Neural precursor cells differentiating in the absence of Rb exhibit delayed terminal mitosis and deregulated E2F 1 and 3 activity

Abstract

The severe neurological deficit in embryos carrying null mutations for the retinoblastoma (Rb) gene suggests that Rb plays a crucial role in neurogenesis. While developing neurons undergo apoptosis in vivo neural precursor cells cultured from Rb-deficient embryos appear to differentiate and survive. To determine whether Rb is an essential regulator of the intrinsic pathway modulating terminal mitosis we examined the terminal differentiation of primary cortical progenitor cells and bFGF-dependent neural stem cells derived from Rb-deficient mice. Although Rb -/- neural precursor cells are able to differentiate in vitro we show that these cells exhibit a significant delay in terminal mitosis relative to wild-type cells. Furthermore, Rb -/- cells surviving in vitro exhibit an upregulation of p107 that is found in complexes with E2F3. This suggests that p107 may partially compensate for the loss of Rb in neural precursor cells. Functional ablation of Rb family proteins by adenovirus-mediated delivery of an E1A N-terminal mutant results in apoptosis in Rb-deficient cells, consistent with the interpretation that other Rb family proteins may facilitate differentiation and survival. While p107 is upregulated and interacts with the putative Rb target E2F3 in neural precursor cells, our results indicate that it clearly cannot restore normal E2F regulation. Rb-deficient cells exhibit a significant enhancement of E2F 1 and 3 activity throughout differentiation concomitant with the aberrant expression of E2F-inducible genes. In these studies we show that Rb is essential for the regulation of E2F 1 and 3 activity as well as the onset of terminal mitosis in neural precursor cells.