Figure 7. Targeting extracellular Hsp90 reduces KSHV lytic gene expression and release of infectious virions by PEL cells.

(A) BCBL-1 cells were prepared for flow cytometry experiments following their incubation with a monoclonal antibody recognizing Hsp90-alpha (771Ab) followed by secondary antibodies conjugated to ALEXA-488 (black histograms). For controls, cells were incubated with secondary antibodies alone (gray histograms). (B) BCBL-1 cells were incubated with 0.5 μM of DNo for 48 h with or without 0.6 mM valproic acid (VA), then immunoblot analyses were performed for signaling molecules as above and β-Actin used as an internal control. (C–D) BCBL-1 cells were incubated with 0.5 μM of DNo for 16 h, or with 10 μM U0126 for 1.5 h, then LANA (C) and RTA (D) transcripts quantified by qRT-PCR. (E) BCBL-1 cells were incubated with 0.6 mM valproic acid (VA) for 5 days along with either vehicle (DMSO), 0.5 μM of DNo or 1.0 μM U0126, and RTA transcripts quantified by qRT-PCR. (F) 200 μl of BCBL-1 culture supernatants were collected from groups of (E) then diluted 1:5 and incubated with HeLa cells in 8-well chamber slides for 16 h for LANA IFA as above. Controls included cells incubated with UV-KSHV (UV-K). Error bars represent the S.E.M. for two independent experiments.