1. Functional dissection of epigenetic modifications of histones during development of the cerebral cortex
Precursor cells residing in ventricular and subventricular zones give rise to the adult cerebral cortex that consists of six layers formed in an “inside-out manner” during development. Functionally specialized areas of the neocortex are formed through a gradient of gene expression in the neuroepithelium. This relies on a time-dependent cell fate decision of precursor cells. We have highlighted the contribution of histone methylation in the neuronal specification of upper layer neurons in an Af9-mutant mouse. Af9-interaction partner Dot1l regulates the expression of the transcription factor Tbr1 through methylation of Histone H3 at position K79. Our interests are to further elucidate the transcriptional network of Af9, Dot1l and Tbr1 and their implication in cortical development. Further aims of the project are to analyse differences in progenitors of motor and visual cortex, especially with regard to gene expression and histone modifications.
2. Functional relevance of Tgfβ-signalling for the development of the cerebral cortex
Transforming growth factor β (Tgfβ) mediated neurogenesis has been reported to be increased in older (E16.5) embryonic cortical progenitor cells as compared to that in E13.5 and adult spheres. In this project we aim to reveal key genes that highlight what function Tgfβ-signalling mediates during cortical and hippocampal development and how Tgfβ affects neurogenesis in an age-dependent manner. We are using microarrays with RNA extracted from Tgfβ-treated primary cortical cells from different embryonic stages. Clusters of regulated genes that are categorized into genes affecting neuronal differentiation, proliferation, apoptosis, transcription factors and genes expressed in the extracellular matrix which are validated for regulations through Tgfβ via semiquantitative and qRT-PCR, as well as in situ hybridizations and immunohistochemical stainings in mutant mice with defects of Tgfβ-signalling at different levels. Virus-mediated overexpression and siRNA-mediated knock down of Tgfβ-target genes will complete our efforts to shed further light into the role of Tgfβ for cortical development. We are also aiming to investigate whether interference with other signalling pathways induces a competence of adult spheres to respond with increased neuronal differentiation upon Tgfβ-signalling.
3. Implication of Tgfβ in coupling neurovascular development in the forebrain
Neurogenesis and angiogenesis are both processes influenced by the Tgfβ-signalling pathway. To investigate functions of Tgfβ in forebrain development in vivo, we use a cre-expressing mouse line to conditionally knock-out Tgfbr2. Mutant mice display severe haemorrhages mostly in the telencephalon and diencephalon beginning around E13.5. The embryos die between E16 to E17, when the entire forebrain is infiltrated with blood cells. To investigate this neurovascular defect we characterise these mutants with regard to angiogenesis and neurogenesis, changes in extracellular matrix and cytoskeleton composition, as well as progenitor proliferation and apoptosis. A further aim is to decipher the network of molecules influenced and connected through Tgfβ-signalling.