Implication of the DotCom complex members AF9 and DOT1L in cerebral cortex development

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Neuronal specification of neural stem cells predominates embryonic development of the stratified cerebral cortex. Neural stem cells reside in two cellular compartments and produce either lower or upper layer neurons. The latter are produced by stem cells from the subventricular zone, which are not well characterised on a molecular level. However, they play a pivotal role during evolution of higher cognitive tasks. In this project we aim to further characterise these determined stem cells.Our preliminary data identified AF9/MLLT3 (myeloid/lymphoid or mixed-lineage leukemia (trithorax homolog, Drosophila); translocated to, 3) as transcription factor that suppresses upper layer fate and influences subventricular progenitor proliferation. We are now addressing the question how AF9 controls progenitor proliferation and differentiation. AF9 acts in different protein complexes: on one hand in the DotCom that contains the histone methyltransferase DOT1L (DOT1-like, histone H3 methyltransferase), and on the other hand in the super elongation complex (SEC) that is implicated in transcriptional elongation. We will generate AF9- and DOT1L-deficient mice and characterise progenitor and neuronal specification in a context of a hierarchical developmental sequence involving a variety of transcription factors. We will further identify target genes for AF9 and DOT1L specifically in subventricular progenitors using high throughput sequencing of RNA. Targets will be classified according to DOT1L-dependent and -independent transcription. Using chromatin-immunoprecipitation we will elucidate genome-wide distribution of specific chromatin modifications, RNA polymerase II, AF9 and DOT1L to define presence of DotCom or SEC proteins at the respective AF9 target gene loci in the context of subventricular progenitor specification. We will thus provide data about transcriptional control through transcription factors that act via chromatin modifications during cortical development. Furthermore we will provide insights into the developmental capacities of a specific stem cell population that gives rise to neuronal networks implicated in higher cognitive tasks.