Background: Unintegrated viral DNA is generally transcriptionally inert but can be expressed under some circumstances and may play a significant role in HIV-1 pathogenesis.

Methods: We determined whether HIV-1 Vpr may be involved in mediating gene expression from unintegrated HIV-1. Cells were infected with VSV-G pseudotyped integrase-defective, vpr-defective HIV-1, and Vpr expressed either de novo from a lentiviral vector or packaged into virions.

Results: We find that Vpr, either expressed de novo or released from virions following viral entry, is essential for unintegrated viral DNA expression. HIV-1 mutants defective for integration in either the integrase catalytic domain or the cis-acting att sites can express at levels similar to wild-type HIV-1 but only in the presence of Vpr. Vpr activation of expression does not correlate with Vpr-mediated cell cycle arrest, nor does it require functional Tat. Vpr-mediated enhancement of expression from integrase-defective HIV-1 is dependent upon the presence of a transcriptionally active HIV-1 and results in increased transcription of viral RNA.

Conclusions: We propose that Vpr facilitates expression of integrase-defective HIV-1 by specific transactivation of unintegrated viral DNA. These results attribute a new function to HIV-1 Vpr and provide a mechanism by which unintegrated HIV-1 DNA expression occurs.