Suppression of a fusion defect by second site mutations in the ecotropic murine leukemia virus surface protein

Entry of ecotropic murine leukemia virus initiates when the envelope surfac e protein recognizes and binds to the virus receptor on host cells. The env elope transmembrane protein then mediates fusion of viral and host cell mem branes and penetration into the cytoplasm. Using a genetic selection, we is olated an infectious retrovirus variant containing three changes in the sur face protein-histidine 8 to arginine, glutamine 227 to arginine, and aspart ate 243 to tyrosine. Single replacement of histidine 8,with arginine (H8R) resulted in almost complete loss of infectivity, even though the mutant env elope proteins were stable and efficiently incorporated into virions. Virio ns carrying H8R envelope were proficient at binding cells expressing recept or but failed to induce cell-cell fusion of XC cells, indicating that the h istidine at position 8 plays an essential role in fusion during penetration of the host cell membrane. Thus, there is at least one domain in SU that i s involved in fusion; the fusion functions do not reside exclusively in TMI . In contrast, envelope with all three changes induced cell-cell fusion of XC cells and produced virions that were 10,000-fold more infectious than th ose containing only the H8R substitution, indicating that changes at positi ons 227 and 243 can suppress a fusion defect caused by loss of histidine 8 function. Moreover, the other two changes acted synergistically, indicating that both compensate for the loss of the same essential function of histid ine 8. The ability of these changes to suppress this fusion defect might pr ovide a means for overcoming postbinding defects found in targeted retrovir al vectors for use in human gene therapy.