[P1.49]: A requirement for pRb during olfactory bulb and hippocampal neurogenesis

Abstract

Recent evidence suggests that cell cycle proteins may have novel functions beyond the control of cell division. We have investigated the role of Rb/E2F pathway in the regulation of neuronal differentiation and migration during late embryonic development.We show that loss of Rb leads to terminal differentiation and radial migration defects as well as loss of specific interneuron subtypes in the developing forebrain and olfactory bulb. This phenotype is linked to a dramatic reduction in the levels of Dlx homeodomain genes that regulate ventral telencephalic development, most significantly Dlx2. To ask if Rb plays a direct role in controlling the induction of Dlx2, we examined the regulatory regions of the Dlx1/Dlx2 locus. Using chromatin immunoprecipitation experiments, we show that Rb modulates Dlx gene expression through interaction with the Dlx forebrain-specific enhancer, I12b, the Dlx2 proximal promoter and 3'UTR region in vivo. This interaction is mediated by E2F functional sites located in I12b that act as repressor sites. Deletion of E2F consensus sites on the I12b-Dlx1/Dlx2 enhancer results in increased reporter activity in the subventricular zone of the developing brain. We demonstrate that in the absence of Rb, E2F7, an Rb-independent repressor, is upregulated in the brain and could ectopically repress the I12b activity and Dlx2 transcription.In conclusion, our data provides the first evidence that cell cycle proteins such as Rb play an essential role to coordinate the transition from proliferation to differentiation and maintain terminal differentiation by regulating the levels of key transcription factors such as Dlx2 during neurogenesis. This work was supported by a grant from CIHR.The specification of unique neuronal and glial sub-types relies upon the restricted expression of cell fate determinants, acting primarily in 'combinatorial codes'. The expression of the proper combinatorial code in each sub-type in turn relies upon the combinatorial action of upstream regulators, acting in a positional and temporal manner. While many players acting at these different levels have been identified, the complex multi-step regulatory flow acting to dictate any one unique cell fate in the nervous system has not been deciphered. Moreover, because each sub-type is generated in distinct numbers, such in-depth decoding of the neural diversification process also requires addressing how upstream and downstream combinatorial codes intersect with the cell cycle and cell death machineries.To address these issues, we are using a specific Drosophila CNS progenitor cell, the neuroblast 5-6, as a model. This neuroblast is presented in all 18 segments of the developing fly CNS, but generates a unique group of neurons, the Apterous cluster of four neurons, only in the three thoracic segments. Two of the Ap cluster cells are furthermore uniquely identifiable by their selective expression of the FMRFa and Nplp1 neuropeptide genes. Specification of Ap neurons requires a complex interplay between positional and temporal cues, whic...