FIG. 1.
Tax enhances replication of the HIV-1 provirus with a truncated nef gene in human T lymphocytes. (A) Schematic representation of the truncated-nef infectious provirus of HIV-1 and the HIV-1 LTR-SEAP reporter. The truncated-nef HIV-1 provirus (pNL4-3neo) is derived from pNL4-3 in which a neo gene fragment was inserted into the XhoI site at the nef locus of pNL4-3 at amino acid position 35. The nef gene is truncated and encodes only the first N-terminal 35 amino acids of the Nef protein. The complete LTR of HIV-1 was derived from pNL4-3 and was constructed by PCR with Pfu DNA polymerase. The SEAP reporter gene (14) is linked to the full-length LTR. (B) Flag-tagged wild-type Tax (Flag-Tax) and frameshift mutant (Flag-TaxFS) were constructed in a phEFneo vector in which the human elongation factor promoter controls the expression of Tax. Tax (1 μg) or TaxFS (1 μg) was cotransfected with or without pNL4-3neo (0.1 μg), together with pHIV-1 LTR-SEAP (0.2 μg) into Jurkat TAg cells by using DMRIE-C liposome transfection reagent. SEAP activity was quantified by using chemiluminescent substrate. The basal activity of the LTR was set at 1. The data are representative of at least two independent experiments.