Thesis (Ph.D.)--University of Washington, 2012Research in the area of Human Immunodeficiency Virus type 1 (HIV-1) vaccine development is hindered by the limitations of the Simian/Human Immunodeficiency Viruses (SHIVs) used in macaque studies. The relevance of SHIV infection of macaques to HIV-1 infection in humans depends on how closely SHIVs mimic aspects of HIV-1 transmission, pathogenesis, and diversity - properties partly determined by the HIV-1 envelope (Env). The inclusion of relevant Envs in SHIVs has proven to be difficult as many HIV-1 Envs are unable to mediate efficient infection of macaque cells. To better understand barriers to HIV-1 Env in macaque cells, we adapted a subtype A based SHIV (SHIV-A) for replication in pig-tailed macaque (Pt) lymphocytes, and identified two mutations in gp120, A204E and G312V, that increased replication by >100-fold. Introduction of these changes into multiple subtype A Envs also greatly increased entry into Pt cells. A204E and G312V Env variants continued to require CCR5 for entry, displayed sensitivity to neutralization by soluble CD4 (sCD4), and all showed a greatly increased ability to use Pt CD4. These findings hinted at the inefficient use of CD4 as an unappreciated restriction to HIV-1 replication in macaque cells and identified changes to subtype A Envs that allowed for increased infection of macaque cells, which has direct implications for the development of a successful SHIV-A. In an effort to develop a SHIV-A for in vivo use, the A204E and G312V Env variants were introduced to SHIVs, which were further assessed for their ability to mediate spreading infection in macaque cells, as well as their neutralization properties. Two SHIV-As were identified that warrant further evaluation in vivo. To establish whether CD4 is a determinant for limited infection of macaque cells by other globally relevant HIV-1 subtypes, HIV-1 Envs representative of diverse circulating strains from subtypes A to D were assayed for their ability to infect cells expressing Pt CD4 or Rhesus (Rh) CD4. Most of the 39 Envs tested (>74%) showed a >10-fold decrease in their ability to mediate infection using macaque CD4 as compared to human CD4. Infectivity using macaque CD4 was highly associated with sensitivity to sCD4 and the ability to mediate infection utilizing low levels of CD4 (p < 0.0001), thus identifying properties of HIV-1 Envs that allow for the increased ability to use macaque CD4. Additionally, the determinants of CD4 sufficient for increased infection by HIV-1 were mapped to a single amino acid difference in the D1 domain. This demonstrates that the inefficient use of macaque CD4 acts as a potent barrier to replication mediated by Envs from circulating strains of HIV-1 and identifies characteristics of HIV-1 Envs that may allow for the successful creation of relevant SHIV models
