Targeted HIV-1 Reservoirs Figure 2. Inhibition of Akt phosphorylation by ALP compounds in HIV-1 vector-transduced or Tat expressing CHME5 cells. (A) CHME5 cells transduced with DHIV-GFP vector (D3) giving ,90% transduction and (B) Tat expressing CHME5 subline cells were exposed to LPS (50 mg/ml) and CHX (10 mg/ml) (‘‘L’’) in the presence and absence of 25 mM ALP compounds for 24 hours. For the transduced cells (A), the ratios between the levels of phospho-specific and total Akt proteins in the western blots were determined in triplicate, and these ratios were normalized by the ratio in the HIV-1 vector transduced control cells without any treatment (‘‘C’’). For the Tat expressing subline cells (B), the phospho-Akt levels were determined by ELISA-based assay, and the phospho-Akt level in the HIV-1 vector transduced control CHME5 cells with no treatment was used for signal normalization. L: LPS and CHX stress treatment, PF: perifosine, EF: edelfosine, MF: miltefosine. doi:10.1371/journal.pone.0013121.g002 Figure 3. Inhibition of Akt kinase activity by ALP compounds in HIV-1 vector transduced or Tat expressing CHME5 cells. (A) CHME5 cells transduced with DHIV-GFP giving ,90% transduction and (B) Tat expressing CHME5 subline cells were exposed to LPS (50 mg/ml)/CHX (10 mg/ ml) in the presence or absence of the ALP compounds for 24 hours. Ratios between the levels of phospho-specific and total GSK3b proteins were determined and normalized as described in Figure 2. C: control cells not treated with stress, L: LPS and CHX stress treatment, PF: perifosine, EF: edelfosine, MF: miltefosine. doi:10.1371/journal.pone.0013121.g003 PLoS ONE | www.plosone.org 4 September 2010 | Volume 5 | Issue 9 | e13121