Nuclear export signal of IκBα interferes with the Rev-dependent posttranscriptional regulation of human immunodeficiency virus type I

De novo synthesized IκBα accumulates transiently in the nucleus where it inhibits NF-κB-dependent transcription and reduces nuclear NF-κB content. A sequence present in the C-terminal domain of IκBα and homologous to the HIV-1 Rev nuclear export signal (NES) has been recently defined as a functional NES conferring on IκBα the ability to export IκBα/NF-κB complexes. Rev utilises its RNAbinding activity and NES sequence to promote specifically the transport of unspliced and monospliced viral RNAs to the cytoplasm. The object of this work was to determine if nuclear IκBα could interfere with Rev-dependent transport of viral RNA from the nucleus to the cytoplasm. We report that accumulation of IκBα in the cell nucleus blocks viral replication. This effect could be dissociated from the capacity of IκBα to inhibit NF-κB-DNA-binding activity and required a functional IκBα NES motif. Indeed, mutation of the NES abrogated the capacity of IκBα to inhibit Rev-dependent mechanisms involved in the replication of either wild-type or NF-κB-mutated HIV-1 molecular clones. Nuclear accumulation of a reporter protein tagged with a nuclear localization signal (NLS) and fused to the IκBα NES motif (NLS-PK-NES) was sufficient to inhibit HIV-1 replication at a post-transcriptional level by specifically blocking the expression of a Rev-dependent gene. Furthermore, in cells pulsed with TNF, a treatment which favors nuclear accumulation of newly synthesized IκBα, NLS-PK-NES expression promoted sustained accumulation of nuclear NF-κB lacking DNA-binding activity. This NES-mediated accumulation of inactive nuclear NF-κB is likely the consequence of interference in the IκBαmediated export of NF-κB. These findings indicate that IκBα and Rev compete for the same nuclear export pathway and suggest that nuclear accumulation of IκBα, which would occur during normal physiological cell activation process, may interfere with the Rev-NES-mediated export pathway of viral RNAs, thus inhibiting HIV-1 replication.