Structural basis for human T-cell leukemia virus type 1 Gag targeting to the plasma membrane for assembly

Abstract

During the late phase of retroviral replication, the virally encoded Gag polyprotein is targeted to the plasma membrane (PM) for assembly. Gag-PM binding is mediated by the N-terminal matrix (MA) domain of Gag. For many retroviruses, Gag binding to the PM was found to be dependent on phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2]. However, it was shown that for human T-cell leukemia virus type 1 (HTLV-1), Gag binding to membranes is less dependent on PI(4,5)P2, suggesting that other factors may modulate Gag assembly. To elucidate the mechanism by which HTLV-1 Gag binds to the PM, we employed NMR techniques to solve the structure of unmyristoylated MA (myr(-)MA) and to characterize its interactions with lipids and liposomes. The MA structure consists of four -helices and unstructured N- and C-termini. We show that myr(-)MA binds to PI(4,5)P2 via the polar head and that myr(-)MA binding to inositol phosphates (IPs) is significantly enhanced by increasing the number of phosphate groups on the inositol ring, indicating that the MA-IP binding is governed by charge-charge interactions. The IP binding site was mapped to a well-defined basic patch formed by lysine and arginine residues. Using a sensitive NMR-based liposome binding assay, we show that myr(-)MA binding to membranes is significantly enhanced by phosphatidylserine (PS). Confocal microscopy data show that Gag is localized to the inner leaflet of the PM of infected cells, while the Gag G2A mutant, lacking myristoylation, is diffuse and cytoplasmic. These findings advance our understanding of a key mechanism in retroviral assembly.