FIG. 10.

HHV-8 utilizes the cytoplasmic dynein motors for trafficking into the cell nucleus. (A) Expression of p50/dynamitin in 293 cells. 293 cells were transfected for 8 h with a p50/dynamitin plasmid expressing a c-Myc-p50 fusion protein and an EGFP gene under the control of the CMV promoter. The expression of c-Myc and EGFP was monitored by IFA with MAbs against c-Myc and GFP. (B) Immunoblot detection of p50 expression in cells transfected with different concentrations of a p50/dynamitin-c-Myc-expressing plasmid by the use of anti-c-Myc antibodies. (C) Overexpression of dynamitin inhibiting the dynein motor blocks infected cell nucleus-associated HHV-8 DNA. 293 cells transfected with the pCMV plasmid (293) and the p50-c-Myc plasmid (293/p50), CV-1 cells transfected with the pCMV plasmid (CV-1) and the p50-c-Myc plasmid (CV-1/p50), and HFF cells preincubated with 100 μM Na₃VO₄ or DMEM were infected with HHV-8 for 3 h. The cells were washed to remove the unbound virus, treated with trypsin-EDTA to remove the bound but noninternalized virus, and subjected to estimates of the HHV-8 DNA copy numbers associated with the infected cell nuclei by real-time DNA PCR as described in the legend to Fig. 3. Each reaction was done in duplicate, and each bar represents the mean ± SD for three experiments.