Fig. 2.

Adenoviral expression of N17Rac1 induces apoptosis of CGNs: dominant-negative mutants of RhoA or Cdc42 do not cause significant cell death. (a) On day 4 in vitro, CGN cultures were infected with adenovirus expressing either GFP alone or in combination with dominant-negative N17Rac1. At 72 h post-infection, CGNs were fixed in paraformaldehyde and nuclei were stained with Hoechst dye. Images shown represent composites of two fields each for GFP or GFP/N17Rac1. Infected cells are indicated by the arrows. CGNs infected with adenovirus expressing GFP alone showed large intact nuclei characteristic of healthy cells (upper panels). In contrast, CGNs infected with adenovirus expressing N17Rac1 displayed marked chromatin condensation and fragmentation indicative of apoptotic cell death (lower panels). Note that non-infected cells in the N17Rac1 condition remained healthy. Scale bar, 10 μm. (b) Quantitation of CGN apoptosis in GFP-expressing cells following 72 h of infection with adenoviruses expressing either GFP alone or in combination with dominant-negative mutants of Rac1, Cdc42, or RhoA at a final titer of 50 infectious particles per cell. Using these conditions, each of the adenoviruses infected approximately 15-20% of the CGNs in the cultures. The results shown represent the means ± SEM for four experiments, each conducted on triplicate coverslips in which approximately 100 GFP-positive CGNs were counted per condition per coverslip. *Significantly different from adenoviral GFP alone (p < 0.01).