2-APB and BAPTA-AM prevent viral-induced FAK phosphorylation. CaSki cells were serum starved overnight, and a synchronized infection with HSV-2(G) was conducted as described in Materials and methods. 100 μM 2-APB, 10 μM nifedipine, or control buffer (5% DMSO) was added to cells at the time of binding or penetration (temperature shift, 15 min, 37°C). Cells were treated with low pH citrate buffer, washed, and cell lysates were prepared 10 min after citrate treatment. Western blots were prepared, probed with anti-FAK pY 397 , and scanned by optical densitometry. Blots were then stripped and reprobed with mAb to total FAK. A representative blot is shown in A. Alternatively, cells were pretreated for 2 h with BAPTA-AM, EGTA, or 5% methanol in PBS (control), and then exposed to HSV-2(G). (B) Lysates were prepared 5 min after infection and analyzed by Western blotting for FAK phosphorylation. (C) Results obtained from three independent experiments are summarized graphically as odu of phosphorylated FAK:odu total FAK as a percentage of control (HSV-2(G) in the absence of any inhibitors). Results are the mean ± SD obtained from three independent experiments; asterisks indicate P