Fig. 5: HSV triggers translocation of PDPK1 and PLCγ1 to the outer leaflet and their subsequent phosphorylation is inhibited by CIMSS.

a HaCat cells were mock-infected or synchronously infected with HSV-2(G) in the absence or presence of 10 µM CIMSS. After 15 min incubation, cell surface proteins were biotinylated and precipitated with streptavidin magnetic beads and analyzed by immunoblotting with Abs to pPDPK1^S241 and total PDPK1, pPLCγ1^Y783 and total PLCγ1, pAkt^T308 and total Akt, and FIC-1 (cytosolic protein); controls include whole cell lysates. Results are representative of two independent experiments. b HaCat cells were mock-infected or infected with HSV-2(G) (MOI = 10 PFU/cell) in the presence of 0.1% DMSO or 10 µM CIMSS and at the indicated time post-infection cells were fixed with or without Triton X-100 permeabilization and stained for pPDPK1^S241 (red, upper panel, ZeissLive/DuoScan, objective 100 × 1.4) or pPLCγ1^Y783 (green, lower panel, LeicaSP8, objective 63 × 1.4). Representative images from 2–3 independent experiments are shown (bars = 10 μm).