Introduction Since the recognition of AIDS in
Introduction Since the recognition of AIDS in 1981 more than thirty-five years ago, nearly 70 million individuals have been infected with human immunodeficiency virus type 1 (HIV) and roughly half have died (http://www.unaids.org/en/resources/documents/2016/AIDS-by-the-numbers). Although the introduction of HAART two decades ago has been truly transformative, HIV disease, once invariably fatal, remains incurable. The development and testing of a safe and efficacious prophylactic HIV vaccine, a long-sought goal, has been stymied by the absence of a small animal model. Although chimpanzees can be infected with HIV there is now a moratorium on their experimentation (Kaiser, 2015), and the use of simian-HIV (SHIV) hybrids in rhesus macaques is limited by animal availability and expense. Ideally, a fully permissive small animal model would allow higher throughput testing of candidate vaccines and correlates of immunity, if any. Unfortunately, current humanized mouse models, although highly sophisticated and informative (Denton and Garcia, 2011), do not allow for vaccine testing. Most rodent species, including the mouse, have multiple blocks to HIV replication, including at the level of viral entry and transcriptional elongation. Even when those obstacles are circumvented using entry factors and human cyclin T1, which allows for high level viral RNA production, very little infectious HIV is produced from murine AZD-5438 australia (Coskun et al., 2007, Coskun et al., 2006, Elinav et al., 2012, Sherer et al., 2011, Swanson et al., 2010). We and others have pinpointed a major post-integration block in mouse cells at the level of unspliced and partially spliced viral RNA nuclear export (Elinav et al., 2012, Sherer et al., 2011, Swanson et al., 2010). All retroviruses require nuclear export of intron-containing mRNA for productive, high level replication, and for the lentiviruses this process requires a cis-acting RNA sequence and trans-acting viral and cellular proteins. HIV Rev is an essential regulatory protein that is highly conserved among all viral isolates and clades Malim et al., 1989a, Malim et al., 1989b, Malim et al., 1989c. It is encoded by a fully spliced mRNA, and is in a different reading frame than Tat, but shares precisely the same major intron. After cytosolic translation, Rev is imported into the nucleus, where it multimerizes on the Rev-response element (RRE, present within that same intron) to allow nuclear export of unspliced and partially spliced viral mRNAs, including genomic RNA (Madore et al., 1994; Malim and Cullen, 1991; Mann et al., 1994; Szilvay et al., 1997; Vercruysse and Daelemans, 2013). Based upon biochemical analysis it is thought that 6–8 Revs cooperatively bind a single RRE (Cook et al., 1991; Daly et al., 1989; Daugherty et al., 2008; Daugherty et al., 2010; Heaphy et al., 1991; Holland et al., 1990; Wingfield et al., 1991); a low resolution structure has demonstrated a Rev dimer binding across a bent RRE (Fang et al., 2013). In the nucleus Rev-RRE complex interacts with host factors chromosome region maintenance 1 (Crm1) and Ran-GTP, and in the cytosol the complex dissociates to be recycled to the nucleus to export additional cargo, leaving RRE-containing viral mRNA in the cytosol. In the absence of Rev multiple viral proteins cannot be synthesized, including Gag, Pol, and Env, and genomic RNA is trapped in the nucleus or spliced. Interestingly there are no cellular homologs of Rev. Only rarely are intron-containing cellular mRNAs exported to the cytosol via an analogous system (Li et al., 2006, Wang et al., 2015), whereas all lentiviruses routinely use the Rev-RRE complex for trafficking of viral mRNAs with retained introns. We had previously observed a decrease in unspliced, intron-containing HIV mRNA in infected murine cells (Coskun et al., 2007, Coskun et al., 2006, Elinav et al., 2012). The presence of human chromosome 2 in murine cells largely reversed this defect and significantly enhanced infectious virus production (Coskun et al., 2006). We and other investigators identified human (h)Crm1 as the likely gene product on chromosome 2 responsible for this effect (Elinav et al., 2012, Nagai-Fukataki et al., 2011, Okada et al., 2009, Sherer et al., 2011). Expression of hCrm1 in murine cells allowed export of intron-containing HIV mRNAs from the nucleus and significantly boosted virus production, whereas murine (m) Crm1 was non-functional. In our hands this defect mapped to HEAT (Huntingtin, elongation factor 3, protein phosphatase 2A, and the yeast kinase TOR1) repeat 9A of hCrm1, specifically to amino acid residues 411, 412, and 414 (Elinav et al., 2012). The effect of hCrm1 was even more pronounced using feline immunodeficiency virus, and hCrm1 acted more than additively with hSRp40, a serine-arginine rich splicing factor, to increase infectious virus production from mouse cells. More recently it was demonstrated that the addition of a second nuclear export signal to HIV Rev allowed for both enhanced HIV Capsid (CA) production and infectious virus release from murine cells (Aligeti et al., 2014), consistent with a fundamental defect in nuclear export of viral mRNAs in rodent cells.