Identification of two sequences in the cytoplasmic tail of the human immunodeficiency virus type 1 envelope glycoprotein that inhibit cell surface expression

During synthesis and export of protein, the majority of the human immunodef iciency virus type 1 (HIV-1) Env glycoprotein gp160 is retained in the endo plasmic reticulum (ER) and subsequently ubiquitinated and degraded by prote asomes. Only a small fraction of gp160 appears to be correctly folded and p rocessed and is transported to the cell surface, which makes it difficult t o identify negative sequence elements regulating steady-state surface expre ssion of Env at the post-ER level. Moreover, poorly localized mRNA retentio n sequences inhibiting the nucleocytoplasmic transport of viral transcripts interfere with the identification of these sequence elements. Using two he terologous systems with CD4 or immunoglobulin extracellular/transmembrane d omains in combination with the gp160 cytoplasmic domain, we were able to id entify two membrane-distal, neighboring motifs, is1 (amino acids 750 to 763 ) and is2 (amino acids 764 to 785), which inhibited surface expression and induced Golgi localization of the chimeric proteins. To prove that these tw o elements act similarly in the homologous context of the Env glycoprotein, we generated a synthetic gp160 gene with synonymous codons, the transcript s of which are not retained within the nucleus. In accordance with the resu lts in heterologous systems, an internal deletion of both elements consider ably increased surface expression of gp160.