Intermolecular masking of the HIV-1 Rev NLS by the cellular protein HIC: Novel insights into the regulation of Rev nuclear import

Abstract

BackgroundThe HIV-1 regulatory protein Rev, which is essential for viral replication, mediates the nuclear export of unspliced viral transcripts. Rev nuclear function requires active nucleocytoplasmic shuttling, and Rev nuclear import is mediated by the recognition of its Nuclear Localisation Signal (NLS) by multiple import factors, which include transportin and importin β. However, it remains unclear which nuclear import pathway(s) predominate in vivo, and the cellular environment that modulates Rev nucleocytoplasmic shuttling remains to be characterised.ResultsIn our study, we have identified the cellular protein HIC (Human I-mfa domain-Containing protein) as a novel interactor of HIV-1 Rev. We demonstrate that HIC selectively interferes with Rev NLS interaction with importin β and impedes its nuclear import and function, but does not affect Rev nuclear import mediated by transportin. Hence, the molecular determinants mediating Rev-NLS recognition by importin β and transportin appear to be distinct. Furthermore, we have employed HIC and M9 M, a peptide specifically designed to inhibit the transportin-mediated nuclear import pathway, to characterise Rev nuclear import pathways within different cellular environments. Remarkably, we could show that in 293T, HeLa, COS7, Jurkat, U937, THP-1 and CEM cells, Rev nuclear import is cell type specific and alternatively mediated by transportin or importin β, in a mutually exclusive fashion.ConclusionsRev cytoplasmic sequestration by HIC may represent a novel mechanism for the control of Rev function. These studies highlight that the multivalent nature of the Rev NLS for different import receptors enables Rev to adapt its nuclear trafficking strategy.

The Bovine Immunodeficiency Virus Rev Protein: Identification of a Novel Nuclear Import Pathway and Nuclear Export Signal among Retroviral Rev/Rev-Like Proteins

“…It is noteworthy that importin ␤ was the first described receptor for HIV-1 Rev nuclear import in vitro through its direct interaction with the atypical monopartite, arginine-rich NLS of Rev (58,68). Importin ␤ also has been shown to be an important Rev receptor in vivo (26). Therefore, we wished to determine whether importin ␤ alone could also mediate the nuclear import of BIV Rev.…”

“…The shuttling of HIV-1 Rev into the nucleus is mediated by the direct binding of the protein to the nuclear transport receptors, mainly importin ␤ but also transportin, importin 5, and importin 7 (3). Recent studies showed that importin ␤ and transportin import pathways are at play for the nuclear import of HIV-1 Rev in vivo (26,31). Moreover, the transportin pathway depends on the Nup358 nucleoporin that acts as a dock station (31).…”

Designer peptide delivery systems for gene therapy

“…Gu et al (77) characterized the nuclear import characteristics of the HIV-1 derived Rev peptide across different cell lines, with results suggesting that HeLa, U937 and THP-1 cell lines employed transportin as the major transport receptor to rev, whereas in 293T, Jurkat or CEM cell lines, importin-β was the primary mediator of nuclear uptake. Intranuclear transport characteristics are also known to alter once cells become cancerous (78,79) .…”

Identification of a Karyopherin β1/β2 Proline-Tyrosine Nuclear Localization Signal in Huntingtin Protein

“…Consistent with the concept of an additional class of karyopherin ␤1-and ␤2-dependent regulated NLSs, recently, the HIV-1 Rev protein was shown to import into the nucleus by both karyopherin ␤1-direct (24) and karyopherin ␤2 pathways, with a binding factor that regulates these NLS-transport factor interactions (25). Other proteins, such as the c-Jun oncoprotein, have been seen to interact with both karyopherin ␤1 and ␤2 as well as other import factors in vitro, via a basic stretch of amino acids, but without the PY epitope of a karyopherin ␤2-dependent NLS (26,27).…”