| Retroviral capsid assembly involves the accumulation of the structural Gag polyprotein in association with genomic RNA at a specific location within an infected cell. Mason-Pfizer monkey virus (M-PMV), the prototypic D-type retrovirus, assembles capsids at the pericentriolar region of the cell. Capsids are transported to the plasma membrane where M-PMV catalyzes the membrane envelopment of the spherical structure during viral release. The assembly unit of a capsid is the myristylated Gag polyprotein, which is cleaved into six proteins upon maturation of the enveloped virion (p10MA, pp 16/18, p12, p27CA, p14NC and p4). Myristylation of the Gag matrix domain (MA) is essential for transport and release but not assembly of M-PMV. A MA mutant, in which threonine residues at positions 41 and 78 were replaced with isoleucine residues, assembled capsids that are transported to the plasma membrane but are delayed in an early stage of budding. Since the nuclear magnetic resonance structure of MA showed that these threonine residues are oriented into the hydrophobic core of the protein, it was reasoned that the myristate moiety is sequestered within MA and that this mutant was defective in release of myristic acid from the more hydrophobic core. Analysis of the MPMV MA structure also identified a positive-charge density on the membrane-proximal surface of MA which is postulated to facilitate the association of MA with acidic phospholipid head groups at the plasma membrane. In this dissertation a genetic approach was used to test the hypothesis that basic amino acids in MA interact with acidic phospholipid head groups on the inner leaflet of the plasma membrane to trigger the release of myristic acid from the hydrophobic core of MA into the hydrophobic environment of the membrane, anchoring the immature capsid and facilitating viral release. It is reported here that the hydrophobicity of the MA core can modulate M-PMV release and that basic amino acids in M-PMV MA define both the cellular location and efficiency of virus release. These findings support a model in which the sequestration of myristic acid in MA is regulated by the positive charge of this domain, likely through specific electrostatic interactions. |
