The human cytidine deaminase APOBEC3G (A3G) is an innate restriction factor that inhibits human immunodeficiency virus, type 1 (HIV-1) replication. Regulation of A3G gene expression plays an important role in this suppression. Currently, an understanding of the mechanism of this gene regulation is largely unknown. Here, we have identified and characterized a TATA-less core promoter with an NFAT/IRF-4 composite binding site that confers cell type-specific transcriptional regulation. We found that A3G expression is critically dependent on NFATc1/NFATc2 and IRF-4. When either NFATc1 or NFATc2 and IRF-4 were co-expressed, A3G promoter activity was observed in cells that normally lack A3G expression and expression was not detected in the presence of the individual factors. This induced A3G expression allowed normally permissive CEMss cells to adopt a nonpermissive state, able to resist an HIV-1Δvif challenge. This represents the first reporting of manipulating the restrictive state of a cell type via gene regulation. Identification of NFAT and IRF family members as critical regulators of A3G expression offers important insight into the transcriptional control mechanisms that regulate innate immune responses and identifies specific targets for therapeutic intervention aimed at effectively boosting our natural immunity, in the form of a host defensive factor, against HIV-1.