EVIDENCE FOR A 2ND FUNCTION OF THE MA SEQUENCE IN THE ROUS-SARCOMA VIRUS GAG PROTEIN

During retrovirus assembly, Gag proteins bind to the inner leaflet of the plasma membrane to initiate the budding process. The molecular bas is of this protein-lipid interaction is poorly understood. For the hum an immunodeficiency virus type 1 Gag protein, we recently reported tha t the membrane-binding domain resides within the N-terminal 31 amino a cids and consists of two components: myristate and a cluster of basic residues, which together promote membrane binding in vitro and budding in vivo (W. Zhou, L. J. Parent, J. W. Wills, and M. D. Resh, J. Virol . 68:2556-2569, 1994). The positively charged residues associate elect rostatically with acidic phospholipids to stabilize membrane binding, while myristate provides membrane-binding energy via hydrophobic inter actions, Here rye demonstrate that the human immunodeficiency virus ty pe 1 Gag membrane-binding domain can fully replace the membrane-target ing function of the N-terminal 100 residues of the nonmyristylated Rou s sarcoma virus (RSV) Gag protein, To further explore the importance o f myristate and basic residues in membrane binding, we developed a gai n-of-function assay whereby budding was restored to defective mutants of RSV Gag, Detailed mutational analysis revealed that the position, n umber, and context of charged residues are crucial to budding, Myrista te provides additional membrane-binding energy, which is critical when a Gag protein is near the threshold of stable membrane association, F inally, viruses with altered matrix (RSV) proteins that are noninfecti ous, even though they produce particles with high efficiency, were ide ntified, Thus, we present the first evidence that the RSV MA sequence plays two distinct roles, membrane binding during particle assembly an d a second, as yet undefined function required for viral infectivity.