A SINGLE AMINO-ACID CHANGE IN THE CYTOPLASMIC DOMAIN OF THE SIMIAN IMMUNODEFICIENCY VIRUS TRANSMEMBRANE MOLECULE INCREASES ENVELOPE GLYCOPROTEIN EXPRESSION ON INFECTED-CELLS

We have described a virus termed CP-MAC, derived from the BK28 molecul ar clone of simian immunodeficiency virus, that was remarkable for its ability to infect Sup-T1 cells with rapid kinetics, cell fusion, and CD4 down-modulation (C. C. LaBranche, M. M. Sauter, B. S. Haggarty, P. J. Vance, J. Romano, T. K. Hart, P. J. Bugelski, and J. A. Hoxie, J. Virol. 68:5509-5522, 1994 [Erratum 68:7665-7667]). Compared with BK28, CP-MAC exhibited a number of changes in its envelope glycoproteins, i ncluding a highly stable association between the external (SU) and tra nsmembrane (TM) molecules, a more rapid electrophoretic mobility of TM , and, of particular interest, a marked increase in the level of envel ope protein expression on the surface of infected cells. These changes were shown to be associated with 11 coding mutations in the env gene (5 in SU and 6 in TM). In this report, we demonstrate that a single am ino acid mutation of a Tyr to a Cys at position 723 (Y723C) in the TM cytoplasmic domain of CP-MAC is the principal determinant for the incr eased expression of envelope glycoproteins on the cell surface. When i ntroduced into the env gene of BK28, the Y723C mutation produced up to a 25-fold increase in the levels of SU and TM on chronically infected cells, as determined by fluorescence-activated cell sorter analysis w ith monoclonal and polyclonal antibodies. A similar effect was observe d when a Tyr-to-Cys change was introduced at the analogous position (a mino acid 721) in the SIVmac239 molecular clone, which, unlike BK28 do es not contain a premature stop codon in its TM cytoplasmic tail. Subs tituting other amino acids, including Ala, Ile, and Ser, at this posit ion produced increases in surface envelope glycoproteins that were sim ilar to that observed for the Cys substitution, while a Tyr-to-Phe mut ation produced a smaller increase. These results could not be accounte d for by differences in the kinetics or efficiency of envelope glycopr otein processing or by shedding of SU from infected cells. However, im muno-electron microscopy demonstrated that the Y723C mutation in BK28 produced a striking redistribution of cell surface envelope molecules from localized patches to a diffuse pattern that covered the entire pl asma membrane. This finding suggests that mutation of a Tyr residue in the simian immunodeficiency virus TM cytoplasmic domain may disrupt a structural element that can modulate envelope glycoprotein expression on the surface of infected cells.