence features and which factors require demethylation to establish stable transcription factor -DNA interactions and to participate in gene regulatory processes. In the current application we propose to study the differentiation of post-mitotic monocytes to address remaining key questions related to the mechanisms of active DNA demethylation. Using a recently established, highly efficient and non-toxic/activating siRNA transfection protocol we will knock-down the expression of key genes (including TET2 as well as candidate targeting transcription factors) and study the effect on transcription, DNA methylation, chromatin accessibility, histone modifications and transcription factor binding. This will ultimately clarify the functional importance of DNA demethylation, and how it is targeted in myeloid cells. The findings will not only be important for the general understanding of monocyte biology but also for our understanding how mutations in the DNA methylation machinery might alter hematopoietic cells to drive leukaemogenesis.

• DFG Programme: Research Grants