Molecular biology of lentivirus-mediated gene transfer.

In order to better appreciate issues regarding the design and the utility of lentiviral vectors, the lentiviral life cycle and, in particular, how the host cell cycle influences lentiviral replication, will be discussed. Since most of the events in lentiviral replication have been best characterized for the primate lentiviruses, including human immunodeficiency virus-1 (HIV-1), HIV-2 and simian immunodeficiency virus (SIV), the discussion will focus on these viruses. The primate lentiviruses contain ten open reading frames (Fig. 1). The gag open reading frame directs the synthesis of structural virion proteins and proteins which direct the encapsidation of genomic viral RNA. The pol open reading frame encodes the viral enzymes which are involved in synthesis of viral cDNA and which direct the integration of viral into cellular DNA. The env open reading frame encodes the structural envelope proteins which mediate attachment of the virion to the cell surface and fusion of viral with target cell membranes. Sequences within the long terminal repeat (LTR) regulate viral gene expression both at the transcriptional and post-transcriptional levels. The LTR contains cis acting regulatory sequences and sequences which mediate the binding of trans-acting viral regulatory proteins. Gag, pol, and env open reading frames are a basic characteristic of retroviral genomes including primate and non-primate lentiviruses as well as simple animal onco-retroviruses such as murine leukemia virus (Fig. 1). A number of additional small open reading frames distinguish the primate and non-primate lentiviruses from simple animal onco-retroviruses. The Tat and Rev proteins regulate lentiviral gene expression at the transcriptional and post-transcriptional levels respectively (CULLEN 1998; JEANG et al. 1999). Tat protein binds to a cis-acting element (TAR), located within the LTR, to up-regulate the activity of the promoter. Rev recognizes a cis-acting element (RRE) located in the central portion of the viral envelope gene to posttranscriptionally regulate viral gene expression. The remaining open reading frames encode what are referred to as the accessory or auxiliary proteins. These terms are somewhat of a misnomer since they imply that these proteins facilitate, but are not essential for, viral replication. However, the Vif protein, which is common to all lentiviruses, with the exception of EIAV, is essential for the replication of the primate lentiviruses (BORMAN et al. 1995; COURCOUL et al. 1995; HARMACHE et al. 1995; REDDY et al. 1995). Activities of these accessory proteins have been comprehensively reviewed elsewhere (TRONO 1995; EMERMAN and MALIM 1998). Although, with the exception of Vif, the accessory proteins appear dispensable for viral replication and pathogenicity in vivo (DESROSIERS et al. 1998) they are likely to contribute to some unique aspects of primate lentiviral biology. For example, and as will be discussed later, the Vpr/Vpx proteins may facilitate the entry of the primate lentiviruses into non-dividing cells by promoting nuclear uptake of the viral genome. Collectively, these unique proteins contribute to viral fitness in that they allow the virus to adapt, or to function within, inhospitable environments. As such, loss of any of these functions would impair the ability of the virus to compete with its wild-type counterpart, ultimately leading to loss of the viral variant from the virus population. This point will become more apparent as the overlapping functions of some of the viral proteins in promoting viral entry into non-dividing cells are discussed. What may be considered redundant features of the viral genome may, in a competitive setting, confer distinct fitness advantages which allow viruses bearing these apparent redundancies to predominate. Ultimately, viral evolution preserves the fittest viruses and the fact that primate lentiviruses have several determinants which may promote nuclear uptake of the viral genome underscores the essential contribution of these proteins to viral replication and persistence in the host.