Cell-to-Cell Transmission Can Overcome Multiple Donor and Target Cell Barriers Imposed on Cell-Free HIV
Figure 6
The actin cytoskeleton of Jurkat cells presents a barrier to cell-free HIV infection.
(A, B) MT4, Jurkat and primary CD4 T cells were inoculated with concentrated cell-free HIVNL4-3-GLuc by spinoculation and incubated at 37°C in the presence or absence of increasing concentrations of latrunculin-A (Lat-A) or japlakinolide (Jas)(µM). Luciferase activity was measured 36 h post-inoculation. Data were normalized to DMSO control. Error bars represent the standard error of the mean from 2 experiments. (C) Phalloidin staining of untreated, Lat-A-, and Jas-treated cells. Cells were exposed to Lat-A (1 µM for MT4 and 0.5 µM for Jurkat) or Jas (0.5 µM for MT-4 and 0.0625 µM for Jurkat) for 1 h at 37°C. Note that phalloidin competes with Jas for binding to polymerized actin and further dilution of drug was required to observe actin staining in Jurkat cells [76]. Size bars correspond to 10 µm. (D) Viral binding was measured by α-p24-ELISA after spinoculating cells in the presence or absence of 1 µM Lat-A or Jas. Error bars represent the standard deviation from 3 measurements. (E) Late reverse transcription (RT) was measured by Q-PCR from cells treated with 1 µM of Lat-A or Jas. Error bars represent the standard deviation of 3 late RT measurements. (F, G) A co-culture experiment as in Figure 5B was performed in the presence of increasing concentrations of Lat-A or Jas (µM). Error bars represent the standard error of the mean from 2 experiments.