Previous studies have shown that in human T-cells (Jurkat) and hepatoma cells (HepG2), exogenous NF-IL6 can activate HIV-1 gene expression even in the absence of its consensus binding elements in the viral long terminal repeat (LTR). To identify the LTR elements that mediate this response, we have analysed constructs containing mutated and deleted LTR sequences. We have also examined heterologous plasmids to evaluate a potential requirement for the natural LTR sequences in producing HIV-1 gene activation by NF-IL6. As observed for Jurkat and HepG2 cells, we find that in the absence of NF-IL6 binding elements, NF-IL6 can elicit LTR-mediated gene expression in cotransient expression assays performed in monocytic (U937) cells. However, we detect distinct modes of regulation depending on cell type. In U937 cells, the basal LTR sequences retain a significant fraction (42%) of NF-IL6 responsiveness in the absence of upstream regulatory elements in the LTR while these elements are required for maximal response. In HepG2 cells, NF-IL6 elicits a relatively low level of gene activation from the basal LTR elements; response to NF-IL6 is restored with either the Sp1 binding sequences or the other upstream regulatory elements in the LTR. In addition, even though NF-IL6 induces a relatively low gene activity from the basal LTR sequences analysed in HepG2 cells, study of a heterologous construct indicates that these sequences are required for the responsiveness of Sp1 elements to NF-IL6 in this cellular background.