Fig 5. Rev function in isogenic viruses differing only in their ability to express Rev1-Vpu.

(A) Gene arrangement of a reporter construct expressing GFP in a Rev-dependent manner. The GFP ORF is flanked by splice donor site 4 (D4) and splice acceptor site 7 (A7). Rev mediates the export of intron-containing GFP expressing mRNA via binding to the RRE. (B) Rev-dependent gene expression was determined by co-transfection of HEK293T cells with increasing amounts of the indicated molecular clones of HIV-1, the GFP reporter construct and a BFP expressing control plasmid. 40 h post transfection, GFP expression levels of BFP positive cells were analyzed by flow cytometry. Examples of primary FACS data are shown in the lower panel. (C) Western blot, showing Rev-dependent expression of p24-capsid and Rev-independent expression of Nef in HEK293T cells transfected with the indicated proviral constructs.