FIG 1.

Experimental timeline of neuronal differentiation, infection, latency, and reactivation. (A) Experimental timeline of acute infection. LUHMES neuronal cells were plated and allowed to proliferate for 3 days, followed by 5 days of differentiation, as described in Materials and Methods. Once differentiated, cells were infected with either 17syn⁺ or KOS(M) at an MOI of 5 for 1 h, with harvesting of DNA/RNA at the indicated times. (B) Experimental timeline of latency and reactivation. LUHMES neuronal cells were plated and allowed to proliferate for 3 days, followed by 5 days of differentiation. Once differentiated, cells were pretreated for 2 h in medium containing 50 μM acyclovir (ACV). Following pretreatment with ACV, cells were infected with either 17syn⁺ or KOS(M) at an MOI of 5 in medium containing 50 μM ACV. Forty-eight hours later, the medium was changed to medium without ACV and the infection was allowed to proceed, with harvesting at the latent time point 8 days postinfection (labeled 0 hours postreactivation). Reactivation was induced at day 8 postinfection (labeled as day 16) with 10 μM final concentration of Ly294002, and reactivation samples were harvested at the indicated times.