Functional dissection of CCR5 coreceptor function through the use of CD4-independent simian immunodeficiency virus strains

With rare exceptions, all simian immunodeficiency virus (SIV) strains can u se CCR5 as a coreceptor along with CD4 for viral infection. In addition, ma ny SN strains are capable of using CCR5 as a primary receptor to infect CD4 -negative cells such as rhesus brain capillary endothelial cells. By using coupled fluorescence-activated cell sorter (FACS) and infection assays, we found that even very low levels of CCR5 expression could support CD4-indepe ndent virus infection. CD4-independent viruses represent valuable tools for finely dissecting interactions between Env and CCR5 which may otherwise be masked due to the stabilization of these contacts by Env-CD4 binding. Base d on the ability of SIV Env to bind to and mediate infection of cells expre ssing CCR5 chimeras and mutants, we identified the N terminus of CCR5 as a critical domain for direct Env binding and for supporting CD4-independent v irus infection, However, the activity of N-terminal domain CCR5 mutants cou ld be rescued by the presence of CD4, indicating that other regions of CCR5 are important for post-binding events that lead to viral entry. Rhesus CCR 5 supported CD4-independent infection and direct Env binding more efficient ly than did human CCR5 due to a single amino acid difference in the N termi nus. Interestingly, uncleaved, oligomeric SIV Env protein bound to both CD4 and CCR5 less efficiently than did monomeric gp120, Finally, several mutat ions present in chronically infected monkey populations are shown to decrea se the ability of CCR5 to serve as a primary viral receptor for the SIV iso lates examined.