Figure 6

TCERG1 is required for HIV-1 replication. A. PBLs and Jurkat cells were transiently transfected with the infectious clone pNL4-3-Renilla along with pGeneClip-shTCERG1-C1 or pGeneClip-shTCERG1-3 and pGeneClip-shTCERG1-4, and the latter pair together with pGeneClip-shTCERG1-1. Renilla was measured 72 hours after transfection, and RLUs are expressed relative to the control cells. B. Expression of TCERG1 protein was assessed by Western blot in both PBLs and Jurkat. C. Jurkat cells were co-transfected with pNL4-3-Renilla and the pEFBOST7-TCERG1 expression vector or empty pcDNA3 as the basal control. Overexpression of TCERG1 was assessed by Western blot. D. Jurkat cells with stable TCERG1 mRNA interference (Jurkat shTCERG1-(3-4) and Jurkat shTCERG1-(1-3-4)) were transfected with pNL4-3-Renilla. Renilla activity was measured in the infected cells (left panel), and p24 was measured in the culture supernatants (right panel); the data are expressed relative to the control cells. E. The TCERG1 interference was analyzed in Jurkat shTCERG1-(3-4) and Jurkat shTCERG1-(1-3-4) by semiquantitative RT-PCR and Western blot. F. The rescue of HIV-1 replication in Jurkat cells with stable TCERG1 mRNA interference (Jurkat shTCERG1-(3-4) and Jurkat shTCERG1-(1-3-4)) transfected with pNL4-3_renilla vector was assessed by co-transfection with a TCERG1 expression vector (right panel), in comparison with cells transfected with pcDNA3 as negative control (left panel). Renilla activity was measured and represented as RLUs; the data are expressed relative to the control cells. The TCERG1 interference and overexpression was determined by Western blot. In all cases, the data from three different experiments is represented in the histograms (mean ± SEM). Statistical analyses were performed and are shown as *, p < 0.05; **, p < 0.01; ***, p < 0.001.