Pseudotyping the Moloney murine leukemia virus with engineered envelope glycoproteins.

Abstract

We were interested in generating an in vivo retroviral gene therapy vector based on the commonly used Moloney murine leukemia virus (MoMLV). This was accomplished by pseudotyping with an engineered influenza A hemagglutinin. Point mutations were introduced to abrogate hemagglutinin's wild type binding and a single chain variable domain antibody fragment (scFv) was added to its amino terminus to provide new binding specificity. The engineered hemagglutinin was able to mediate binding of pseudotyped retrovirus to a scFv specific peptide but was unable mediate infection of target cells. To rescue the infectivity of the pseudotyped retrovirus the role of lipid rafts in the lifecycle of the MoMLV was examined. Lipid raft isolation from transfected cells and virus particles revealed that retroviral proteins were not associated with lipid rafts. Using a panel of hemagglutinin mutants with reduced lipid raft affinity we also determined that this parameter did not affect pseudotyping efficiency.