Figure 5
From: HIV-1-encoded antisense RNA suppresses viral replication for a prolonged period
Expression of the HIV-1 antisense RNA in various types of HIV-1-infected cells. (A) A schematic representation of positions of ASP-L and ASP mRNA in the antisense strand of HIV-1NL4-3 genome. Arrows indicate the regions of the antisense-specific RT-PCR (R6, R7, R9, and R10). (B and C) Results of antisense-specific RT-PCR at R6 region. Total RNAs from HIV-1NL4-3-newly infected Molt-4 and MAGIC-5A cells, and ACH-2 and OM10.1 chronically infected cell lines were analyzed by the antisense-specific RT-PCR (B). Results of HIV-1NL4-3 infected-PBMCs from three healthy individuals (HIV-1-PHA-blast #1–#3) are shown in (C). Lane1, uninfected cell line or PBMC; Lane 2, HIV-1 infected cell line or PBMC; Lane 3, no RTase control; Lane 4, no RT primer control, Lane 5; amplified from cDNA synthesized with random primers; Lane 6, positive control (amplified from pNL4-3 plasmid DNA or cellular genomic DNA). (D) Termination sites of HIVIIIBASP-L. Summary of 3' RACE analyses in OM10.1 and TPA-stimulated ACH-2 is shown. The upper letters mean transcription termination sites in black (ACH-2), green (OM10.1) and red (HIV-1NL4-3 infected MAGIC-5A cells). The black and green arrows are the major transcript in ACH-2 and OM10.1, respectively. (E) Determination of ASP-L TSS in HIVIIIB. The results of antisense-specific RT-PCR at R9 and R10 are shown. Lanes 1–6 are the same as in Figure 2B. (F) Relative expression levels of the antisense and sense transcripts. Results of the strand-specific qRT-PCR at R7 region are shown to measure the expression levels of the antisense and sense transcripts (sense RNA). Expression levels were normalized by the levels of β-actin gene expression. Results of triplicated experiments are shown with mean ± S.D.