A comprehensive approach to mapping the interacting surfaces of murine amphotropic and feline subgroup B leukemia viruses with their cell surface receptors

Because mutations in envelope glycoproteins of retroviruses or in their cel l surface receptors can eliminate function by multiple mechanisms, it has b een difficult to unambiguously identify sites for their interactions by sit e-directed mutagenesis. Recently, we developed a gain-of-function approach to overcome this problem. Our strategy relies on the fact that feline leuke mia virus subgroup B (FeLV-B) and amphotropic murine leukemia virus (A-MLV) have closely related gp70 surface envelope glycoproteins and use related N a+-dependent phosphate symporters, Pit1 and Pit2, respectively, as their re ceptors. We previously observed that FeLV-B/A-MLV envelope glycoprotein chi meras spliced between the variable regions VRA and VRB were unable to use P it1 or Pit2 as a receptor but could efficiently use specific Pit1/Pit2 chim eras. The latter study suggested that the VRA of A-MLV and FeLV-B functiona lly interact with the presumptive extracellular loops 4 and 5 (ECL4 and -5) of their respective receptors, whereas VRB interacts with ECL2. We also fo und that FeLV-B gp70 residues F60 and P61 and A-MLV residues Y60 and V61 in the first disulfide-bonded loop of VRA were important for functional inter action with the receptor's ECL4 or -5, We have now extended this approach t o identify additional VRA and VRB residues that are involved in receptor re cognition. Our studies imply that FeLV-B VRA residues F60 and P61 interact with the Pitl ECL5 region, whereas VRA residues 66 to 78 interact with Pitl ECL4. Correspondingly, A-MLV VRA residues Y60 and V61 interact with the Pi t2 ECL5 region, whereas residues 66 to 78 interact with Pit2 ECL4. Similar studies that focused on the gp70 VRB implicated residues 129 to 139 as cont ributing to specific interactions with the receptor ECL2. These results ide ntify three regions of gp70 that interact in a specific manner with distinc t portions of their receptors, thereby providing a map of the functionally interacting surfaces.