A CRITICAL TEMPORAL REQUIREMENT FOR THE RETINOBLASTOMA PROTEIN FAMILYDURING NEURONAL DETERMINATION

In this report, we have examined the requirement for the retinoblastom a (Rb) gene family in neuronal determination with a focus on the devel oping neocortex. To determine whether pRb is required for neuronal det ermination in vivo, we crossed the Rb-/- mice with transgenic mice exp ressing beta-galactosidase from the early, panneuronal T alpha 1 alpha -tubulin promoter (T alpha 1:nlacZ). In E12.5 Rb-/- embryos, the T alp ha 1:nlacZ transgene was robustly expressed throughout the developing nervous system. However, by E14.5, there were perturbations in T alpha 1:nlacZ expression throughout the nervous system, including deficits in the forebrain and retina. To more precisely define the temporal req uirement for pRb in neuronal determination, we functionally ablated th e pRb family in wild-type cortical progenitor cells that undergo the t ransition to postmitotic neurons in vitro by expression of a mutant ad enovirus E1A protein. These studies revealed that induction of T alpha 1:nlacZ did not require proteins of the pRb family. However, in their absence, determined, T alpha 1:nlacZ-positive cortical neurons underw ent apoptosis, presumably as a consequence of ''mixed signals'' derivi ng from their inability to undergo terminal mitosis. In contrast, when the pRb family was ablated in postmitotic cortical neurons, there was no effect on neuronal survival, nor did it cause the postmitotic neur ons to reenter the cell cycle. Together, these studies define a critic al temporal window of requirement for the pRb family; these proteins a re not required for induction of neuronal gene expression or for the m aintenance of postmitotic neurons, but are essential for determined ne urons to exit the cell cycle and survive.