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. 2014 Jan 9;10(1):e1003891. doi: 10.1371/journal.ppat.1003891

Figure 3. Chk2-mediated S-phase arrest facilitates MVM replication.

Figure 3

(A) siRNA depletion of Chk2 reduces MVM replication. A9 cells were transfected twice with 40 nM of control siRNA (lane 1) or siRNA against Chk2 (lane 2) as described in the Materials and Methods. At the time of release from para-synchronization, cells were infected at an MOI of 2. 24 hours later cells were harvested and split in two for Southern and western blotting. Top panel shows a Southern blot demonstrating reduced monomer replicative form (RF, compare lane 2 to 1). Bottom panel shows a western blot showing Chk2 knockdown (compare lane 2 to 1). Actin was blotted as a loading control. There was a two-fold reduction in viral replication following Chk2 depletion by siRNA (B) Chk2 inhibition reduces MVM replication. A9 cells were pretreated with DMSO (lane 1) or Chk2 inhibitor II (Chk2i, lane 2)) before infection at an MOI of 2. 24 hours later cells were harvested and Southern blotting was performed. Bottom panel was probed using a mitochondrial DNA probe for standardization. There was a 4.5 fold reduction in viral replication in the presence of Chk2 inhibitor compared with DMSO treated cells.