The Effects of MAPK Signaling on the Development of Cerebellar Granule Cells

The granule cells are the most abundant neuronal type in the human brain. Rapid proliferation of granule cell progenitors results in dramatic expansion and folding of the cerebellar cortex during postnatal development. Mis-regulation of this proliferation process causes medulloblastoma, the most prevalent childhood brain tumor. In the developing cerebellum, granule cells are derived from Atoh1-expressing cells, which arise from the upper rhombic lip (the interface between the roof plate and neuroepithelium). In addition to granule cells, the Atoh1 lineage also gives rise to different types of neurons including cerebellar nuclei neurons. In the current study, I have investigated the mechanisms that regulate the proliferation of cerebellar granule cells and the diversification of the Atoh1 lineage. The research presented in Chapter I elucidates a previously underappreciated role of FGF/MAPK pathway signaling in granule cell (GC) development and cerebellar morphogenesis. Here, we consider the involvement of MAPK signaling in the development of the Atoh1 lineage of granule cell precursors (GCP) in the cerebellum. Using mice with upregulation of the MAPK intermediate, MEK1, and mice with downregulation of the proliferative readout Etv4 gene, the regulatory effects of MAPK signaling were observed and interpreted through phenotype analysis. The results showed distinct morphological differences between the mutants and the wildtypes, including irregularities in foliation patterning of the central lobe, changes in lobule sizes, and discontinuities in the developing external granular layer (EGL) and internal granular layer (IGL). The measured cerebellar foliation index was indeed increased in MEK gain-of-function (MEK-GOF) mutants, but oddly enough the internal granular layer (IGL) area at maturity was decreased compared to the WT. Further investigation of the MEK-GOF mutants revealed ectopic expression of a neural progenitor gene called Sox2 in the EGL of late stage postnatal mice and ectopic expression of the MAPK gene Tlx3, indicating that GCP continue to proliferate longer than expected due to induction of MAPK activity in non-endogenous tissues. Furthermore, analyses showed that sustained GCP presence in the EGL did not seem to affect total GC number, but may contribute to the foliation and expansion phenotypes in seen in MEK-GOF and may also provide insight for division mechanisms in overexpression of MAPK signaling. In addition to examining the signaling and effects of GCP populations during development, the origin of Atoh1 neural precursor lineages was investigated in lineage tracing experiments presented in Chapter II. While the rhombic lip has historically been identified as the sole progenitor region for Atoh1 glutamatergic stem cell lineages, we propose an early developmental origin for subpopulations of glutamatergic Atoh1 cells in the ventricular zone (VZ). Early embryonic analyses of Atoh1 and Cre expression at E10.5 revealed clonal expansion of a novel Atoh1 lineage demonstrating oscillating expression from the VZ prior to commitment as deep cerebellar nuclei (DCN).