The role of the retinoblastoma protein in cortical neurogenesis

Abstract

The retinoblastoma (Rb) protein was known to be a critical cell cycle regulator in tumour cells, however, little was known regarding its role in normal neural development. To determine the importance of the Rb signalling pathway in neural precursor proliferation and differentiation, I first characterized the key cell cycle regulators and examined the requirement for Rb in these processes. In contrast to previous studies, dominant negative CDK4/6 mutants induced mitotic arrest in neural precursors, despite the binding and sequestration of the Cip/Kip CKIs, p21Cip1 and p27 Kip1. The activity of these mutants was Rb-dependent, indicating that activation of the Rb pathway was sufficient to cause cell cycle arrest in neural precursor cells. While the phenotype of Rb germline knockouts indicated a requirement for Rb in neural precursor differentiation and survival, embryonic lethality and contradictions in the literature precluded a definitive answer. To assess the neural requirement for Rb in the absence of pleiotropic defects, I examined a conditional knockout in which Rb was specifically deleted in the developing telencephalon. These mutants survived to birth and exhibited minimal apoptosis. Although Rb-/- progenitor cells divided ectopically, they were able to survive and differentiate. Due to increased neuroblast proliferation, mutant telencephalic lobes were significantly enlarged. These studies demonstrated that cell cycle deregulation during differentiation does not necessitate apoptosis and terminal mitosis may not be required to initiate differentiation. I next examined the impact of Rb deficiency on cortical development and neuronal differentiation. By histological examination and in situ analyses, Rb mutants were found to exhibit defective laminar organization and region-specific cellularity. Although the majority of cortical neurons survived in the absence of Rb, specific populations, such as Cajal-Retzius neurons, required Rb for survival. By the selective reduction of calbindin- and Lhx6-positive interneurons from the cortical MZ, Rb was shown to regulate these tangentially migrating populations. These studies revealed a role for Rb in the development of cortical architecture, neuronal migration and its requirement in diverse neuronal populations. Together, my studies demonstrated critical functions for Rb in the developing cortex, from the regulation of precursor differentiation to cell type-specific requirements in neuronal survival and migration.