Abstract
Human Immunodeficiency Virus-1 Vpr, a nonstructural protein incorporated into virus particles, possesses several features contributing to virus replication and cytopathic effects. Vpr induced effects are mediated through its interactions with viral and/or host cellular proteins, in particular, the host protein, Glucocorticoid Receptor (GR). Though GR is known to increase Vpr-mediated HIV-1 transactivation, Vpr-GR interaction and its subcellular localization have not been studied in cells. Towards this, we evaluated Vpr interaction with GR in cells using Bimolecular Fluorescence Complementation analysis by generating chimeric Vpr or GR with the N- and C-terminal fragments of Venus protein. Our results showed that interaction between Vpr and GR requires certain Vpr and GR domains. Specifically, leucine residues in the third helical domain of Vpr and N-terminal domain of GR is involved in Vpr-GR interaction. Altering these residues not only interferes with Vpr-GR interaction, but also prevents translocation of this complex into the nucleus. Further, utilizing a mutant Vpr unable to oligomerize, we show that Vpr oligomerization is essential for optimal interaction with GR. In conclusion, by taking advantage of BiFC system, specific residues in Vpr have been found to be associated with binding GR and the subcellular distribution of Vpr-GR complex.